Donna M. Bouley, DVM, PhD

All Publications

• The glucosyltransferase activity of C. difficile Toxin B is required for disease pathogenesis. PLoS pathogens Bilverstone, T. W., Garland, M., Cave, R. J., Kelly, M. L., Tholen, M., Bouley, D. M., Kaye, P., Minton, N. P., Bogyo, M., Kuehne, S. A., Melnyk, R. A. 2020; 16 (9): e1008852

  Abstract

  Enzymatic inactivation of Rho-family GTPases by the glucosyltransferase domain of Clostridioides difficile Toxin B (TcdB) gives rise to various pathogenic effects in cells that are classically thought to be responsible for the disease symptoms associated with C. difficile infection (CDI). Recent in vitro studies have shown that TcdB can, under certain circumstances, induce cellular toxicities that are independent of glucosyltransferase (GT) activity, calling into question the precise role of GT activity. Here, to establish the importance of GT activity in CDI disease pathogenesis, we generated the first described mutant strain of C. difficile producing glucosyltransferase-defective (GT-defective) toxin. Using allelic exchange (AE) technology, we first deleted tcdA in C. difficile 630Deltaerm and subsequently introduced a deactivating D270N substitution in the GT domain of TcdB. To examine the role of GT activity in vivo, we tested each strain in two different animal models of CDI pathogenesis. In the non-lethal murine model of infection, the GT-defective mutant induced minimal pathology in host tissues as compared to the profound caecal inflammation seen in the wild-type and 630DeltaermDeltatcdA (DeltatcdA) strains. In the more sensitive hamster model of CDI, whereas hamsters in the wild-type or DeltatcdA groups succumbed to fulminant infection within 4 days, all hamsters infected with the GT-defective mutant survived the 10-day infection period without primary symptoms of CDI or evidence of caecal inflammation. These data demonstrate that GT activity is indispensable for disease pathogenesis and reaffirm its central role in disease and its importance as a therapeutic target for small-molecule inhibition.

  View details for DOI 10.1371/journal.ppat.1008852

  View details for PubMedID 32960931

• Histologic safety of transcranial focused ultrasound neuromodulation and magnetic resonance acoustic radiation force imaging in rhesus macaques and sheep. Brain stimulation Gaur, P., Casey, K. M., Kubanek, J., Li, N., Mohammadjavadi, M., Saenz, Y., Glover, G. H., Bouley, D. M., Pauly, K. B. 2020; 13 (3): 804–14

  Abstract

  BACKGROUND: Neuromodulation by transcranial focused ultrasound (FUS) offers the potential to non-invasively treat specific brain regions, with treatment location verified by magnetic resonance acoustic radiation force imaging (MR-ARFI).OBJECTIVE: To investigate the safety of these methods prior to widespread clinical use, we report histologic findings in two large animal models following FUS neuromodulation and MR-ARFI.METHODS: Two rhesus macaques and thirteen Dorset sheep were studied. FUS neuromodulation was targeted to the primary visual cortex in rhesus macaques and to subcortical locations, verified by MR-ARFI, in eleven sheep. Both rhesus macaques and five sheep received a single FUS session, whereas six sheep received repeated sessions three to six days apart. The remaining two control sheep did not receive ultrasound but otherwise underwent the same anesthetic and MRI procedures as the eleven experimental sheep. Hematoxylin and eosin-stained sections of brain tissue (harvested zero to eleven days following FUS) were evaluated for tissue damage at FUS and control locations as well as tissue within the path of the FUS beam. TUNEL staining was used to evaluate for the presence of apoptosis in sheep receiving high dose FUS.RESULTS: No FUS-related pre-mortem histologic findings were observed in the rhesus macaques or in any of the examined sheep. Extravascular red blood cells (RBCs) were present within the meninges of all sheep, regardless of treatment group. Similarly, small aggregates of perivascular RBCs were rarely noted in non-target regions of neural parenchyma of FUS-treated (8/11) and untreated (2/2) sheep. However, no concurrent histologic abnormalities were observed, consistent with RBC extravasation occurring as post-mortem artifact following brain extraction. Sheep within the high dose FUS group were TUNEL-negative at the targeted site of FUS.CONCLUSIONS: The absence of FUS-related histologic findings suggests that the neuromodulation and MR-ARFI protocols evaluated do not cause tissue damage.

  View details for DOI 10.1016/j.brs.2020.02.017

  View details for PubMedID 32289711

• Histologic evaluation of activation of acute inflammatory response in a mouse model following ultrasound-mediated blood-brain barrier using different acoustic pressures and microbubble doses. Nanotheranostics Pascal, A. n., Li, N. n., Lechtenberg, K. J., Rosenberg, J. n., Airan, R. D., James, M. L., Bouley, D. M., Pauly, K. B. 2020; 4 (4): 210–23

  Abstract

  Rationale: Localized blood-brain barrier (BBB) opening can be achieved with minimal to no tissue damage by applying pulsed focused ultrasound alongside a low microbubble (MB) dose. However, relatively little is known regarding how varying treatment parameters affect the degree of neuroinflammation following BBB opening. The goal of this study was to evaluate the activation of an inflammatory response following BBB opening as a function of applied acoustic pressure using two different microbubble doses. Methods: Mice were treated with 650 kHz ultrasound using varying acoustic peak negative pressures (PNPs) using two different MB doses, and activation of an inflammatory response, in terms of microglial and astrocyte activation, was assessed one hour following BBB opening using immunohistochemical staining. Harmonic and subharmonic acoustic emissions (AEs) were monitored for all treatments with a passive cavitation detector, and contrast-enhanced magnetic resonance imaging (CE-MRI) was performed following BBB opening to quantify the degree of opening. Hematoxylin and eosin-stained slides were assessed for the presence of microhemorrhage and edema. Results: For each MB dose, BBB opening was achieved with minimal activation of microglia and astrocytes using a PNP of 0.15 MPa. Higher PNPs were associated with increased activation, with greater increases associated with the use of the higher MB dose. Additionally, glial activation was still observed in the absence of histopathological findings. We found that CE-MRI was most strongly correlated with the degree of activation. While acoustic emissions were not predictive of microglial or astrocyte activation, subharmonic AEs were strongly associated with marked and severe histopathological findings. Conclusions: Our study demonstrated that there were mild histologic changes and activation of the acute inflammatory response using PNPs ranging from 0.15 MPa to 0.20 MPa, independent of MB dose. However, when higher PNPs of 0.25 MPa or above were applied, the same applied PNP resulted in more severe and widespread histological findings and activation of the acute inflammatory response when using the higher MB dose. The potential activation of the inflammatory response following ultrasound-mediated BBB opening should be considered when treating patients to maximize therapeutic benefit.

  View details for DOI 10.7150/ntno.49898

  View details for PubMedID 32802731

  View details for PubMedCentralID PMC7425053

• Axial Skeletal Malformations in Genetically Modified Xenopus laevis and Xenopus tropicalis. Comparative medicine Zlatow, A. L., Wilson, S. S., Bouley, D. M., Tetens-Woodring, J. n., Buchholz, D. R., Green, S. L. 2020; 70 (6): 532–41

  Abstract

  Skeletal malformations in captive-bred, adult Xenopus spp., have not previously been reported. Here we describe 10 sexually mature, genetically modified laboratory frogs (6 Xenopus laevis and 4 Xenopus tropicalis) with axial skeletal abnormalities. The young adult frogs were described by veterinary staff as presenting with "hunchbacks," but were otherwise considered to be in good health. All affected frogs were genetically engineered using various techniques: transcription activator-like effector nucleases (TALEN) editing using thyroid hormone receptor α TALEN mRNA, restriction enzyme-mediated integration methods involving insertion of the inducible transgene pCAR/TRDN, or via I-SceI meganuclease transgenesis using either pDRTREdpTR-HS4 or pDPCrtTA-TREG-HS4 plasmid sequences. Radiographic findings (6 frogs) and gross necropsy (10 frogs) revealed vertebral column malformations and sacroiliac deformities that resulted in moderate to severe kyphosis and kyphoscoliosis. These findings were confirmed and additional skeletal abnormalities were identified using computed tomography to create a 3D reconstruction of 4 frogs. Additional findings visible on the 3D reconstructions included incomplete vertebral segmentation, malformed transverse processes, and a short and/or curved urostyle. Histopathologic findings included misshapen intervertebral joints with nonconforming articular surfaces, narrowed joint cavities, flattened or irregularly-formed articular cartilage, irregular maturation lines and nonpolarized chondrocytes, excess fibrocartilage, and evidence of irregular bone resorption and growth. While the specific etiology of the vertebral skeletal abnormalities remains unclear, possibilities include: 1) egg/oocyte physical manipulation (dejellying, microinjection, fertilization, etc.), 2) induction and expression of the transgenes, 3) inactivation (knockout) of existing genes by insertional mutagenesis, or 4) a combination of the above. Furthermore, the possibility of undetected changes in the macro or microenvironment, or a feature of the genetic background of the affected frogs cannot be ruled out.

  View details for DOI 10.30802/AALAS-CM-20-000069

  View details for PubMedID 33203505

• A Gut Commensal-Produced Metabolite Mediates Colonization Resistance to Salmonella Infection. Cell host & microbe Jacobson, A., Lam, L., Rajendram, M., Tamburini, F., Honeycutt, J., Pham, T., Van Treuren, W., Pruss, K., Stabler, S. R., Lugo, K., Bouley, D. M., Vilches-Moure, J. G., Smith, M., Sonnenburg, J. L., Bhatt, A. S., Huang, K. C., Monack, D. 2018

  Abstract

  The intestinal microbiota provides colonization resistance against pathogens, limiting pathogen expansion and transmission. These microbiota-mediated mechanisms were previously identified by observing loss of colonization resistance after antibiotic treatment or dietary changes, which severely disrupt microbiota communities. We identify a microbiota-mediated mechanism of colonization resistance against Salmonella enterica serovar Typhimurium (S. Typhimurium) by comparing high-complexity commensal communities with different levels of colonization resistance. Using inbred mouse strains with different infection dynamics and S. Typhimurium intestinal burdens, we demonstrate that Bacteroides species mediate colonization resistance against S. Typhimurium by producing the short-chain fatty acid propionate. Propionate directly inhibits pathogen growth invitro by disrupting intracellular pH homeostasis, and chemically increasing intestinal propionate levels protects mice from S.Typhimurium. In addition, administering susceptible mice Bacteroides, but not a propionate-production mutant, confers resistance to S. Typhimurium. This work provides mechanistic understanding into the role of individualized microbial communities in host-to-host variability of pathogen transmission.

  View details for PubMedID 30057174

• Transient Osmotic Perturbation Causes Long-Term Alteration to the Gut Microbiota. Cell Tropini, C., Moss, E. L., Merrill, B. D., Ng, K. M., Higginbottom, S. K., Casavant, E. P., Gonzalez, C. G., Fremin, B., Bouley, D. M., Elias, J. E., Bhatt, A. S., Huang, K. C., Sonnenburg, J. L. 2018; 173 (7): 1742

  Abstract

  Osmotic diarrhea is a prevalent condition in humans caused by food intolerance, malabsorption, and widespread laxative use. Here, we assess the resilience of the gut ecosystem to osmotic perturbation at multiple length and timescales using mice as model hosts. Osmotic stress caused reproducible extinction of highly abundant taxa and expansion of less prevalent members in human and mouse microbiotas. Quantitative imaging revealed decimation of the mucus barrier during osmotic perturbation, followed by recovery. The immune system exhibited temporary changes in cytokine levels and a lasting IgG response against commensal bacteria. Increased osmolality prevented growth of commensal strains invitro, revealing one mechanism contributing to extinction. Environmental availability of microbiota members mitigated extinction events, demonstrating how species reintroduction can affect community resilience. Our findings (1) demonstrate that even mild osmotic diarrhea can cause lasting changes to the microbiota and host and (2) lay the foundation for interventions that increase system-wide resilience.

  View details for PubMedID 29906449

• Transient Osmotic Perturbation Causes Long-Term Alteration to the Gut Microbiota CELL Tropini, C., Moss, E., Merrill, B., Ng, K., Higginbottom, S., Casavant, E., Gonzalez, C., Fremin, B., Bouley, D., Elias, J., Bhatt, A., Huang, K., Sonnenburg, J. 2018; 173 (7): 1742-+

  View details for DOI 10.1016/j.cell.2018.05.008

  View details for Web of Science ID 000437004000021

• Microbiota-accessible carbohydrates suppress Clostridium difficile infection in a murine model. Nature microbiology Hryckowian, A. J., Van Treuren, W. n., Smits, S. A., Davis, N. M., Gardner, J. O., Bouley, D. M., Sonnenburg, J. L. 2018

  Abstract

  Clostridium difficile is an opportunistic diarrhoeal pathogen, and C. difficile infection (CDI) represents a major health care concern, causing an estimated 15,000 deaths per year in the United States alone 1 . Several enteric pathogens, including C. difficile, leverage inflammation and the accompanying microbial dysbiosis to thrive in the distal gut 2 . Although diet is among the most powerful available tools for affecting the health of humans and their relationship with their microbiota, investigation into the effects of diet on CDI has been limited. Here, we show in mice that the consumption of microbiota-accessible carbohydrates (MACs) found in dietary plant polysaccharides has a significant effect on CDI. Specifically, using a model of antibiotic-induced CDI that typically resolves within 12 days of infection, we demonstrate that MAC-deficient diets perpetuate CDI. We show that C. difficile burdens are suppressed through the addition of either a diet containing a complex mixture of MACs or a simplified diet containing inulin as the sole MAC source. We show that switches between these dietary conditions are coincident with changes to microbiota membership, its metabolic output and C. difficile-mediated inflammation. Together, our data demonstrate the outgrowth of MAC-utilizing taxa and the associated end products of MAC metabolism, namely, the short-chain fatty acids acetate, propionate and butyrate, are associated with decreased C. difficile fitness despite increased C. difficile toxin expression in the gut. Our findings, when placed into the context of the known fibre deficiencies of a human Western diet, provide rationale for pursuing MAC-centric dietary strategies as an alternate line of investigation for mitigating CDI.

  View details for PubMedID 29686297

• Toxoplasma gondii infection triggers chronic cachexia and sustained commensal dysbiosis in mice. PloS one Hatter, J. A., Kouche, Y. M., Melchor, S. J., Ng, K., Bouley, D. M., Boothroyd, J. C., Ewald, S. E. 2018; 13 (10): e0204895

  Abstract

  Toxoplasma gondii is a protozoan parasite with a predation-mediated transmission cycle between rodents and felines. Intermediate hosts acquire Toxoplasma by eating parasite cysts which invade the small intestine, disseminate systemically and finally establish host life-long chronic infection in brain and muscles. Here we show that Toxoplasma infection can trigger a severe form of sustained cachexia: a disease of progressive lean weight loss that is a causal predictor of mortality in cancer, chronic disease and many infections. Toxoplasma cachexia is characterized by acute anorexia, systemic inflammation and loss of 20% body mass. Although mice recover from symptoms of peak sickness, they fail to regain muscle mass or visceral adipose depots. We asked whether the damage to the intestinal microenvironment observed at acute time points was sustained in chronic infection and could thereby play a role in sustaining cachexia. We found that parasites replicate in the same region of the distal jejunum/proximal ileum throughout acute infection, inducing the development of secondary lymphoid structures and severe, regional inflammation. Small intestine pathology was resolved by 5 weeks post-infection. However, changes in the commensal populations, notably an outgrowth of Clostridia spp., were sustained in chronic infection. Importantly, uninfected animals co-housed with infected mice display similar changes in commensal microflora but never display symptoms of cachexia, indicating that altered commensals are not sufficient to explain the cachexia phenotype alone. These studies indicate that Toxoplasma infection is a novel and robust model to study the immune-metabolic interactions that contribute to chronic cachexia development, pathology and potential reversal.

  View details for PubMedID 30379866

• Pseudogenization of the Secreted Effector Gene sseI Confers Rapid Systemic Dissemination of S. Typhimurium ST313 within Migratory Dendritic Cells. Cell host & microbe Carden, S. E., Walker, G. T., Honeycutt, J., Lugo, K., Pham, T., Jacobson, A., Bouley, D., Idoyaga, J., Tsolis, R. M., Monack, D. 2017; 21 (2): 182-194

  Abstract

  Genome degradation correlates with host adaptation and systemic disease in Salmonella. Most lineages of the S. enterica subspecies Typhimurium cause gastroenteritis in humans; however, the recently emerged ST313 lineage II pathovar commonly causes systemic bacteremia in sub-Saharan Africa. ST313 lineage II displays genome degradation compared to gastroenteritis-associated lineages; yet, the mechanisms and causal genetic differences mediating these infection phenotypes are largely unknown. We find that the ST313 isolate D23580 hyperdisseminates from the gut to systemic sites, such as the mesenteric lymph nodes (MLNs), via CD11b(+) migratory dendritic cells (DCs). This hyperdissemination was facilitated by the loss of sseI, which encodes an effector that inhibits DC migration in gastroenteritis-associated isolates. Expressing functional SseI in D23580 reduced the number of infected migratory DCs and bacteria in the MLN. Our study reveals a mechanism linking pseudogenization of effectors with the evolution of niche adaptation in a bacterial pathogen.

  View details for DOI 10.1016/j.chom.2017.01.009

  View details for PubMedID 28182950

  View details for PubMedCentralID PMC5325708

• Acute and Chronic Outcomes of Gas-Bubble Disease in a Colony of African Clawed Frogs (Xenopus laevis) COMPARATIVE MEDICINE Tsai, J. Y., Felt, S. A., Bouley, D. M., Green, S. L. 2017; 67 (1): 4-10

  Abstract

  Gas-bubble disease occurs in aquatic species that are exposed to water that is supersaturated with gases. In February 2007, municipal water supersaturated with gas was inadvertently pumped into the vivarium's aquatic housing systems and affected approximately 450 adult female Xenopus laevis. The inflow of supersaturated water was stopped immediately, the holding tanks aggressively aerated, and all experimental manipulations and feeding ceased. Within the first 6 h after the event, morbidity approached 90%, and mortality reached 3.5%. Acutely affected frogs showed clinical signs of gas-bubble disease: buoyancy problems, micro- and macroscopic bubbles in the foot webbing, hyperemia in foot webbing and leg skin, and loss of the mucous slime coat. All of the frogs that died or were euthanized had areas of mesenteric infarction, which resulted in intestinal epithelial necrosis and degeneration of the muscular tunic. Over the subsequent 2 wk, as gas saturation levels returned to normal, the clinical symptoms resolved completely in the remaining frogs. However, 3 mo later, 85% of them failed to lay eggs or produce oocytes, and the remaining 15% produced oocytes of low number and poor quality, yielding cytosolic extracts with poor to no enzymatic activity. Histology of the egg mass from a single 2- to 3-y-old frog at 3 mo after disease resolution revealed irregularly shaped oocytes, few large mature oocytes, and numerous small, degenerating oocytes. At 6 mo after the incident, the remaining frogs continued to fail to produce eggs of sufficient quantity or quality after hormonal priming. The researchers consequently opted to cull the remainder of the colony and repopulate with new frogs.

  View details for Web of Science ID 000394079600001

  View details for PubMedCentralID PMC5310619

• Endoluminal ultrasound applicators for MR-guided thermal ablation of pancreatic tumors: Preliminary design and evaluation in a porcine pancreas model MEDICAL PHYSICS Adams, M. S., Salgaonkar, V. A., Plata-Camargo, J., Jones, P. D., Pascal-Tenorio, A., Chen, H., Bouley, D. M., Sommer, G., Pauly, K. B., Diederich, C. J. 2016; 43 (7): 4184-4197

  Abstract

  Endoluminal ultrasound may serve as a minimally invasive option for delivering thermal ablation to pancreatic tumors adjacent to the stomach or duodenum. The objective of this study was to explore the basic feasibility of this treatment strategy through the design, characterization, and evaluation of proof-of-concept endoluminal ultrasound applicators capable of placement in the gastrointestinal (GI) lumen for volumetric pancreas ablation under MR guidance.Two variants of the endoluminal applicator, each containing a distinct array of two independently powered transducers (10 × 10 mm 3.2 MHz planar; or 8 × 10 × 20 mm radius of curvature 3.3 MHz curvilinear geometries) at the distal end of a meter long flexible catheter assembly, were designed and fabricated. Transducers and circulatory water flow for acoustic coupling and luminal cooling were contained by a low-profile polyester balloon covering the transducer assembly fixture. Each applicator incorporated miniature spiral MR coils and mechanical features (guiding tips and hinges) to facilitate tracking and insertion through the GI tract under MRI guidance. Acoustic characterization of each device was performed using radiation force balance and hydrophone measurements. Device delivery into the upper GI tract, adjacent to the pancreas, and heating characteristics for treatment of pancreatic tissue were evaluated in MR-guided ex vivo and in vivo porcine experiments. MR guidance was utilized for anatomical target identification, tracking/positioning of the applicator, and MR temperature imaging (MRTI) for PRF-based multislice thermometry, implemented in the real-time RTHawk software environment.Force balance and hydrophone measurements indicated efficiencies of 48.8% and 47.8% and -3 dB intensity beam-widths of 3.2 and 1.2 mm for the planar and curvilinear transducers, respectively. Ex vivo studies on whole-porcine carcasses revealed capabilities of producing ablative temperature rise (ΔT > 15 °C) contours in pancreatic tissue 4-40 mm long and 4-28 mm wide for the planar transducer applicator (1-13 min sonication duration, ∼4 W/cm(2) applied acoustic intensity). Curvilinear transducers produced more selective heating, with a narrower ΔT > 15 °C contour length and width of up to 1-24 mm and 2-7 mm, respectively (1-7 min sonication duration, ∼4 W/cm(2) applied acoustic intensity). Active tracking of the miniature spiral coils was achieved using a Hadamard encoding tracking sequence, enabling real-time determination of each coil's coordinates and automated prescription of imaging planes for thermometry. In vivo MRTI-guided heating trials in three pigs demonstrated capability of ∼20 °C temperature elevation in pancreatic tissue at 2 cm depths from the applicator, with 5-7 W/cm(2) applied intensity and 6-16 min sonication duration. Dimensions of thermal lesions in the pancreas ranged from 12 to 28 mm, 3 to 10 mm, and 5 to 10 mm in length, width, and depth, respectively, as verified through histological analysis of tissue sections. Multiple-baseline reconstruction and respiratory-gated acquisition were demonstrated to be effective strategies in suppressing motion artifacts for clear evolution of temperature profiles during MRTI in the in vivo studies.This study demonstrates the technical feasibility of generating volumetric ablation in pancreatic tissue using endoluminal ultrasound applicators positioned in the stomach lumen. MR guidance facilitates target identification, device tracking/positioning, and treatment monitoring through real-time multislice PRF-based thermometry.

  View details for DOI 10.1118/1.4953632

  View details for Web of Science ID 000379171900027

  View details for PubMedID 27370138

  View details for PubMedCentralID PMC4912561

• The adherens junctions control susceptibility to Staphylococcus aureus a-toxin. Proceedings of the National Academy of Sciences of the United States of America Popov, L. M., Marceau, C. D., Starkl, P. M., Lumb, J. H., Shah, J., Guerrera, D., Cooper, R. L., Merakou, C., Bouley, D. M., Meng, W., Kiyonari, H., Takeichi, M., Galli, S. J., Bagnoli, F., Citi, S., Carette, J. E., Amieva, M. R. 2015; 112 (46): 14337-14342

  Abstract

  Staphylococcus aureus is both a transient skin colonizer and a formidable human pathogen, ranking among the leading causes of skin and soft tissue infections as well as severe pneumonia. The secreted bacterial α-toxin is essential for S. aureus virulence in these epithelial diseases. To discover host cellular factors required for α-toxin cytotoxicity, we conducted a genetic screen using mutagenized haploid human cells. Our screen identified a cytoplasmic member of the adherens junctions, plekstrin-homology domain containing protein 7 (PLEKHA7), as the second most significantly enriched gene after the known α-toxin receptor, a disintegrin and metalloprotease 10 (ADAM10). Here we report a new, unexpected role for PLEKHA7 and several components of cellular adherens junctions in controlling susceptibility to S. aureus α-toxin. We find that despite being injured by α-toxin pore formation, PLEKHA7 knockout cells recover after intoxication. By infecting PLEKHA7(-/-) mice with methicillin-resistant S. aureus USA300 LAC strain, we demonstrate that this junctional protein controls disease severity in both skin infection and lethal S. aureus pneumonia. Our results suggest that adherens junctions actively control cellular responses to a potent pore-forming bacterial toxin and identify PLEKHA7 as a potential nonessential host target to reduce S. aureus virulence during epithelial infections.

  View details for DOI 10.1073/pnas.1510265112

  View details for PubMedID 26489655

• A feasibility study on monitoring the evolution of apparent diffusion coefficient decrease during thermal ablation MEDICAL PHYSICS Plata, J. C., Holbrook, A. B., Marx, M., Salgaonkar, V., Jones, P., Pascal-Tenorio, A., Bouley, D., Diederich, C., Sommer, G., Pauly, K. B. 2015; 42 (9): 5130-5137

  Abstract

  Evaluate whether a decrease in apparent diffusion coefficient (ADC), associated with loss of tissue viability (LOTV), can be observed during the course of thermal ablation of the prostate.Thermal ablation was performed in a healthy in vivo canine prostate model (N = 2, ages: 5 yr healthy, mixed breed, weights: 13-14 kg) using a transurethral high-intensity ultrasound catheter and was monitored using a strategy that interleaves diffusion weighted images and gradient-echo images. The two sequences were used to measure ADC and changes in temperature during the treatment. Changes in temperature were used to compute expected changes in ADC. The difference between expected and measured ADC, ADCDIFF, was analyzed in regions ranging from moderate hyperthermia to heat fixation. A receiver operator characteristic (ROC) curve analysis was used to select a threshold of detection of LOTV. Time of threshold activation, tLOTV, was compared with time to reach CEM43 = 240, tDOSE.The observed relationship between temperature and ADC in vivo (2.2%/ °C, 1.94%-2.47%/ °C 95% confidence interval) was not significantly different than the previously reported value of 2.4%/ °C in phantom. ADCDIFF changes after correction for temperature showed a mean decrease of 25% in ADC 60 min post-treatment in regions where sufficient thermal dose (CEM43 > 240) was achieved. Following our ROC analysis, a threshold of 2.25% decrease in ADCDIFF for three consecutive time points was chosen as an indicator of LOTV. The ADCDIFF was found to decrease quickly (1-2 min) after reaching CEM43 = 240 in regions associated with heat fixation and more slowly (10-20 min) in regions that received slower heating.Simultaneous monitoring of ADC and temperature during treatment might allow for a more complete tissue viability assessment of ablative thermal treatments in the prostate. ADCDIFF decreases during the course of treatment may be interpreted as loss of tissue viability.

  View details for DOI 10.1118/1.4928155

  View details for Web of Science ID 000360645000015

  View details for PubMedCentralID PMC4537482

• A feasibility study on monitoring the evolution of apparent diffusion coefficient decrease during thermal ablation. Medical physics Plata, J. C., Holbrook, A. B., Marx, M., Salgaonkar, V., Jones, P., Pascal-Tenorio, A., Bouley, D., Diederich, C., Sommer, G., Pauly, K. B. 2015; 42 (9): 5130-5137

  Abstract

  Evaluate whether a decrease in apparent diffusion coefficient (ADC), associated with loss of tissue viability (LOTV), can be observed during the course of thermal ablation of the prostate.Thermal ablation was performed in a healthy in vivo canine prostate model (N = 2, ages: 5 yr healthy, mixed breed, weights: 13-14 kg) using a transurethral high-intensity ultrasound catheter and was monitored using a strategy that interleaves diffusion weighted images and gradient-echo images. The two sequences were used to measure ADC and changes in temperature during the treatment. Changes in temperature were used to compute expected changes in ADC. The difference between expected and measured ADC, ADCDIFF, was analyzed in regions ranging from moderate hyperthermia to heat fixation. A receiver operator characteristic (ROC) curve analysis was used to select a threshold of detection of LOTV. Time of threshold activation, tLOTV, was compared with time to reach CEM43 = 240, tDOSE.The observed relationship between temperature and ADC in vivo (2.2%/ °C, 1.94%-2.47%/ °C 95% confidence interval) was not significantly different than the previously reported value of 2.4%/ °C in phantom. ADCDIFF changes after correction for temperature showed a mean decrease of 25% in ADC 60 min post-treatment in regions where sufficient thermal dose (CEM43 > 240) was achieved. Following our ROC analysis, a threshold of 2.25% decrease in ADCDIFF for three consecutive time points was chosen as an indicator of LOTV. The ADCDIFF was found to decrease quickly (1-2 min) after reaching CEM43 = 240 in regions associated with heat fixation and more slowly (10-20 min) in regions that received slower heating.Simultaneous monitoring of ADC and temperature during treatment might allow for a more complete tissue viability assessment of ablative thermal treatments in the prostate. ADCDIFF decreases during the course of treatment may be interpreted as loss of tissue viability.

  View details for DOI 10.1118/1.4928155

  View details for PubMedID 26328964

• Helicobacter pylori Activates and Expands Lgr5(+) Stem Cells Through Direct Colonization of the Gastric Glands. Gastroenterology Sigal, M., Rothenberg, M. E., Logan, C. Y., Lee, J. Y., Honaker, R. W., Cooper, R. L., Passarelli, B., Camorlinga, M., Bouley, D. M., Alvarez, G., Nusse, R., Torres, J., Amieva, M. R. 2015; 148 (7): 1392-404 e21

  Abstract

  Helicobacter pylori infection is the main risk factor for gastric cancer. We characterized the interactions of H pylori with gastric epithelial progenitor and stem cells in humans and mice and investigated how these interactions contribute to H pylori-induced pathology.We used quantitative confocal microscopy and 3-dimensional reconstruction of entire gastric glands to determine the localizations of H pylori in stomach tissues from humans and infected mice. Using lineage tracing to mark cells derived from leucine-rich repeat-containing G-protein coupled receptor 5-positive (Lgr5(+)) stem cells (Lgr5-eGFP-IRES-CreERT2/Rosa26-TdTomato mice) and in situ hybridization, we analyzed gastric stem cell responses to infection. Isogenic H pylori mutants were used to determine the role of specific virulence factors in stem cell activation and pathology.H pylori grow as distinct bacterial microcolonies deep in the stomach glands and interact directly with gastric progenitor and stem cells in tissues from mice and humans. These gland-associated bacteria activate stem cells, increasing the number of stem cells, accelerating Lgr5(+) stem cell proliferation, and up-regulating expression of stem cell-related genes. Mutant bacteria with defects in chemotaxis that are able to colonize the stomach surface but not the antral glands in mice do not activate stem cells. In addition, bacteria that are unable to inject the contact-dependent virulence factor CagA into the epithelium colonized stomach glands in mice, but did not activate stem cells or produce hyperplasia to the same extent as wild-type H pylori.H pylori colonize and manipulate the progenitor and stem cell compartments, which alters turnover kinetics and glandular hyperplasia. Bacterial ability to alter the stem cells has important implications for gastrointestinal stem cell biology and H pylori-induced gastric pathology.

  View details for DOI 10.1053/j.gastro.2015.02.049

  View details for PubMedID 25725293

• Helicobacter pylori Activates and Expands Lgr5(+) Stem Cells Through Direct Colonization of the Gastric Glands GASTROENTEROLOGY Sigal, M., Rothenberg, M. E., Logan, C. Y., Lee, J. Y., Honaker, R. W., Cooper, R. L., Passarelli, B., Camorlinga, M., Bouley, D. M., Alvarez, G., Nusse, R., Torres, J., Amieva, M. R. 2015; 148 (7): 1392-?

  Abstract

  Helicobacter pylori infection is the main risk factor for gastric cancer. We characterized the interactions of H pylori with gastric epithelial progenitor and stem cells in humans and mice and investigated how these interactions contribute to H pylori-induced pathology.We used quantitative confocal microscopy and 3-dimensional reconstruction of entire gastric glands to determine the localizations of H pylori in stomach tissues from humans and infected mice. Using lineage tracing to mark cells derived from leucine-rich repeat-containing G-protein coupled receptor 5-positive (Lgr5(+)) stem cells (Lgr5-eGFP-IRES-CreERT2/Rosa26-TdTomato mice) and in situ hybridization, we analyzed gastric stem cell responses to infection. Isogenic H pylori mutants were used to determine the role of specific virulence factors in stem cell activation and pathology.H pylori grow as distinct bacterial microcolonies deep in the stomach glands and interact directly with gastric progenitor and stem cells in tissues from mice and humans. These gland-associated bacteria activate stem cells, increasing the number of stem cells, accelerating Lgr5(+) stem cell proliferation, and up-regulating expression of stem cell-related genes. Mutant bacteria with defects in chemotaxis that are able to colonize the stomach surface but not the antral glands in mice do not activate stem cells. In addition, bacteria that are unable to inject the contact-dependent virulence factor CagA into the epithelium colonized stomach glands in mice, but did not activate stem cells or produce hyperplasia to the same extent as wild-type H pylori.H pylori colonize and manipulate the progenitor and stem cell compartments, which alters turnover kinetics and glandular hyperplasia. Bacterial ability to alter the stem cells has important implications for gastrointestinal stem cell biology and H pylori-induced gastric pathology.

  View details for DOI 10.1053/j.gastro.2015.02.049

  View details for Web of Science ID 000355014700031

  View details for PubMedID 25725293

• Biochemical and Hematologic Reference Intervals for Aged Xenopus laevis in a Research Colony. Journal of the American Association for Laboratory Animal Science Chang, A. G., Hu, J., Lake, E., Bouley, D. M., Johns, J. L. 2015; 54 (5): 465-470

  Abstract

  Xenopus laevis, the African clawed frog, is commonly used in developmental and toxicology research studies. Little information is available on aged X. laevis; however, with the complete mapping of the genome and the availability of transgenic animal models, the number of aged animals in research colonies is increasing. The goals of this study were to obtain biochemical and hematologic parameters to establish reference intervals for aged X. laevis and to compare results with those from young adult X. laevis. Blood samples were collected from laboratory reared, female frogs (n = 52) between the ages of 10 and 14 y. Reference intervals were generated for 30 biochemistry analytes and full hematologic analysis; these data were compared with prior results for young X. laevis from the same vendor. Parameters that were significantly higher in aged compared with young frogs included calcium, calcium:phosphorus ratio, total protein, albumin, HDL, amylase, potassium, CO2, and uric acid. Parameters found to be significantly lower in aged frogs included glucose, AST, ALT, cholesterol, BUN, BUN:creatinine ratio, phosphorus, triglycerides, LDL, lipase, sodium, chloride, sodium:potassium ratio, and anion gap. Hematology data did not differ between young and old frogs. These findings indicate that chemistry reference intervals for young X. laevis may be inappropriate for use with aged frogs.

  View details for PubMedID 26424243

• Gut Microbiota-Produced Succinate Promotes C-difficile Infection after Antibiotic Treatment or Motility Disturbance CELL HOST & MICROBE Ferreyra, J. A., Wu, K. J., Hryckowian, A. J., Bouley, D. M., Weimer, B. C., Sonnenburg, J. L. 2014; 16 (6): 770-777

  Abstract

  Clostridium difficile is a leading cause of antibiotic-associated diarrhea. The mechanisms underlying C. difficile expansion after microbiota disturbance are just emerging. We assessed the gene expression profile of C. difficile within the intestine of gnotobiotic mice to identify genes regulated in response to either dietary or microbiota compositional changes. In the presence of the gut symbiont Bacteroides thetaiotaomicron, C. difficile induces a pathway that metabolizes the microbiota fermentation end-product succinate to butyrate. The low concentration of succinate present in the microbiota of conventional mice is transiently elevated upon antibiotic treatment or chemically induced intestinal motility disturbance, and C. difficile exploits this succinate spike to expand in the perturbed intestine. A C. difficile mutant compromised in succinate utilization is at a competitive disadvantage during these perturbations. Understanding the metabolic mechanisms involved in microbiota-C. difficile interactions may help to identify approaches for the treatment and prevention of C. difficile-associated diseases.

  View details for DOI 10.1016/j.chom.2014.11.003

  View details for Web of Science ID 000346211100010

  View details for PubMedID 25498344

• m(6)A RNA Modification Controls Cell Fate Transition in Mammalian Embryonic Stem Cells. Cell stem cell Batista, P. J., Molinie, B., Wang, J., Qu, K., Zhang, J., Li, L., Bouley, D. M., Lujan, E., Haddad, B., Daneshvar, K., Carter, A. C., Flynn, R. A., Zhou, C., Lim, K., Dedon, P., Wernig, M., Mullen, A. C., Xing, Y., Giallourakis, C. C., Chang, H. Y. 2014; 15 (6): 707-719

  Abstract

  N6-methyl-adenosine (m(6)A) is the most abundant modification on messenger RNAs and is linked to human diseases, but its functions in mammalian development are poorly understood. Here we reveal the evolutionary conservation and function of m(6)A by mapping the m(6)A methylome in mouse and human embryonic stem cells. Thousands of messenger and long noncoding RNAs show conserved m(6)A modification, including transcripts encoding core pluripotency transcription factors. m(6)A is enriched over 3' untranslated regions at defined sequence motifs and marks unstable transcripts, including transcripts turned over upon differentiation. Genetic inactivation or depletion of mouse and human Mettl3, one of the m(6)A methylases, led to m(6)A erasure on select target genes, prolonged Nanog expression upon differentiation, and impaired ESC exit from self-renewal toward differentiation into several lineages in vitro and in vivo. Thus, m(6)A is a mark of transcriptome flexibility required for stem cells to differentiate to specific lineages.

  View details for DOI 10.1016/j.stem.2014.09.019

  View details for PubMedID 25456834

• Role of disease-associated tolerance in infectious superspreaders PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Gopinath, S., Lichtman, J. S., Bouley, D. M., Elias, J. E., Monack, D. M. 2014; 111 (44): 15780-15785

  Abstract

  Natural populations show striking heterogeneity in their ability to transmit disease. For example, a minority of infected individuals known as superspreaders carries out the majority of pathogen transmission events. In a mouse model of Salmonella infection, a subset of infected hosts becomes superspreaders, shedding high levels of bacteria (>10(8) cfu per g of feces) but remain asymptomatic with a dampened systemic immune state. Here we show that superspreader hosts remain asymptomatic when they are treated with oral antibiotics. In contrast, nonsuperspreader Salmonella-infected hosts that are treated with oral antibiotics rapidly shed superspreader levels of the pathogen but display signs of morbidity. This morbidity is linked to an increase in inflammatory myeloid cells in the spleen followed by increased production of acute-phase proteins and proinflammatory cytokines. The degree of colonic inflammation is similar in antibiotic-treated superspreader and nonsuperspreader hosts, indicating that the superspreader hosts are tolerant of antibiotic-mediated perturbations in the intestinal tract. Importantly, neutralization of acute-phase proinflammatory cytokines in antibiotic-induced superspreaders suppresses the expansion of inflammatory myeloid cells and reduces morbidity. We describe a unique disease-associated tolerance to oral antibiotics in superspreaders that facilitates continued transmission of the pathogen.

  View details for DOI 10.1073/pnas.1409968111

  View details for Web of Science ID 000344088100047

  View details for PubMedCentralID PMC4226084

• Role of disease-associated tolerance in infectious superspreaders. Proceedings of the National Academy of Sciences of the United States of America Gopinath, S., Lichtman, J. S., Bouley, D. M., Elias, J. E., Monack, D. M. 2014; 111 (44): 15780-15785

  Abstract

  Natural populations show striking heterogeneity in their ability to transmit disease. For example, a minority of infected individuals known as superspreaders carries out the majority of pathogen transmission events. In a mouse model of Salmonella infection, a subset of infected hosts becomes superspreaders, shedding high levels of bacteria (>10(8) cfu per g of feces) but remain asymptomatic with a dampened systemic immune state. Here we show that superspreader hosts remain asymptomatic when they are treated with oral antibiotics. In contrast, nonsuperspreader Salmonella-infected hosts that are treated with oral antibiotics rapidly shed superspreader levels of the pathogen but display signs of morbidity. This morbidity is linked to an increase in inflammatory myeloid cells in the spleen followed by increased production of acute-phase proteins and proinflammatory cytokines. The degree of colonic inflammation is similar in antibiotic-treated superspreader and nonsuperspreader hosts, indicating that the superspreader hosts are tolerant of antibiotic-mediated perturbations in the intestinal tract. Importantly, neutralization of acute-phase proinflammatory cytokines in antibiotic-induced superspreaders suppresses the expansion of inflammatory myeloid cells and reduces morbidity. We describe a unique disease-associated tolerance to oral antibiotics in superspreaders that facilitates continued transmission of the pathogen.

  View details for DOI 10.1073/pnas.1409968111

  View details for PubMedID 25331868

• Next-Generation NAMPT Inhibitors Identified by Sequential High-Throughput Phenotypic Chemical and Functional Genomic Screens. Chemistry & biology Matheny, C. J., Wei, M. C., Bassik, M. C., Donnelly, A. J., Kampmann, M., Iwasaki, M., Piloto, O., Solow-Cordero, D. E., Bouley, D. M., Rau, R., Brown, P., McManus, M. T., Weissman, J. S., Cleary, M. L. 2013; 20 (11): 1352-1363

  Abstract

  Phenotypic high-throughput chemical screens allow for discovery of small molecules that modulate complex phenotypes and provide lead compounds for novel therapies; however, identification of the mechanistically relevant targets remains a major experimental challenge. We report the application of sequential unbiased high-throughput chemical and ultracomplex small hairpin RNA (shRNA) screens to identify a distinctive class of inhibitors that target nicotinamide phosphoribosyl transferase (NAMPT), a rate-limiting enzyme in the biosynthesis of nicotinamide adenine dinucleotide, a crucial cofactor in many biochemical processes. The lead compound STF-118804 is a highly specific NAMPT inhibitor, improves survival in an orthotopic xenotransplant model of high-risk acute lymphoblastic leukemia, and targets leukemia stem cells. Tandem high-throughput screening using chemical and ultracomplex shRNA libraries, therefore, provides a rapid chemical genetics approach for seamless progression from small-molecule lead identification to target discovery and validation.

  View details for DOI 10.1016/j.chembiol.2013.09.014

  View details for PubMedID 24183972

• Applicators for magnetic resonance-guided ultrasonic ablation of benign prostatic hyperplasia. Investigative radiology Sommer, G., Pauly, K. B., Holbrook, A., Plata, J., Daniel, B., Bouley, D., Gill, H., Prakash, P., Salgaonkar, V., Jones, P., Diederich, C. 2013; 48 (6): 387-394

  Abstract

  The aims of this study were to evaluate in a canine model applicators designed for ablation of human benign prostatic hyperplasia (BPH) in vivo under magnetic resonance imaging (MRI) guidance, including magnetic resonance thermal imaging (MRTI), determine the ability of MRI techniques to visualize ablative changes in prostate, and evaluate the acute and longer term histologic appearances of prostate tissue ablated during these studies.An MRI-compatible transurethral device incorporating a tubular transducer array with dual 120° sectors was used to ablate canine prostate tissue in vivo, in zones similar to regions of human BPH (enlarged transition zones). Magnetic resonance thermal imaging was used for monitoring of ablation in a 3-T environment, and postablation MRIs were performed to determine the visibility of ablated regions. Three canine prostates were ablated in acute studies, and 2 animals were rescanned before killing at 31 days postablation. Acute and chronic appearances of ablated prostate tissue were evaluated histologically and were correlated with the MRTI and postablation MRI scans.It was possible to ablate regions similar in size to enlarged transition zone in human BPH in 6 to 18 minutes. Regions of acute ablation showed a central "heat-fixed" region surrounded by a region of more obvious necrosis with complete disruption of tissue architecture. After 31 days, ablated regions demonstrated complete apparent resorption of ablated tissue with formation of cystic regions containing fluid. The inherent cooling of the urethra using the technique resulted in complete urethral preservation in all cases.Prostatic ablation of zones of size and shape corresponding to human BPH is possible using appropriate transurethral applicators using MRTI, and ablated tissue may be depicted clearly in contrast-enhanced magnetic resonance images. The ability accurately to monitor prostate tissue heating, the apparent resorption of ablated regions over 1 month, and the inherent urethral preservation suggest that the magnetic resonance-guided techniques described are highly promising for the in vivo ablation of symptomatic human BPH.

  View details for DOI 10.1097/RLI.0b013e31827fe91e

  View details for PubMedID 23462673

• Focal ablation of prostate cancer: four roles for magnetic resonance imaging guidance CANADIAN JOURNAL OF UROLOGY Sommer, G., Bouley, D., Gill, H., Daniel, B., Pauly, K. B., Diederich, C. 2013; 20 (2): 6672-6681

  Abstract

  There is currently a great deal of interest in the possible use of focal therapies for prostate cancer, since such treatments offer the prospect for control or cure of the primary disease with minimal side effects. Many forms of thermal therapy have been proposed for focal ablation of prostate cancer, including laser, high intensity ultrasound and cryotherapy. This review will demonstrate the important roles that magnetic resonance imaging (MRI) guidance can offer to such focal ablation, focusing on the use of high intensity ultrasonic applicators as an example of one promising technique.Transurethral and interstitial high intensity ultrasonic applicators, designed specifically for ablation of prostate tissue were tested extensively in vivo in a canine model. The roles of MRI in positioning the devices, monitoring prostate ablation, and depicting ablated tissue were assessed using appropriate MRI sequences.MRI guidance provides a very effective tool for the positioning of ablative devices in the prostate, and thermal monitoring successfully predicted ablation of prostate tissue when a threshold of 52 ºC was achieved. Contrast enhanced MRI accurately depicted the distribution of ablated prostate tissue, which is resorbed at 30 days.Guidance of thermal therapies for focal ablation of prostate cancer will likely prove critically dependent on MRI functioning in four separate roles. Our studies indicate that in three roles: device positioning; thermal monitoring of prostate ablation; and depiction of ablated prostate tissue, MR techniques are highly accurate and likely to be of great benefit in focal prostate cancer ablation. A fourth critical role, identification of cancer within the gland for targeting of thermal therapy, is more problematic at present, but will likely become practical with further technological advances.

  View details for PubMedID 23587506

• Loss of Anterior Gradient 2 (Agr2) Expression Results in Hyperplasia and Defective Lineage Maturation in the Murine Stomach JOURNAL OF BIOLOGICAL CHEMISTRY Gupta, A., Wodziak, D., Tun, M., Bouley, D. M., Lowe, A. W. 2013; 288 (6): 4321-4333

  Abstract

  Recent studies of epithelial tissues have revealed the presence of tissue-specific stem cells that are able to establish multiple cell lineages within an organ. The stem cells give rise to progenitors that replicate before differentiating into specific cell lineages. The mechanism by which homeostasis is established between proliferating stem or progenitor cells and terminally differentiated cells is unclear. This study demonstrates that Agr2 expression by mucous neck cells in the stomach promotes the differentiation of multiple cell lineages while also inhibiting the proliferation of stem or progenitor cells. When Agr2 expression is absent, gastric mucous neck cells increased in number as does the number of proliferating cells. Agr2 expression loss also resulted in the decline of terminally differentiated cells, which was supplanted by cells that exhibited nuclear SOX9 labeling. Sox9 expression has been associated with progenitor and stem cells. Similar effects of the Agr2 null on cell proliferation in the intestine were also observed. Agr2 consequently serves to maintain the balance between proliferating and differentiated epithelial cells.

  View details for DOI 10.1074/jbc.M112.433086

  View details for Web of Science ID 000314845000060

  View details for PubMedID 23209296

  View details for PubMedCentralID PMC3567683

• Caspase-11 increases susceptibility to Salmonella infection in the absence of caspase-1 NATURE Broz, P., Ruby, T., Belhocine, K., Bouley, D. M., Kayagaki, N., Dixit, V. M., Monack, D. M. 2012; 490 (7419): 288-?

  Abstract

  Inflammasomes are cytosolic multiprotein complexes assembled by intracellular nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs) and they initiate innate immune responses to invading pathogens and danger signals by activating caspase-1 (ref. 1). Caspase-1 activation leads to the maturation and release of the pro-inflammatory cytokines interleukin (IL)-1β and IL-18, as well as lytic inflammatory cell death known as pyroptosis. Recently, a new non-canonical inflammasome was described that activates caspase-11, a pro-inflammatory caspase required for lipopolysaccharide-induced lethality. This study also highlighted that previously generated caspase-1 knockout mice lack a functional allele of Casp11 (also known as Casp4), making them functionally Casp1 Casp11 double knockouts. Previous studies have shown that these mice are more susceptible to infections with microbial pathogens, including the bacterial pathogen Salmonella enterica serovar Typhimurium (S. typhimurium), but the individual contributions of caspase-1 and caspase-11 to this phenotype are not known. Here we show that non-canonical caspase-11 activation contributes to macrophage death during S. typhimurium infection. Toll-like receptor 4 (TLR4)-dependent and TIR-domain-containing adaptor-inducing interferon-β (TRIF)-dependent interferon-β production is crucial for caspase-11 activation in macrophages, but is only partially required for pro-caspase-11 expression, consistent with the existence of an interferon-inducible activator of caspase-11. Furthermore, Casp1(-/-) mice were significantly more susceptible to infection with S. typhimurium than mice lacking both pro-inflammatory caspases (Casp1(-/-) Casp11(-/-)). This phenotype was accompanied by higher bacterial counts, the formation of extracellular bacterial microcolonies in the infected tissue and a defect in neutrophil-mediated clearance. These results indicate that caspase-11-dependent cell death is detrimental to the host in the absence of caspase-1-mediated innate immunity, resulting in extracellular replication of a facultative intracellular bacterial pathogen.

  View details for DOI 10.1038/nature11419

  View details for Web of Science ID 000309733300055

  View details for PubMedID 22895188

  View details for PubMedCentralID PMC3470772

• Shape Matters: Intravital Microscopy Reveals Surprising Geometrical Dependence for Nanoparticles in Tumor Models of Extravasation NANO LETTERS Smith, B. R., Kempen, P., Bouley, D., Xu, A., Liu, Z., Melosh, N., Dai, H., Sinclair, R., Gambhir, S. S. 2012; 12 (7): 3369-3377

  Abstract

  Delivery is one of the most critical obstacles confronting nanoparticle use in cancer diagnosis and therapy. For most oncological applications, nanoparticles must extravasate in order to reach tumor cells and perform their designated task. However, little understanding exists regarding the effect of nanoparticle shape on extravasation. Herein we use real-time intravital microscopic imaging to meticulously examine how two different nanoparticles behave across three different murine tumor models. The study quantitatively demonstrates that high-aspect ratio single-walled carbon nanotubes (SWNTs) display extravasational behavior surprisingly different from, and counterintuitive to, spherical nanoparticles although the nanoparticles have similar surface coatings, area, and charge. This work quantitatively indicates that nanoscale extravasational competence is highly dependent on nanoparticle geometry and is heterogeneous.

  View details for DOI 10.1021/nl204175t

  View details for Web of Science ID 000306296200004

  View details for PubMedID 22650417

  View details for PubMedCentralID PMC3495189

• A Novel Clinically Translatable Fluorescent Nanoparticle for Targeted Molecular Imaging of Tumors in Living Subjects NANO LETTERS Gao, J., Chen, K., Luong, R., Bouley, D. M., Mao, H., Qiao, T., Gambhir, S. S., Cheng, Z. 2012; 12 (1): 281-286

  Abstract

  The use of quantum dots (QDs) in biomedical research has grown tremendously, yet successful examples of clinical applications are absent due to many clinical concerns. Here, we report on a new type of stable and biocompatible dendron-coated InP/ZnS core/shell QD as a clinically translatable nanoprobe for molecular imaging applications. The QDs (QD710-Dendron) were demonstrated to hold several significant features: near-infrared (NIR) emission, high stability in biological media, suitable size with possible renal clearance, and ability of extravasation. More importantly, a pilot mouse toxicity study confirmed that QD710-Dendron lacks significant toxicity at the doses tested. The acute tumor uptake of QD710-Dendron resulted in good contrast from the surrounding nontumorous tissues, indicating the possibility of passive targeting of the QDs. The highly specific targeting of QD710-Dendron-RGD(2) to integrin α(v)β(3)-positive tumor cells resulted in high tumor uptake and long retention of the nanoprobe at tumor sites. In summary, QD710-Dendron and RGD-modified nanoparticles demonstrate small size, high stability, biocompatibility, favorable in vivo pharmacokinetics, and successful tumor imaging properties. These features satisfy the requirements for clinical translation and should promote efforts to further investigate the possibility of using QD710-Dendron-based nanoprobes in the clinical setting in the near future.

  View details for DOI 10.1021/nl203526f

  View details for Web of Science ID 000298943100049

  View details for PubMedID 22172022

  View details for PubMedCentralID PMC3256290

• Targeting Galectin-1, a Hypoxia Induced Protein, in Non-Small Cell Lung Cancers European Multidisciplinary Cancer Congress on Integrating Basic and Translational Science, Surgery, Radiotherapy, Medical oncology, Advocacy and Care Banh, A., Zhang, J., Cao, H., Bouley, D., Kwok, S., Kong, C., Giaccia, A., Koong, A., Le, Q. ELSEVIER SCI LTD. 2011: S60–S60

  View details for Web of Science ID 000295752800212

• Targeting GLUT1 and the Warburg Effect in Renal Cell Carcinoma by Chemical Synthetic Lethality SCIENCE TRANSLATIONAL MEDICINE Chan, D. A., Sutphin, P. D., Nguyen, P., Turcotte, S., Lai, E. W., Banh, A., Reynolds, G. E., Chi, J., Wu, J., Solow-Cordero, D. E., Bonnet, M., Flanagan, J. U., Bouley, D. M., Graves, E. E., Denny, W. A., Hay, M. P., Giaccia, A. J. 2011; 3 (94)

  Abstract

  Identifying new targeted therapies that kill tumor cells while sparing normal tissue is a major challenge of cancer research. Using a high-throughput chemical synthetic lethal screen, we sought to identify compounds that exploit the loss of the von Hippel-Lindau (VHL) tumor suppressor gene, which occurs in about 80% of renal cell carcinomas (RCCs). RCCs, like many other cancers, are dependent on aerobic glycolysis for ATP production, a phenomenon known as the Warburg effect. The dependence of RCCs on glycolysis is in part a result of induction of glucose transporter 1 (GLUT1). Here, we report the identification of a class of compounds, the 3-series, exemplified by STF-31, which selectively kills RCCs by specifically targeting glucose uptake through GLUT1 and exploiting the unique dependence of these cells on GLUT1 for survival. Treatment with these agents inhibits the growth of RCCs by binding GLUT1 directly and impeding glucose uptake in vivo without toxicity to normal tissue. Activity of STF-31 in these experimental renal tumors can be monitored by [(18)F]fluorodeoxyglucose uptake by micro-positron emission tomography imaging, and therefore, these agents may be readily tested clinically in human tumors. Our results show that the Warburg effect confers distinct characteristics on tumor cells that can be selectively targeted for therapy.

  View details for DOI 10.1126/scitranslmed.3002394

  View details for PubMedID 21813754

• Tumor Galectin-1 Mediates Tumor Growth and Metastasis through Regulation of T-Cell Apoptosis CANCER RESEARCH Banh, A., Zhang, J., Cao, H., Bouley, D. M., Kwok, S., Kong, C., Giaccia, A. J., Koong, A. C., Le, Q. 2011; 71 (13): 4423-4431

  Abstract

  Galectin-1 (Gal-1), a carbohydrate-binding protein whose secretion is enhanced by hypoxia, promotes tumor aggressiveness by promoting angiogenesis and T-cell apoptosis. However, the importance of tumor versus host Gal-1 in tumor progression is undefined. Here we offer evidence that implicates tumor Gal-1 and its modulation of T-cell immunity in progression. Comparing Gal-1-deficient mice as hosts for Lewis lung carcinoma cells where Gal-1 levels were preserved or knocked down, we found that tumor Gal-1 was more critical than host Gal-1 in promoting tumor growth and spontaneous metastasis. Enhanced growth and metastasis associated with Gal-1 related to its immunomodulatory function, insofar as the benefits of Gal-1 expression to Lewis lung carcinoma growth were abolished in immunodeficient mice. In contrast, angiogenesis, as assessed by microvessel density count, was similar between tumors with divergent Gal-1 levels when examined at a comparable size. Our findings establish that tumor rather than host Gal-1 is responsible for mediating tumor progression through intratumoral immunomodulation, with broad implications in developing novel targeting strategies for Gal-1 in cancer.

  View details for DOI 10.1158/0008-5472.CAN-10-4157

  View details for PubMedID 21546572

• Identification of an Ire1alpha endonuclease specific inhibitor with cytotoxic activity against human multiple myeloma BLOOD Papandreou, I., Denko, N. C., Olson, M., Van Melckebeke, H., Lust, S., Tam, A., Solow-Cordero, D. E., Bouley, D. M., Offner, F., Niwa, M., Koong, A. C. 2011; 117 (4): 1311-1314

  Abstract

  Activation of the adaptive Ire1-XBP1 pathway has been identified in many solid tumors and hematologic malignancies, including multiple myeloma (MM). Here, we report the identification of STF-083010, a novel small-molecule inhibitor of Ire1. STF-083010 inhibited Ire1 endonuclease activity, without affecting its kinase activity, after endoplasmic reticulum stress both in vitro and in vivo. Treatment with STF-083010 showed significant antimyeloma activity in model human MM xenografts. Similarly, STF-083010 was preferentially toxic to freshly isolated human CD138(+) MM cells compared with other similarly isolated cell populations. The identification of this novel Ire1 inhibitor supports the hypothesis that the Ire1-XBP1 axis is a promising target for anticancer therapy, especially in the context of MM.

  View details for DOI 10.1182/blood-2010-08-303099

  View details for Web of Science ID 000286623400029

  View details for PubMedID 21081713

  View details for PubMedCentralID PMC3056474

• EFFECTS OF RAINFALL, HOST DEMOGRAPHY, AND MUSTH ON STRONGYLE FECAL EGG COUNTS IN AFRICAN ELEPHANTS (LOXODONTA AFRICANA) IN NAMIBIA JOURNAL OF WILDLIFE DISEASES Thurber, M. I., O'Connell-Rodwell, C. E., Turner, W. C., Nambandi, K., KINZLEY, C., Rodwel, T. C., Faulkner, C. T., Felt, S. A., Bouley, D. M. 2011; 47 (1): 172-181

  Abstract

  Wild African elephants (Loxodonta africana) are commonly infected with intestinal strongyle parasites. Our objective was to determine baseline fecal strongyle egg counts for elephants in the northeast region of Etosha National Park, Namibia and determine if these numbers were affected by annual rainfall, elephant demography (age of individuals and composition of groups), and hormonal state of males. We found that matriarchal family group members have significantly higher fecal egg counts than male elephants (bulls). Among family group members, strongyle egg counts increased with age, whereas among bulls, strongyle egg counts decreased with age. Years of higher rainfall were correlated with decreased numbers of strongyle eggs among bulls. Finally, bulls were not affected by their physiologic (hormonal) status (musth vs. nonmusth). These results suggest that infection by strongyle parasites in Namibian African elephants is a dynamic process affected by intrinsic and extrinsic factors including host demography and rainfall.

  View details for Web of Science ID 000287280200016

  View details for PubMedID 21270006

• Loss of polyubiquitin gene Ubb leads to metabolic and sleep abnormalities in mice NEUROPATHOLOGY AND APPLIED NEUROBIOLOGY Ryu, K., Fujiki, N., Kazantzis, M., Garza, J. C., Bouley, D. M., Stahl, A., Lu, X., Nishino, S., Kopito, R. R. 2010; 36 (4): 285-299

  Abstract

  Ubiquitin performs essential roles in a myriad of signalling pathways required for cellular function and survival. Recently, we reported that disruption of the stress-inducible ubiquitin-encoding gene Ubb reduces ubiquitin content in the hypothalamus and leads to adult-onset obesity coupled with a loss of arcuate nucleus neurones and disrupted energy homeostasis in mice. Neuropeptides expressed in the hypothalamus control both metabolic and sleep behaviours. In order to demonstrate that the loss of Ubb results in broad hypothalamic abnormalities, we attempted to determine whether metabolic and sleep behaviours were altered in Ubb knockout mice.Metabolic rate and energy expenditure were measured in a metabolic chamber, and sleep stage was monitored via electroencephalographic/electromyographic recording. The presence of neurodegeneration and increased reactive gliosis in the hypothalamus were also evaluated.We found that Ubb disruption leads to early-onset reduced activity and metabolic rate. Additionally, we have demonstrated that sleep behaviour is altered and sleep homeostasis is disrupted in Ubb knockout mice. These early metabolic and sleep abnormalities are accompanied by persistent reactive gliosis and the loss of arcuate nucleus neurones, but are independent of neurodegeneration in the lateral hypothalamus.Ubb knockout mice exhibit phenotypes consistent with hypothalamic dysfunction. Our data also indicate that Ubb is essential for the maintenance of the ubiquitin levels required for proper regulation of metabolic and sleep behaviours in mice.

  View details for DOI 10.1111/j.1365-2990.2009.01057.x

  View details for Web of Science ID 000277712100003

  View details for PubMedID 20002312

• The RGD Domain of Human Osteopontin Promotes Tumor Growth and Metastasis through Activation of Survival Pathways PLOS ONE Courter, D., Cao, H., Kwok, S., Kong, C., Banh, A., Kuo, P., Bouley, D. M., Vice, C., Brustugun, O. T., Denko, N. C., Koong, A. C., Giaccia, A., Le, Q. 2010; 5 (3)

  Abstract

  Human osteopontin (OPN), a known tumor associated protein, exists in different isoforms, whose function is unclear. It also possesses a RGD domain, which has been implicated in diverse function. Here, we use genetic approaches to systematically investigate the function of the RGD domain in different OPN isoforms on tumor progression and metastasis for 2 different solid tumor models.Using isoform-specific qRT-PCR, we found that OPN-A and B were the main isoforms overexpressed in evaluated human tumors, which included 4 soft tissue sarcomas, 24 lung and 30 head and neck carcinomas. Overexpression of either OPN-A or B in two different cell types promoted local tumor growth and lung metastasis in SCID mouse xenografts. However, expression of either isoform with the RGD domain either mutated or deleted decreased tumor growth and metastasis, and resulted in increased apoptosis by TUNEL staining. In vitro, whereas mutation of the RGD domain did not affect cell-cell adhesion, soft agar growth or cell migration, it increased apoptosis under hypoxia and serum starvation. This effect could be mitigated when the RGD mutant cells were treated with condition media containing WT OPN. Mechanistically, the RGD region of OPN inhibited apoptosis by inducing NF-kappaB activation and FAK phosphorylation. Inhibition of NF-kappaB (by siRNA to the p65 subunit) or FAK activation (by a inhibitor) significantly increased apoptosis under hypoxia in WT OPN cells, but not in RGD mutant cells.Unlike prior reports, our data suggest that the RGD domain of both OPN-A and B promote tumor growth and metastasis mainly by protecting cells against apoptosis under stressed conditions and not via migration or invasion. Future inhibitors directed against OPN should target multiple isoforms and should inhibit cell survival mechanisms that involve the RGD domain, FAK phosphorylation and NF-kappaB activation.

  View details for DOI 10.1371/journal.pone.0009633

  View details for PubMedID 20224789

• Circulating tumour cells demonstrate an altered response to hypoxia and an aggressive phenotype BRITISH JOURNAL OF CANCER Ameri, K., Luong, R., Zhang, H., Powell, A. A., Montgomery, K. D., Espinosa, I., Bouley, D. M., Harris, A. L., Jeffrey, S. S. 2010; 102 (3): 561-569

  Abstract

  Tumours contain hypoxic regions that select for an aggressive cell phenotype; tumour hypoxia induces metastasis-associated genes. Treatment refractory patients with metastatic cancer show increased numbers of circulating tumour cells (CTCs), which are also associated with disease progression. The aim of this study was to examine the as yet unknown relationship between hypoxia and CTCs.We generated human MDA-MB-231 orthotopic xenografts and, using a new technology, isolated viable human CTCs from murine blood. The CTCs and parental MDA-MB-231 cells were incubated at 21 and 0.2% (hypoxia) oxygen, respectively. Colony formation was assayed and levels of hypoxia- and anoxia-inducible factors were measured. Xenografts generated from CTCs and parental cells were compared.MDA-MB-231 xenografts used to generate CTCs were hypoxic, expressing hypoxia factors: hypoxia-inducible factor1 alpha (HIF1alpha) and glucose transporter protein type 1 (GLUT1), and anoxia-induced factors: activating transcription factor 3 and 4 (ATF3 and ATF4). Parental MDA-MB-231 cells induced ATF3 in hypoxia, whereas CTCs expressed it constitutively. Asparagine synthetase (ASNS) expression was also higher in CTCs. Hypoxia induced ATF4 and the HIF1alpha target gene apelin in CTCs, but not in parental cells. Hypoxia induced lower levels of carbonic anhydrase IX (CAIX), GLUT1 and BCL2/adenovirus E1B 19-KD protein-interacting protein 3 (BNIP3) proteins in CTCs than in parental cells, supporting an altered hypoxia response. In chronic hypoxia, CTCs demonstrated greater colony formation than parental cells. Xenografts generated from CTCs were larger and heavier, and metastasised faster than MDA-MB-231 xenografts.CTCs show an altered hypoxia response and an enhanced aggressive phenotype in vitro and in vivo.

  View details for DOI 10.1038/sj.bjc.6605491

  View details for Web of Science ID 000274194700015

  View details for PubMedID 20051957

  View details for PubMedCentralID PMC2805847

• Short-term stress enhances cellular immunity and increases early resistance to squamous cell carcinoma BRAIN BEHAVIOR AND IMMUNITY Dhabhar, F. S., Saul, A. N., Daugherty, C., Holmes, T. H., Bouley, D. M., Oberyszyn, T. M. 2010; 24 (1): 127-137

  Abstract

  In contrast to chronic/long-term stress that suppresses/dysregulates immune function, an acute/short-term fight-or-flight stress response experienced during immune activation can enhance innate and adaptive immunity. Moderate ultraviolet-B (UV) exposure provides a non-invasive system for studying the naturalistic emergence, progression and regression of squamous cell carcinoma (SCC). Because SCC is an immunoresponsive cancer, we hypothesized that short-term stress experienced before UV exposure would enhance protective immunity and increase resistance to SCC. Control and short-term stress groups were treated identically except that the short-term stress group was restrained (2.5h) before each of nine UV-exposure sessions (minimum erythemal dose, 3-times/week) during weeks 4-6 of the 10-week UV exposure protocol. Tumors were measured weekly, and tissue collected at weeks 7, 20, and 32. Chemokine and cytokine gene expression was quantified by real-time PCR, and CD4+ and CD8+ T cells by flow cytometry and immunohistochemistry. Compared to controls, the short-term stress group showed greater cutaneous T-cell attracting chemokine (CTACK)/CCL27, RANTES, IL-12, and IFN-gamma gene expression at weeks 7, 20, and 32, higher skin infiltrating T cell numbers (weeks 7 and 20), lower tumor incidence (weeks 11-20) and fewer tumors (weeks 11-26). These results suggest that activation of short-term stress physiology increased chemokine expression and T cell trafficking and/or function during/following UV exposure, and enhanced Type 1 cytokine-driven cell-mediated immunity that is crucial for resistance to SCC. Therefore, the physiological fight-or-flight stress response and its adjuvant-like immuno-enhancing effects, may provide a novel and important mechanism for enhancing immune system mediated tumor-detection/elimination that merits further investigation.

  View details for DOI 10.1016/j.bbi.2009.09.004

  View details for Web of Science ID 000272676400017

  View details for PubMedID 19765644

  View details for PubMedCentralID PMC2788066

• Where's the Mouse Pathology Training? VETERINARY PATHOLOGY Bouley, D., Meyerholz, D., Sellers, R., Boyd, K., La Perle, K. 2009; 46 (6): 1245-1247

  View details for DOI 10.1354/vp.09-VP-0185-B-COM

  View details for Web of Science ID 000271801600025

  View details for PubMedID 19890103

• MRI-Guided Cryoablation: In Vivo Assessment of Focal Canine Prostate Cryolesions JOURNAL OF MAGNETIC RESONANCE IMAGING Josan, S., Bouley, D. M., van den Bosch, M., Daniel, B. L., Pauly, K. B. 2009; 30 (1): 169-176

  Abstract

  To analyze the appearance of acute and chronic canine prostate cryolesions on T1-weighted (T1w) and T2-weighted (T2w) magnetic resonance imaging (MRI) and compare them with contrast-enhanced (CE) MRI and histology for a variety of freezing protocols.Three different freezing protocols were used in canine prostate cryoablation experiments. Six acute and seven chronic (survival times ranging between 4-53 days) experiments were performed. The change in T2w signal intensity was correlated with freezing protocol parameters. The lesion area on T2w MRI was compared to CE-MRI. Histopathologic evaluation of the cryolesions was performed and visually compared to the appearance on MRI.The T2w signal increased from pre- to postfreeze at the site of the cryolesion, and the enhancement was higher for smaller freeze area and duration. The T2w lesion area was between the CE nonperfused area and the hyperenhancing CE rim. The appearance of the lesion on T1w and T2w imaging over time correlated with outcome on pathology.T1w and T2w MRI can potentially be used to assess cryolesions and to monitor tissue response over time following cryoablation.

  View details for DOI 10.1002/jmri.21827

  View details for PubMedID 19557805

• MR Imaging-guided Percutaneous Cryoablation of the Prostate in an Animal Model: In Vivo Imaging of Cryoablation-induced Tissue Necrosis with Immediate Histopathologic Correlation 32nd Annual Meeting of the Society-of-Interventional-Radiology (SIR) van den Bosch, M. A., Josan, S., Bouley, D. M., Chen, J., Gill, H., Rieke, V., Butts-Pauly, K., Daniel, B. L. ELSEVIER SCIENCE INC. 2009: 252–58

  Abstract

  To evaluate the feasibility of magnetic resonance (MR) imaging-guided percutaneous cryoablation of normal canine prostates and to identify MR imaging features that accurately predict the area of tissue damage at a microscopic level.Six adult male mixed-breed dogs were anesthetized, intubated, and placed in a 0.5-T open MR imaging system. A receive-only endorectal coil was placed, and prostate location and depth were determined on T1-weighted fast spin-echo (FSE) MR imaging. After placement of cryoprobes and temperature sensors, three freezing protocols were used to ablate prostate tissue. Ice ball formation was monitored with T1-weighted FSE imaging. Tissue necrosis area was assessed with contrast-enhanced weighted MR imaging and compared with histopathologic findings.A total of 12 cryolesions (mean size, 1.2 cm) were bilaterally created in six prostates. Ice ball formation was oval and signal-free on T1-weighted FSE sequences in all cases. Postprocedural contrast-enhanced MR imaging typically showed a nonenhancing area of low signal intensity centrally located within the frozen area, surrounded by a bright enhancing rim in all cases. On histopathologic examination, two distinct zones were identified within cryolesions. Centrally, a necrotic zone with complete cellular destruction and hemorrhage was found. Between this necrotic zone and normal glandular tissue, a zone of fragmented and intact glands, interstitial edema, and rare acute inflammatory cells was seen. Correlation between nonenhancement on contrast-enhanced weighted MR images and tissue necrosis on pathologic examination was consistent within all six dogs.MR imaging-guided cryoablation of the prostate is technically feasible. The nonenhancing area on postablation contrast-enhanced weighted MR imaging accurately predicts the area of cryoablation-induced tissue necrosis on pathologic analysis.

  View details for DOI 10.1016/j.jvir.2008.10.030

  View details for PubMedID 19091600

• Catheter-Based Ultrasound for 3D Control of Thermal Therapy 8th International Symposium on Therapeutic Ultras Diederich, C., Chen, X., Wootton, J., Juang, T., Nau, W. H., Kinsey, A., Hsu, I., Rieke, V., Pauly, K. B., Sommer, G., Bouley, D. AMER INST PHYSICS. 2009: 318–322

  View details for Web of Science ID 000266425300061

• Characterization of a novel anthropomorphic plastinated lung phantom MEDICAL PHYSICS Yoon, S., Henry, R. W., Bouley, D. M., Bennett, N. R., Fahrig, R. 2008; 35 (12): 5934-5943

  Abstract

  Phantoms are widely used during the development of new imaging systems and algorithms. For development and optimization of new imaging systems such as tomosynthesis, where conventional image quality metrics may not be applicable, a realistic phantom that can be used across imaging systems is desirable. A novel anthropomorphic lung phantom was developed by plastination of an actual pig lung. The plastinated phantom is characterized and compared with reference to in vivo images of the same tissue prior to plastination using high resolution 3D CT. The phantom is stable over time and preserves the anatomical features and relative locations of the in vivo sample. The volumes for different tissue types in the phantom are comparable to the in vivo counterparts, and CT numbers for different tissue types fall within a clinically useful range. Based on the measured CT numbers, the phantom cardiac tissue experienced a 92% decrease in bulk density and the phantom pulmonary tissue experienced a 78% decrease in bulk density compared to their in vivo counterparts. By-products in the phantom from the room temperature vulcanizing silicone and plastination process are also identified. A second generation phantom, which eliminates most of the by-products, is presented. Such anthropomorphic phantoms can be used to evaluate a wide range of novel imaging systems.

  View details for DOI 10.1118/1.3016524

  View details for PubMedID 19175148

• Monitoring prostate thermal therapy with diffusion-weighted MRI MAGNETIC RESONANCE IN MEDICINE Chen, J., Daniel, B. L., Diederich, C. J., Bouley, D. M., Van den Bosch, M. A., Kinsey, A. M., Sommer, G., Pauly, K. B. 2008; 59 (6): 1365-1372

  Abstract

  For MR-guided minimally invasive therapies, it is important to have a repeatable and reliable tissue viability evaluation method. The use of diffusion-weighted MRI (DWI) to evaluate tissue damage was assessed in 19 canine prostates with cryoablation or high-intensity ultrasound (HIU) ablation. The apparent diffusion coefficient (ADC) trace value was measured in the treated tissue immediately upon the procedure and on the posttreatment follow-up. For the acute lesions, the ADC value decreased to (1.05+/-0.25)x10(-3) mm2/s, as compared to (1.64+/-0.24)x10(-3) mm2/s before the treatment. There was no statistical difference between previously frozen or previously ultrasound-heated lesions in terms of the 36% ADC reduction (P=0.66). The ADC decrease occurred early during the course of the treatment, which appears to complicate DWI-based thermometry. Over time, the ADC value increased as the tissue recovered and regenerated. This study shows that DWI could be a promising method to monitor prostate thermal therapies and to provide insight on tissue damage and tissue remodeling after injury.

  View details for DOI 10.1002/mrm.21589

  View details for PubMedID 18506801

• Transurethral ultrasound applicators with dynamic multi-sector control for prostate thermal therapy: In vivo evaluation under MR guidance MEDICAL PHYSICS Kinsey, A. M., Diederich, C. J., Rieke, V., Nau, W. H., Pauly, K. B., Bouley, D., Sommer, G. 2008; 35 (5): 2081-2093

  Abstract

  The purpose of this study was to explore the feasibility and performance of a multi-sectored tubular array transurethral ultrasound applicator for prostate thermal therapy, with potential to provide dynamic angular and length control of heating under MR guidance without mechanical movement of the applicator. Test configurations were fabricated, incorporating a linear array of two multi-sectored tubular transducers (7.8-8.4 MHz, 3 mm OD, 6 mm length), with three 120 degrees independent active sectors per tube. A flexible delivery catheter facilitated water cooling (100 ml min(-1)) within an expandable urethral balloon (35 mm long x 10 mm diameter). An integrated positioning hub allows for rotating and translating the transducer assembly within the urethral balloon for final targeting prior to therapy delivery. Rotational beam plots indicate approximately 90 degrees-100 degrees acoustic output patterns from each 120 degrees transducer sector, negligible coupling between sectors, and acoustic efficiencies between 41% and 53%. Experiments were performed within in vivo canine prostate (n = 3), with real-time MR temperature monitoring in either the axial or coronal planes to facilitate control of the heating profiles and provide thermal dosimetry for performance assessment. Gross inspection of serial sections of treated prostate, exposed to TTC (triphenyl tetrazolium chloride) tissue viability stain, allowed for direct assessment of the extent of thermal coagulation. These devices created large contiguous thermal lesions (defined by 52 degrees C maximum temperature, t43 = 240 min thermal dose contours, and TTC tissue sections) that extended radially from the applicator toward the border of the prostate (approximately15 mm) during a short power application (approximately 8-16 W per active sector, 8-15 min), with approximately 200 degrees or 360 degrees sector coagulation demonstrated depending upon the activation scheme. Analysis of transient temperature profiles indicated progression of lethal temperature and thermal dose contours initially centered on each sector that coalesced within approximately 5 min to produce uniform and contiguous zones of thermal destruction between sectors, with smooth outer boundaries and continued radial propagation in time. The dimension of the coagulation zone along the applicator was well-defined by positioning and active array length. Although not as precise as rotating planar and curvilinear devices currently under development for MR-guided procedures, advantages of these multi-sectored transurethral applicators include a flexible delivery catheter and that mechanical manipulation of the device using rotational motors is not required during therapy. This multi-sectored tubular array transurethral ultrasound technology has demonstrated potential for relatively fast and reasonably conformal targeting of prostate volumes suitable for the minimally invasive treatment of BPH and cancer under MR guidance, with further development warranted.

  View details for DOI 10.1118/1.2900131

  View details for Web of Science ID 000255456500049

  View details for PubMedID 18561684

  View details for PubMedCentralID PMC2673638

• Experimental study of intracranial hematoma detection with flat panel detector C-arm CT AMERICAN JOURNAL OF NEURORADIOLOGY Arakawa, H., Marks, M. P., Do, H. M., Bouley, D. M., Strobel, N., Moore, T., Fahrig, R. 2008; 29 (4): 766-772

  Abstract

  Intracranial hemorrhage is a commonly acknowledged complication of interventional neuroradiology procedures, and the ability to image hemorrhage at the time of the procedure would be very beneficial. A new C-arm system with 3D functionality extends the capability of C-arm imaging to include soft-tissue applications by facilitating the detection of low-contrast objects. We evaluated its ability to detect small intracranial hematomas in a swine model.Intracranial hematomas were created in 7 swine by autologous blood injection of various hematocrits (19%-37%) and volumes (1.5-5 mL). Four animals received intravascular contrast before obtaining autologous blood (group 1), and 3 did not (group 2). We scanned each animal by using the C-arm CT system, acquiring more than 500 images during a 20-second rotation through more than 200 degrees . Multiplanar reformatted images with isotropic resolution were reconstructed on the workstation by using product truncation, scatter, beam-hardening, and ring-artifact correction algorithms. The brains were harvested and sliced for hematoma measurement and compared with imaging findings.Five intracranial hematomas were created in group 1 animals, and all were visualized. Six were created in group 2, and 3 were visualized. One nonvisualized hematoma was not confirmed at necropsy. All the others in both groups were confirmed. In group 1 (with contrast), small hematomas were detectable even when the hematocrit was 19%-20%. In group 2 (without contrast) C-arm CT was able to detect small hematomas (<1.0 cm(2)) created with hematocrits of 29%-37%. The area of hematoma measured from the C-arm CT data was, on average, within 15% of the area measured from harvested brain.The image quality obtained with this implementation of C-arm CT was sufficient to detect experimentally created small intracranial hematomas. This capability should provide earlier detection of hemorrhagic complications that may occur during neurointerventional procedures.

  View details for DOI 10.3174/ajnr.A0898

  View details for Web of Science ID 000255129700029

  View details for PubMedID 18202240

• The mouse polyubiquitin gene Ubb is essential for meiotic progression MOLECULAR AND CELLULAR BIOLOGY Ryu, K., Sinnar, S. A., Reinholdt, L. G., Vaccari, S., Hall, S., Garcia, M. A., Zaitseva, T. S., Bouley, D. M., Boekelheide, K., Handel, M. A., Conti, M., Kopito, R. R. 2008; 28 (3): 1136-1146

  Abstract

  Ubiquitin is encoded in mice by two polyubiquitin genes, Ubb and Ubc, that are considered to be stress inducible and two constitutively expressed monoubiquitin (Uba) genes. Here we report that targeted disruption of Ubb results in male and female infertility due to failure of germ cells to progress through meiosis I and hypogonadism. In the absence of Ubb, spermatocytes and oocytes arrest during meiotic prophase, before metaphase of the first meiotic division. Although cellular ubiquitin levels are believed to be maintained by a combination of functional redundancy among the four ubiquitin genes, stress inducibility of the two polyubiquitin genes, and ubiquitin recycling by proteasome-associated isopeptidases, our results indicate that ubiquitin is required for and consumed during meiotic progression. The striking similarity of the meiotic phenotype in Ubb(-/-) germ cells to the sporulation defect in fission yeast (Schizosaccharomyces pombe) lacking a polyubiquitin gene suggests that a meiotic role of the polyubiquitin gene has been conserved throughout eukaryotic evolution.

  View details for DOI 10.1128/MCB.01566-07

  View details for Web of Science ID 000252606100024

  View details for PubMedID 18070917

  View details for PubMedCentralID PMC2223379

• Host transmission of Salmonella enterica serovar Typhimurium is controlled by virulence factors and indigenous intestinal microbiota INFECTION AND IMMUNITY Lawley, T. D., Bouley, D. A., Hoy, Y. E., Gerke, C., Relman, D. A., Monack, D. M. 2008; 76 (1): 403-416

  Abstract

  Transmission is an essential stage of a pathogen's life cycle and remains poorly understood. We describe here a model in which persistently infected 129X1/SvJ mice provide a natural model of Salmonella enterica serovar Typhimurium transmission. In this model only a subset of the infected mice, termed supershedders, shed high levels (>10(8) CFU/g) of Salmonella serovar Typhimurium in their feces and, as a result, rapidly transmit infection. While most Salmonella serovar Typhimurium-infected mice show signs of intestinal inflammation, only supershedder mice develop colitis. Development of the supershedder phenotype depends on the virulence determinants Salmonella pathogenicity islands 1 and 2, and it is characterized by mucosal invasion and, importantly, high luminal abundance of Salmonella serovar Typhimurium within the colon. Immunosuppression of infected mice does not induce the supershedder phenotype, demonstrating that the immune response is not the main determinant of Salmonella serovar Typhimurium levels within the colon. In contrast, treatment of mice with antibiotics that alter the health-associated indigenous intestinal microbiota rapidly induces the supershedder phenotype in infected mice and predisposes uninfected mice to the supershedder phenotype for several days. These results demonstrate that the intestinal microbiota plays a critical role in controlling Salmonella serovar Typhimurium infection, disease, and transmissibility. This novel model should facilitate the study of host, pathogen, and intestinal microbiota factors that contribute to infectious disease transmission.

  View details for DOI 10.1128/IAI.01189-07

  View details for Web of Science ID 000252126000043

  View details for PubMedID 17967858

  View details for PubMedCentralID PMC2223630

• Catheter-Based Ultrasound Devices and MR Thermal Monitoring for Conformal Prostate Thermal Therapy 30th Annual International Conference of the IEEE-Engineering-in-Medicine-and-Biology-Society Diederich, C. J., Nau, W. H., Kinsey, A., Ross, T., Wootton, J., Juang, T., Butts-Pauly, K., Rieke, V., Chen, J., Bouley, D. M., Sommer, G. IEEE. 2008: 3664–3668

  Abstract

  Catheter-based ultrasound applicators have been developed for delivering hyperthermia or high-temperature thermal ablation of cancer and benign disease of the prostate. These devices allow for control of heating along the length and angular expanse during therapy delivery. Four types of transurethral applicators were devised for thermal treatment of prostate combined with MR thermal monitoring: sectored tubular transducer devices with directional heating patterns and rotation; planar and curvilinear devices with narrow heating patterns and rotation; and multi-sectored tubular devices capable of dynamic angular control without applicator movement. Interstitial devices (2.4 mm OD) have been developed for percutaneous implantation with directional or dynamic angular control. In vivo experiments in canine prostate under MR temperature imaging were used to evaluate these devices and develop treatment delivery strategies. MR thermal imaging was used to monitor temperature and thermal dose in multiple slices through the target volume. Multi-sectored transurethral applicators can dynamically control the angular heating profile and target large regions of the gland in short treatment times without applicator manipulation. The sectored tubular, planar, and curvilinear transurethral devices produce directional coagulation zones, extending 15-20 mm radial distance to the outer prostate capsule. Sequential rotation under motor control and modulated dwell time can be used to tightly conform thermal ablation to selected regions. Interstitial implants with directional devices can be used to effectively ablate targeted regions of the gland while protecting the rectum. The MR derived 52 degrees C and lethal thermal dose contours (t43=240 min) effectively defined the extent of thermal damage and provided a means for real-time control of the applicators. Catheter-based ultrasound devices, combined with MR thermal monitoring, can produce relatively fast (5-40 min) and precise thermal ablation of prostate.

  View details for Web of Science ID 000262404502111

  View details for PubMedID 19163505

• The distribution, density and three-dimensional histomorphology of Pacinian corpuscles in the foot of the Asian elephant (Elephas maximus) and their potential role in seismic communication JOURNAL OF ANATOMY Bouley, D. M., Alarcon, C. N., Hildebrandt, T., O'Connell-Rodwell, C. E. 2007; 211 (4): 428-435

  Abstract

  Both Asian (Elephas maximus) and African (Loxodonta africana) elephants produce low-frequency, high-amplitude rumbles that travel well through the ground as seismic waves, and field studies have shown that elephants may utilize these seismic signals as one form of communication. Unique elephant postures observed in field studies suggest that the elephants use their feet to 'listen' to these seismic signals, but the exact sensory mechanisms used by the elephant have never been characterized. The distribution, morphology and tissue density of Pacinian corpuscles, specialized mechanoreceptors, were studied in a forefoot and hindfoot of Asian elephants. Pacinian corpuscles were located in the dermis and distal digital cushion and were most densely localized to the anterior, posterior, medial and lateral region of each foot, with the highest numbers in the anterior region of the forefoot (52.19%) and the posterior region of the hindfoot (47.09%). Pacinian corpuscles were encapsulated, had a typical lamellar structure and were most often observed in large clusters. Three-dimensional reconstruction through serial sections of the dermis revealed that individual Pacinian corpuscles may be part of a cluster. By studying the distribution and density of these mechanoreceptors, we propose that Pacinian corpuscles are one possible anatomic mechanism used by elephants to detect seismic waves.

  View details for DOI 10.1111/j.1469-7580.2007.00792.x

  View details for Web of Science ID 000249826500002

  View details for PubMedID 17711421

  View details for PubMedCentralID PMC2375831

• The mouse polyubiquitin gene UbC is essential for fetal liver development, cell-cycle progression and stress tolerance EMBO JOURNAL Ryu, K., Maehr, R., Gilchrist, C. A., Long, M. A., Bouley, D. M., Mueller, B., Ploegh, H. L., Kopito, R. R. 2007; 26 (11): 2693-2706

  Abstract

  UbC is one of two stress-inducible polyubiquitin genes in mammals and is thought to supplement the constitutive UbA genes in maintaining cellular ubiquitin (Ub) levels during episodes of cellular stress. We have generated mice harboring a targeted disruption of the UbC gene. UbC(-/-) embryos die between embryonic days 12.5 and 14.5 in utero, most likely owing to a severe defect in liver cell proliferation. Mouse embryonic fibroblasts from UbC(-/-) embryos exhibit reduced growth rates, premature senescence, increased apoptosis and delayed cell-cycle progression, with slightly, but significantly, decreased steady-state Ub levels. UbC(-/-) fibroblasts are hypersensitive to proteasome inhibitors and heat shock, and unable to adequately increase Ub levels in response to these cellular stresses. Most, but not all of the UbC(-/-) phenotypes can be rescued by providing additional Ub from a poly hemagglutinin-tagged Ub minigene expressed from the Hprt locus. We propose that UbC is regulated by a process that senses Ub pool dynamics. These data establish that UbC constitutes an essential source of Ub during cell proliferation and stress that cannot be compensated by other Ub genes.

  View details for DOI 10.1038/sj.emboj.7601722

  View details for Web of Science ID 000247084100008

  View details for PubMedID 17491588

  View details for PubMedCentralID PMC1888680

• Ovarian hyperstimulation syndrome in gonadotropin-treated laboratory South African clawed frogs (Xenopus laevis) JOURNAL OF THE AMERICAN ASSOCIATION FOR LABORATORY ANIMAL SCIENCE Green, S. L., Parker, J., Davis, C., Bouley, D. M. 2007; 46 (3): 64-67

  Abstract

  Ovarian hyperstimulation syndrome (OHS) is a rare but sometimes fatal iatrogenic complication of ovarian stimulation associated with the administration of exogenous gonadotropins to women undergoing treatment for infertility. Laboratory Xenopus spp are commonly treated with human chorionic gonadotropin (hCG) to stimulate ovulation and optimize the number of oocytes harvested for use in biomedical research. Here we report cases of OHS in 2 gonadotropin-treated laboratory Xenopus laevis. After receiving hCG, the frogs developed severe subcutaneous accumulation of fluid, coelomic distention, and whole-body edema and were unable to dive, although they continued to eat and swim. At postmortem examination, extensive subcutaneous edema was present; ascites and massive numbers of free-floating eggs were found in the coelomic cavity and in aberrant locations: around the heart-sac and adhered to the liver capsule. Whole-body edema, gross enlargement of the ovaries, ascites, and abdominal distention are findings comparable to those observed in women with OHS. The pathophysiology of OHS is thought to be related to hormonally induced disturbances of vasoactive mediators, one of which may be vascular endothelial growth factor secreted by theca and granulosa cells. We know of no other report describing OHSlike symptoms in gonadotropin-treated frogs, and the cases described here are 2 of the 3 we have observed at our respective institutions over the last 6 y. According to these results, OHS appears to be rare in gonadotropin-treated laboratory Xenopus. However, the condition should be included in the differential diagnosis for the bloated frog.

  View details for Web of Science ID 000246535200013

  View details for PubMedID 17487957

• Determination of 3-dimensional zonal renal volumes using contrast-enhanced computed tomography JOURNAL OF COMPUTER ASSISTED TOMOGRAPHY Sommer, G., Bouley, D., Frisoli, J., Pierce, L., Sandner-Porkristl, D., Fahrig, R. 2007; 31 (2): 209-213

  Abstract

  To determine the accuracy with which total and cortical renal volumes may be measured in vivo with contrast-enhanced computed tomography (CT) in an animal model.Seven female Yorkshire pigs were scanned in vivo using both a 64-channel multidetector row computed tomography (MDCT) scanner (Siemens Medical Solutions, Erlangen, Germany) and a C-arm CT scanner (Siemens AXIOM). Kidneys were scanned in corticomedullary and nephrographic phases after contrast injection. Three-dimensional volumetric analysis of CT data was performed to determine renal cortical and overall renal volumes, and renal lengths. We measured renal weights, water displacement volumes, and overall and cortical renal volumes of resected kidneys.Overall and cortical renal volumes by CT showed excellent correlation (R2 values varying from 0.77 to 0.92) with anatomic measurements of renal volume. There was poor correlation of anatomic renal volumes with renal lengths by CT.Three-dimensional analysis of contrast-enhanced CT provides accurate measurement of overall and cortical renal volumes in vivo.

  View details for Web of Science ID 000245456700009

  View details for PubMedID 17414755

• Correlation of contrast-enhanced MR images with the histopathology of minimally invasive thermal and cryoablation cancer treatments in normal dog prostates. Proceedings of SPIE--the International Society for Optical Engineering Bouley, D. M., Daniel, B., Pauly, K. B., Liu, E., Kinsey, A., NAU, W., Diederich, C. J., Sommer, G. 2007; 6440: 644006-?

  Abstract

  Magnetic Resonance Imaging (MRI) is a promising tool for visualizing the delivery of minimally invasive cancer treatments such as high intensity ultrasound (HUS) and cryoablation. We use an acute dog prostate model to correlate lesion histopathology with contrast-enhanced (CE) T1 weighted MR images, to aid the radiologists in real time interpretation of in vivo lesion boundaries and pre-existing lesions. Following thermal or cryo treatments, prostate glands are removed, sliced, stained with the vital dye triphenyl tetrazolium chloride, photographed, fixed and processed in oversized blocks for routine microscopy. Slides are scanned by Trestle Corporation at .32 microns/pixel resolution, the various lesions traced using annotation software, and digital images compared to CE MR images. Histologically, HUS results in discrete lesions characterized by a "heat-fixed" zone, in which glands subjected to the highest temperatures are minimally altered, surrounded by a rim or "transition zone" composed of severely fragmented, necrotic glands, interstitial edema and vascular congestion. The "heat-fixed" zone is non-enhancing on CE MRI while the "transition zone" appears as a bright, enhancing rim. Likewise, the CE MR images for cryo lesions appear similar to thermally induced lesions, yet the histopathology is significantly different. Glands subjected to prolonged freezing appear totally disrupted, coagulated and hemorrhagic, while less intensely frozen glands along the lesion edge are partially fragmented and contain apoptotic cells. In conclusion, thermal and cryo-induced lesions, as well as certain pre-existing lesions (cystic hyperplasia - non-enhancing, chronic prostatitis - enhancing) have particular MRI profiles, useful for treatment and diagnostic purposes.

  View details for PubMedID 25076818

• Evaluation of thermal and cryo lesions by diffusion-weighted MRI Conference on Thermal Treatment of Tissue - Energy Delivery and Assessment IV Chen, J., Daniel, B., Bouley, D., Sommer, G., Butts Pauly, K. SPIE-INT SOC OPTICAL ENGINEERING. 2007

  View details for DOI 10.1117/12.703255

  View details for Web of Science ID 000246494100006

• Correlation of contrast-enhanced NM images with the histopathology of minimally invasive thermal and cryoablation cancer treatments in normal dog prostates Conference on Thermal Treatment of Tissue - Energy Delivery and Assessment IV Bouley, D. M., Daniel, B., Butts Pauly, K., Liu, E., Kinse, A., NAU, W., Diederich, C. J., Sommer, G. SPIE-INT SOC OPTICAL ENGINEERING. 2007

  Abstract

  Magnetic Resonance Imaging (MRI) is a promising tool for visualizing the delivery of minimally invasive cancer treatments such as high intensity ultrasound (HUS) and cryoablation. We use an acute dog prostate model to correlate lesion histopathology with contrast-enhanced (CE) T1 weighted MR images, to aid the radiologists in real time interpretation of in vivo lesion boundaries and pre-existing lesions. Following thermal or cryo treatments, prostate glands are removed, sliced, stained with the vital dye triphenyl tetrazolium chloride, photographed, fixed and processed in oversized blocks for routine microscopy. Slides are scanned by Trestle Corporation at .32 microns/pixel resolution, the various lesions traced using annotation software, and digital images compared to CE MR images. Histologically, HUS results in discrete lesions characterized by a "heat-fixed" zone, in which glands subjected to the highest temperatures are minimally altered, surrounded by a rim or "transition zone" composed of severely fragmented, necrotic glands, interstitial edema and vascular congestion. The "heat-fixed" zone is non-enhancing on CE MRI while the "transition zone" appears as a bright, enhancing rim. Likewise, the CE MR images for cryo lesions appear similar to thermally induced lesions, yet the histopathology is significantly different. Glands subjected to prolonged freezing appear totally disrupted, coagulated and hemorrhagic, while less intensely frozen glands along the lesion edge are partially fragmented and contain apoptotic cells. In conclusion, thermal and cryo-induced lesions, as well as certain pre-existing lesions (cystic hyperplasia - non-enhancing, chronic prostatitis - enhancing) have particular MRI profiles, useful for treatment and diagnostic purposes.

  View details for DOI 10.1117/12.701049

  View details for Web of Science ID 000246494100005

  View details for PubMedCentralID PMC4112763

• Prostate thermal therapy with catheter-based ultrasound devices and MR thermal monitoring Conference on Thermal Treatment of Tissue - Energy Delivery and Assessment IV Diederich, C. J., Nau, W. H., Kinsey, A., Ross, T., Wootton, J., Juang, T., Butts-Pauly, K., Ricke, V., Liu, E. H., Chen, J., Bouley, D. M., van den Bosch, M., Sommer, G. SPIE-INT SOC OPTICAL ENGINEERING. 2007

  View details for DOI 10.1117/12.703184

  View details for Web of Science ID 000246494100009

• Breakpoints in immunoregulation required for Th1 cells to induce diabetes EUROPEAN JOURNAL OF IMMUNOLOGY Neighbors, M., Hartley, S. B., Xu, X., Castro, A. G., Bouley, D. M., O'Garra, A. 2006; 36 (9): 2315-2323

  Abstract

  We describe a novel TCR-transgenic mouse line, TCR7, where MHC class II-restricted, CD4+ T cells are specific for the subdominant H-2b epitope (HEL74-88) of hen egg lysozyme (HEL), and displayed an increased frequency in the thymus and in peripheral lymphoid compartments over that seen in non-transgenic littermate controls. CD4+ T cells responded vigorously to HEL or HEL74-88 epitope presented on APC and could develop into Th1 or Th2 cells under appropriate conditions. Adoptive transfer of TCR7 Ly5.1 T cells into Ly5.2 rat insulin promoter (RIP)-HEL transgenic recipient hosts did not lead to expansion of these cells or result in islet infiltration, although these TCR7 cells could expand upon transfer into mice expressing high levels of HEL in the serum. Islet cell infiltration only occurred when the TCR7 cells had been polarized to either a Th1 or Th2 phenotype prior to transfer, which led to insulitis. Progression from insulitis to autoimmune diabetes only occurred in these recipients when Th1 but not Th2 TCR7 cells were transferred and CTLA-4 signaling was simultaneously blocked. These findings show that regulatory pathways such as CTLA-4 can hold in check already differentiated autoreactive effector Th1 cells, to inhibit the transition from tolerance to autoimmune diabetes.

  View details for DOI 10.1002/eji.200636432

  View details for Web of Science ID 000240586100006

  View details for PubMedID 16933361

• Magnetic resonance-guided high-intensity ultrasound ablation of the prostate. Topics in magnetic resonance imaging Pauly, K. B., Diederich, C. J., Rieke, V., Bouley, D., Chen, J., Nau, W. H., Ross, A. B., Kinsey, A. M., Sommer, G. 2006; 17 (3): 195-207

  Abstract

  This paper describes our work in developing techniques and devices for magnetic resonance (MR)-guided high-intensity ultrasound ablation of the prostate and includes review of relevant literature.Catheter-based high-intensity ultrasound applicators, in interstitial and transurethral configurations, were developed to be used under MR guidance. Magnetic resonance thermometry and the relevant characteristics and artifacts were evaluated during in vivo thermal ablation of the prostate in 10 animals. Contrast-enhanced MR imaging (MRI) and diffusion-weighted MRI were used to assess tissue damage and compared with histology.During evaluation of these applicators, MR thermometry was used to monitor the temperature distributions in the prostate in real time. Magnetic resonance-derived maximum temperature thresholds of 52 degrees C and thermal dose thresholds of 240 minutes were used to control the extent of treatment and qualitatively correlated well with posttreatment imaging studies and histology. The directional transurethral devices are selective in their ability to target well-defined regions of the prostate gland and can be rotated in discrete steps to conform treatment to prescribed boundaries. The curvilinear applicator is the most precise of these directional techniques. Multisectored transurethral applicators, with dynamic angular control of heating and no rotation requirements, offer a fast and less complex means of treatment with less selective contouring.The catheter-based ultrasound devices can produce spatially selective regions of thermal destruction in prostate. The MR thermal imaging and thermal dose maps, obtained in multiple slices through the target volume, are useful for controlling therapy delivery (rotation, power levels, duration). Contrast-enhanced T1-weighted MRI and diffusion-weighted imaging are useful tools for assessing treatment.

  View details for PubMedID 17414077

• Characterization of a novel anthropomorphic plastinated lung phantom 48th Annual Meeting of the American-Association-of-Physicists-in-Medicine Yoon, S., Henry, R., Bouley, D., Fahrig, R. AMER ASSOC PHYSICISTS MEDICINE AMER INST PHYSICS. 2006: 1998–98

  View details for Web of Science ID 000238688500107

• Spontaneous murine neuroaxonal dystrophy: a model of infantile neuroaxonal dystrophy JOURNAL OF COMPARATIVE PATHOLOGY Bouley, D. M., McIntire, J. J., Harris, B. T., Tolwani, R. J., Otto, G. M., DeKruyff, R. H., Hayflick, S. J. 2006; 134 (2-3): 161-170

  Abstract

  The neuroaxonal dystrophies (NADs) in human beings are fatal, inherited, neurodegenerative diseases with distinctive pathological features. This report describes a new mouse model of NAD that was identified as a spontaneous mutation in a BALB/c congenic mouse strain. The affected animals developed clinical signs of a sensory axonopathy consisting of hindlimb spasticity and ataxia as early as 3 weeks of age, with progression to paraparesis and severe morbidity by 6 months of age. Hallmark histological lesions consisted of spheroids (swollen axons), in the grey and white matter of the midbrain, brain stem, and all levels of the spinal cord. Ultrastructural analysis of the spheroids revealed accumulations of layered stacks of membranes and tubulovesicular elements, strongly resembling the ultrastructural changes seen in the axons of human patients with endogenous forms of NAD. Mouse NAD would therefore seem a potentially valuable model of human NADs.

  View details for DOI 10.1016/j.jcpa.2005.10.002

  View details for Web of Science ID 000237336100006

  View details for PubMedID 16542671

  View details for PubMedCentralID PMC2099456

• Assessment of MR thermometry during high intensity ultrasound ablation of the canine prostate 5th International Symposium on Therapeutic Ultrasound Pauly, K. B., Rieke, V., Pisani, L., Sommer, G., Bouley, D., Diederich, C., Ross, A., Nau, W., Kinsey, A., Dumoulin, C., Watkins, R. AMER INST PHYSICS. 2006: 76–80

  View details for Web of Science ID 000238329700015

• The role of antigenic drive and tumor-infiltrating accessory cells in the pathogenesis of Helicobacter-induced mucosa-associated lymphoid tissue lymphoma AMERICAN JOURNAL OF PATHOLOGY Mueller, A., O'Rourke, J., Chu, P., Chu, A., Dixon, M. F., Bouley, D. M., Lee, A., FALKOW, S. 2005; 167 (3): 797-812

  Abstract

  Gastric B-cell lymphoma of mucosa-associated lymphoid tissue type is closely linked to chronic Helicobacter pylori infection. Most clinical and histopathological features of the tumor can be reproduced by prolonged Helicobacter infection of BALB/c mice. In this study, we have addressed the role of antigenic stimulation in the pathogenesis of the lymphoma by experimental infection with Helicobacter felis, followed by antibiotic eradication therapy and subsequent re-infection. Antimicrobial therapy was successful in 75% of mice and led to complete histological but not "molecular" tumor remission. Although lympho-epithelial lesions disappeared and most gastric lymphoid aggregates resolved, transcriptional profiling revealed the long-term mucosal persistence of residual B cells. Experimental re-introduction of Helicobacter led to very rapid recurrence of the lymphomas, which differed from the original lesions by higher proliferative indices and more aggressive behavior. Immunophenotyping of tumor cells revealed massive infiltration of lesions by CD4(+) T cells, which express CD 28, CD 69, and interleukin-4 but not interferon-gamma, suggesting that tumor B-cell proliferation was driven by Th 2-polarized, immunocompetent, and activated T cells. Tumors were also densely colonized by follicular dendritic cells, whose numbers were closely associated with and predictive of treatment outcome.

  View details for Web of Science ID 000231514500015

  View details for PubMedID 16127158

• Curvilinear transurethral ultrasound applicator for selective prostate thermal therapy MEDICAL PHYSICS Ross, A. B., Diederich, C. J., Nau, W. H., Rieke, V., Butts, R. K., Sommer, G., Gill, H., Bouley, D. M. 2005; 32 (6): 1555-1565

  Abstract

  Thermal therapy offers a minimally invasive option for treating benign prostatic hyperplasia (BPH) and localized prostate cancer. In this study we investigated a transurethral ultrasound applicator design utilizing curvilinear, or slightly focused, transducers to heat prostatic tissue rapidly and controllably. The applicator was constructed with two independently powered transducer segments operating at 6.5 MHz and measuring 3.5 mm x 10 mm with a 15 mm radius of curvature across the short axis. The curvilinear applicator was characterized by acoustic efficiency measurements, acoustic beam plots, biothermal simulations of human prostate, ex vivo heating trials in bovine liver, and in vivo heating trials in canine prostate (n=3). Each transducer segment was found to emit a narrow acoustic beam (max width <3 mm), which extended the length of the transducer, with deeper penetration than previously developed planar or sectored tubular transurethral ultrasound applicators. Acoustic and biothermal simulations of human prostate demonstrated three treatment schemes for the curvilinear applicator: single shot (10 W, 60 s) schemes to generate narrow ablation zones (13 x 4 mm, 52 degrees C at the lesion boundary), incremental rotation (10 W, 10 degrees/45 s) to generate larger sector-shaped ablation zones (16 mm x 180 degrees sector), and rotation with variable sonication times (10 W, 10 degrees/15-90 s) to conform the ablation zone to a predefined boundary (9-17 mm x 180 degrees sector, 13 min total treatment time). During in vivo canine prostate experiments, guided by MR temperature imaging, single shot sonications (6 W/transducer, 2-3 min) with the curvilinear applicator ablated 20 degree sections of tissue to the prostate boundary (9-15 mm). Multiple adjacent sonications ("sweeping") ablated large sections of the prostate (180 degrees) by using the MR temperature imaging to adjust the power (4-6.4 W/transducer) and sonication time (30-180 s) at each 10 degrees rotation such that the periphery of the prostate reached 52 degrees C before the next rotation. The conclusion of this study was that the curvilinear applicator produces a narrow and penetrating ultrasound beam that, when combined with image guidance, can provide a precise technique for ablating target regions with a contoured outer boundary, such as the prostate capsule, by rotating in small steps while dynamically adjusting the net applied electrical power and sonication time at each position.

  View details for DOI 10.1118/1.1924314

  View details for Web of Science ID 000229908600015

  View details for PubMedID 16013714

• An MRI-compatible semiautomated vacuum assisted breast biopsy system: Initial feasibility study JOURNAL OF MAGNETIC RESONANCE IMAGING Daniel, B. L., Freeman, L. J., Pyzoha, J. M., McCoy, T. D., Birdwell, R. L., Bouley, D. M., Movius, B., Hibner, J. A. 2005; 21 (5): 637-644

  Abstract

  To develop an MR-compatible vacuum-assisted core needle breast biopsy system.A vacuum-assisted breast biopsy system (Mammotome Hand Held; Ethicon Endo-Surgery, USA) was modified for freehand MRI-guided biopsy in an open, interventional 0.5-T scanner (Signa SP; GE, USA). Probes (11 gauge [G]) were fabricated without significant susceptibility artifact. These mate with an electromechanical hand piece and control system that were modified for use within the MRI scanner. A total of 62 breast lesions were simulated in the mammary tissues of six recently lactating sows by injecting between 0.1 and 1.0 mL of an aqueous gel containing dilute gadopentetate dimeglumine (Gd-DTPA) that formed a bright mass on T1-weighted imaging.Mechanical performance was satisfactory. Magnetic susceptibility and radiofrequency (RF) artifacts from the 11-G probe were negligible. T1-weighted fast spin echo (FSE) was used to guide biopsy. Up to eight samples were removed per lesion. Overall, 461 samples were obtained in 493 attempts (94%). Sample weights averaged 54 mg (N = 493) compared to 4.6 mg (N = 24) from 14-G titanium MRI-compatible needles. On average, 59% of the attempted samples yielded target lesion material.Preliminary results demonstrate the feasibility of a vacuum-assisted breast biopsy system in the MRI environment. Small 0.1-mL lesions can be biopsied without needle artifacts obscuring the target.

  View details for Web of Science ID 000228653600018

  View details for PubMedID 15834914

• MRI-guided interstitial ultrasound thermal therapy of the prostate: A feasibility study in the canine model MEDICAL PHYSICS Nau, W. H., Diederich, C. J., Ross, A. B., Butts, K., Rieke, V., Bouley, D. M., Gill, H., Daniel, B., Sommer, G. 2005; 32 (3): 733-743

  Abstract

  The feasibility of MRI-guided interstitial ultrasound thermal therapy of the prostate was evaluated in an in vivo canine prostate model. MRI compatible, multielement interstitial ultrasound applicators were developed using 1.5 mm diameter cylindrical piezoceramic transducers (7 to 8 MHz) sectored to provide 180 degrees of angular directional heating. Two in vivo experiments were performed in canine prostate. The first using two interstitial ultrasound applicators, the second using three ultrasound applicators in conjunction with rectal and urethral cooling. In both experiments, the applicators were inserted transperineally into the prostate with the energy directed ventrally, away from the rectum. Electrical power levels of 5-17 W per element (approximately 1.6-5.4 W acoustic output power) were applied for heating periods of 18 and 48 min. Phase-sensitive gradient-echo MR imaging was used to monitor the thermal treatment in real-time on a 0.5 T interventional MRI system. Contrast-enhanced T1-weighted images and vital-stained serial tissue sections were obtained to assess thermal damage and correlate to real-time thermal contour plots and calculated thermal doses. Results from these studies indicated a large volume of ablated (nonstained) tissue within the prostate, extending 1.2 to 2.0 cm from the applicators to the periphery of the gland, with the dorsal margin of coagulation well-defined by the applicator placement and directionality. The shape of the lesions correlated well to the hypointense regions visible in the contrast-enhanced T1-weighted images, and were also in good agreement with the contours of the 52 degrees C threshold temperature and t43 > 240 min. This study demonstrates the feasibility of using directional interstitial ultrasound in conjunction with MRI thermal imaging to monitor and possibly control thermal coagulation within a targeted tissue volume while potentially protecting surrounding tissue, such as rectum, from thermal damage.

  View details for DOI 10.1118/1.1861163

  View details for Web of Science ID 000227910600010

  View details for PubMedID 15839345

• Biothermal modeling of transurethral ultrasound applicators for MR-guided prostate thermal therapy Conference on Treatment of Tissue: Energy Delivery and Assessment III Ross, A. B., Diederich, C. J., Nau, W. H., Tyreus, D., Gill, H., Bouley, D., Butts, R. K., Rieke, V., Daniel, B., Sommer, G. SPIE-INT SOC OPTICAL ENGINEERING. 2005: 220–227

  View details for DOI 10.1117/12.592491

  View details for Web of Science ID 000229738000020

• Magnetic resonance imaging and surgical repair of cleft palate in a four-week-old canine (Canis familiaris): An animal model for cleft palate repair CONTEMPORARY TOPICS IN LABORATORY ANIMAL SCIENCE Tolwani, R. J., Hagan, C. E., Runstadler, J. A., Lyons, H., Green, V. L., Bouley, D. M., Rodriguez, L. F., Schendel, S. A., Moseley, M. E., Daunt, D. A., Otto, G., CORK, L. C. 2004; 43 (6): 17-21

  Abstract

  Successful cleft palate repair (palatoplasty) was accomplished in a male canine pup from a kindred with autosomal recessive transmission for a complete cleft palate phenotype. This case represents the potential application of a new animal model for cleft palate repair. This reproducible congenital defect provides a clinically relevant model to improve research into the human anomaly, as compared with previous iatrogenic or teratogenically induced animal models. This case report presents the basis for new repair techniques and for studying the genetic basis of the cleft palate defect.

  View details for Web of Science ID 000225806200004

  View details for PubMedID 15636550

• Neonatal lethality of LGR5 null mice is associated with ankyloglossia and gastrointestinal distension MOLECULAR AND CELLULAR BIOLOGY Morita, H., Mazerbourg, S., Bouley, D. M., Luo, C. W., Kawamura, K., Kuwabara, Y., Baribault, H., Tian, H., Hsueh, A. J. 2004; 24 (22): 9736-9743

  Abstract

  The physiological role of an orphan G protein-coupled receptor, LGR5, was investigated by targeted deletion of this seven-transmembrane protein containing a large N-terminal extracellular domain with leucine-rich repeats. LGR5 null mice exhibited 100% neonatal lethality characterized by gastrointestinal tract dilation with air and an absence of milk in the stomach. Gross and histological examination revealed fusion of the tongue to the floor of oral cavity in the mutant newborns and immunostaining of LGR5 expression in the epithelium of the tongue and in the mandible of the wild-type embryos. The observed ankyloglossia phenotype provides a model for understanding the genetic basis of this craniofacial defect in humans and an opportunity to elucidate the physiological role of the LGR5 signaling system during embryonic development.

  View details for DOI 10.1128/MCB.24.22.9736-9743.2004

  View details for Web of Science ID 000224823300004

  View details for PubMedID 15509778

  View details for PubMedCentralID PMC525477

• Outbreak of Mycobacterium bovis in a conditioned colony of rhesus (Macaca mulatta) and cynomolgus (Macaca fascicularis) Macaques COMPARATIVE MEDICINE Garcia, M. A., Bouley, D. M., Larson, M. J., Lifland, B., Moorhead, R., Simkins, M. D., Borie, D. C., Tolwani, R., Otto, G. 2004; 54 (5): 578-584

  Abstract

  We describe a tuberculosis outbreak caused by Mycobacterium bovis in a conditioned colony of rhesus (Macaca mulatta) and cynomolgus (Macaca fascicularis) macaques. Animals in five rooms were exposed, but most (16/27) infections were confined to the room that housed a mixed population of cynomolgus and rhesus macaques. In this room, rhesus (8/8) and cynomolgus (10/11) macaques naturally exposed to M. bovis were infected at nearly identical rates (Fisher exact test, 2-tailed P = 1). The clinical signs of disease and pathologic lesions in infected macaques, however, were moderately different between the two species. Rhesus macaques were more likely (5/8) to exhibit clinical signs of persistent coughing and inappetance, and had more severe pulmonary lesions. By contrast, clinical signs of disease were seen in only 1 of 19 cynomolgus macaques, and overall, the pulmonary lesions were often focal and less severe, although some still had severe involvement of the lungs similar to that seen in rhesus macaques. These differences should be taken into consideration when developing or evaluating a tuberculosis-screening program. On the basis of observations made during this outbreak, we recommend that alternative screening methods such as the PRIMAGAM test and the ESAT-6 ELISA, be incorporated into the screening program to aid in the identification of infected animals.

  View details for Web of Science ID 000225357200015

  View details for PubMedID 15575373

• Leucine-rich repeat-containing, G protein-coupled receptor 4 null mice exhibit intrauterine growth retardation associated with embryonic and perinatal lethality MOLECULAR ENDOCRINOLOGY Mazerbourg, S., Bouley, D. M., Sudo, S., Klein, C. A., Zhang, J. V., Kawamura, K., Goodrich, L. V., Rayburn, H., Tessier-Lavigne, M., Hsueh, A. J. 2004; 18 (9): 2241-2254

  Abstract

  Leucine-rich repeat-containing, G protein-coupled receptors (LGRs) belong to the largest mammalian superfamily of proteins with seven-transmembrane domains. LGRs can be divided into three subgroups based on their unique domain arrangement. Although two subgroups have been found to be receptors for glycoprotein hormones and relaxin-related ligands, respectively, the third LGR subgroup, consisting of LGR4-6, are orphan receptors with unknown physiological roles. To elucidate the functions of this subgroup of LGRs, LGR4 null mice were generated using a secretory trap approach to delete the majority of the LGR4 gene after the insertion of a beta-galactosidase reporter gene immediately after exon 1. Tissues expressing LGR4 were analyzed based on histochemical staining of the transgene driven by the endogenous LGR4 promoter. LGR4 was widely expressed in kidney, adrenal gland, stomach, intestine, heart, bone/cartilage, and other tissues. The expression of LGR4 in these tissues was further confirmed by immunohistochemical studies in wild-type animals. Analysis of the viability of 250 newborn animals suggested a skewed inheritance pattern, indicating that only 40% of the expected LGR4 null mice were born. For the LGR4 null mice viable at birth, most of them died within 2 d. Furthermore, the LGR4 null mice showed intrauterine growth retardation as reflected by a 14% decrease in body weight at birth, together with 30% and 40% decreases in kidney and liver weights, respectively. The present findings demonstrate the widespread expression of LGR4, and an essential role of LGR4 for embryonic growth, as well as kidney and liver development. The observed pre- and postnatal lethality of LGR4 null mice illustrates the importance of the LGR4 signaling system for the survival and growth of animals during the perinatal stage.

  View details for DOI 10.1210/me.2004-0133

  View details for PubMedID 15192078

• RabGEF1 is a negative regulator of mast cell activation and skin inflammation NATURE IMMUNOLOGY Tam, S. Y., Tsai, M., Snouwaert, J. N., Kalesnikoff, J., Scherrer, D., Nakae, S., Chatterjea, D., Bouley, D. M., Galli, S. J. 2004; 5 (8): 844-852

  Abstract

  Mast cell activation induced by aggregation of Fc epsilon RI receptors with immunoglobulin E and antigen is mediated through the activation of multiple protein kinase cascades. Here we report that the regulatory protein RabGEF1 bound to Ras and negatively regulated Ras activation and its 'downstream' effector pathways in Fc epsilon RI-dependent mast cell activation. RabGEF1-deficient mast cells showed enhanced degranulation and release of lipid mediators and cytokines in response to Fc epsilon RI aggregation. RabGEF1-deficient mice developed severe skin inflammation and had increased numbers of mast cells. Thus, RabGEF1 is a negative regulator of Fc epsilon RI-dependent mast cell activation, and a lack of RabGEF1 results in the development of skin inflammation in vivo.

  View details for DOI 10.1038/ni1093

  View details for Web of Science ID 000222955600016

  View details for PubMedID 15235600

• Topical treatment with inhibitors of the phosphatidylinositol 3 '-kinase/akt and raf/mitogen-activated protein kinase kinase/extracellular signal-regulated kinase pathways reduces melanoma development in severe combined immunodericient mice CANCER RESEARCH Bedogni, B., O'Neill, M. S., Welford, S. M., Bouley, D. M., Giaccia, A. J., Denko, N. C., Powell, M. B. 2004; 64 (7): 2552-2560

  Abstract

  Topical treatment with inhibitors of the phosphatidylinositol 3'-kinase/Akt and Raf/mitogen-activated protein kinase kinase/extracellular signal-regulated kinase pathways inhibited the growth of TPras transgenic melanomas in severe combined immunodeficient mice, blocked invasive behavior, and reduced angiogenesis. The inhibitor Ly294002, which is specific for phosphatidylinositol 3'-kinase, effectively reduced melanoma cell growth both in vitro and in vivo. Both Ly294002 and U0126, a mitogen-activated protein kinase kinase 1/2 inhibitor, reduced invasion, which correlated with reduction of the metalloproteinase matrix metalloproteinase 2. Tumor angiogenesis was disrupted through inhibition of vascular endothelial growth factor production from the tumor cells and antiangiogenic effects on endothelial cells. Observations with TPras melanoma cells that express dominant negative Deltap85 or kinase-inactive Raf(301) supported the specificity of the phenomena observed with the chemical inhibitors. These studies demonstrate that topical treatment targeting Ras effectors is efficacious, without systemic toxicities, and may prove to be useful in treating and preventing the progression of cutaneous melanoma.

  View details for Web of Science ID 000220586500035

  View details for PubMedID 15059911

• The antinociceptive effect of transcranial electrostimulation with combined direct and alternating current in freely moving rats ANESTHESIA AND ANALGESIA Nekhendzy, V., Fender, C. P., Davies, M. F., Lemmens, H. J., Kim, M. S., Bouley, D. M., Maze, M. 2004; 98 (3): 730-737

  Abstract

  Transcranial electrostimulation (TES) has been reported to elicit significant analgesia, allowing a substantial reduction of intraoperative opioids. Acceptance of TES into clinical practice is hampered by lack of controlled clinical trials and inconclusive animal data regarding the TES antinociceptive action. This inconclusive data may be explained, in part, by failure in rat experiments to simulate the variables used in humans when TES electrodes are positioned on the skin. In this study we validated the TES antinociceptive effect in a novel animal model of cutaneously administered TES, when the stimulating conditions mimic the ones used in clinical practice. The antinociceptive effect was assessed by measuring nociceptive thresholds in the tail-flick and hot-plate latency tests in awake, unrestrained male rats. Data were analyzed by analysis of variance and mixed-effects population modeling. The administration of TES at 2.25 mA produced an almost immediate, sustained, frequency-dependent (40-60 Hz) antinociceptive effect, reaching approximately 50% of the maximal possible value. We conclude that an antinociceptive effect of cutaneously administered TES can be demonstrated in the rat. Some characteristics of the effect suggest an important role of the sensory nerves of the rat's scalp in mediating the TES antinociceptive response.Transcranial electrostimulation produces a significant, frequency-dependent antinociceptive effect that may be mediated by cutaneous nerves of the scalp.

  View details for DOI 10.1213/01.ANE.0000096007.12845.70

  View details for PubMedID 14980928

• Diagnosis of tuberculosis in macaques, using whole-blood in vitro interferon-gamma (PRIMAGAM) testing COMPARATIVE MEDICINE Garcia, M. A., Yee, J., Bouley, D. M., Moorhead, R., LERCHE, N. W. 2004; 54 (1): 86-92

  Abstract

  During the fall of 2001, a tuberculosis outbreak caused by Mycobacterium bovis occurred in a conditioned colony of rhesus (Macaca mulatta) and cynomolgus (Macaca fascicularis) macaques at Stanford University School of Medicine. During this outbreak, we evaluated the diagnostic performance of a new in vitro tuberculosis screening test (PRIMAGAM). The PRIMAGAM test measures the interferon-gamma (IFNgamma) response to purified protein derivatives (PPDs) of M. bovis and M. avium. On the basis of the results of the last test administered before necropsy, the PRIMAGAM test had good sensitivity (68%) and excellent specificity (97%), compared with the disease status, as determined by the presence or absence of gross and/or histologic lesions indicative of tuberculosis. By contrast, sensitivity and specificity of the tuberculin skin test (TST) was 84 and 87%, respectively. Both tests suffered from intermittent positive and negative reactions on repeat testing. Overall, however, there was no significant difference (P = 0.09, McNemar's chi2-test) and moderate agreement (kappa = 0.52) between these two tests. Lastly, the IFNgamma response to bovine PPD was significantly lower in infected cynomolgus macaques. Moreover, each test failed to detect tuberculosis in three cynomolgus macaques. Fortunately, they were different animals; therefore, we recommend the parallel use of the TST and PRIMAGAM test for maximal overall sensitivity in a tuberculosis screening program, especially for cynomolgus macaques.

  View details for Web of Science ID 000189358000015

  View details for PubMedID 15027623

• Highly directional transurethral ultrasound applicators with rotational control for MRI-guided prostatic thermal therapy PHYSICS IN MEDICINE AND BIOLOGY Ross, A. B., Diederich, C. J., Nau, W. H., Gill, H., Bouley, D. M., Daniel, B., Rieke, V., Butts, R. K., Sommer, G. 2004; 49 (2): 189-204

  Abstract

  Transurethral ultrasound applicators with highly directional energy deposition and rotational control were investigated for precise treatment of benign prostatic hyperplasia (BPH) and adenocarcinoma of the prostate (CaP). Two types of catheter-based applicators were fabricated, using either 90 degrees sectored tubular (3.5 mm OD x 10 mm) or planar transducers (3.5 mm x 10 mm). They were constructed to be MRI compatible, minimally invasive and allow for manual rotation of the transducer array within a 10 mm cooling balloon. In vivo evaluations of the applicators were performed in canine prostates (n = 3) using MRI guidance (0.5 T interventional magnet). MR temperature imaging (MRTI) utilizing the proton resonance frequency shift method was used to acquire multiple-slice temperature overlays in real time for monitoring and guiding the thermal treatments. Post-treatment T1-weighted contrast-enhanced imaging and triphenyl tetrazolium chloride stained tissue sections were used to define regions of tissue coagulation. Single sonications with the 90 degrees tubular applicator (9-15 W, 12 min, 8 MHz) produced coagulated zones covering an 80 degrees wedge of the prostate extending from 1-2 mm outside the urethra to the outer boundary of the gland (16 mm radial coagulation). Single sonications with the planar applicator (15-20 W, 10 min, approximately 8 MHz) generated thermal lesions of approximately 30 degrees extending to the prostate boundary. Multiple sequential sonications (sweeping) of a planar applicator (12 W with eight rotations of 30 degrees each) demonstrated controllable coagulation of a 270 degrees contiguous section of the prostate extending to the capsule boundary. The feasibility of using highly directional transurethral ultrasound applicators with rotational capabilities to selectively coagulate regions of the prostate while monitoring and controlling the treatments with MRTI was demonstrated in this study.

  View details for DOI 10.1088/0031-9155/49/2/0020

  View details for Web of Science ID 000220421700002

  View details for PubMedID 15083666

• Salmonella typhimurium persists within macrophages in the mesenteric lymph nodes of chronically infected Nramp1(+/+) mice and can be reactivated by IFN gamma neutralization JOURNAL OF EXPERIMENTAL MEDICINE Monack, D. M., Bouley, D. M., FALKOW, S. 2004; 199 (2): 231-241

  Abstract

  Host-adapted strains of Salmonella are capable of establishing a persistent infection in their host often in the absence of clinical disease. The mouse model of Salmonella infection has primarily been used as a model for the acute systemic disease. Therefore, the sites of long-term S. typhimurium persistence in the mouse are not known nor are the mechanisms of persistent infection clearly understood. Here, we show that S. typhimurium can persist for as long as 1 yr in the mesenteric lymph nodes (MLNs) of 129sv Nramp1(+)(/)(+) (Slc11a1(+)(/)(+)) mice despite the presence of high levels of anti-S. typhimurium antibody. Tissues from 129sv mice colonized for 60 d contain numerous inflammatory foci and lesions with features resembling S. typhi granulomas. Tissues from mice infected for 365 d have very few organized inflammatory lesions, but the bacteria continue to persist within macrophages in the MLN and the animals generally remain disease-free. Finally, chronically infected mice treated with an interferon-gamma neutralizing antibody exhibited symptoms of acute systemic infection, with evidence of high levels of bacterial replication in most tissues and high levels of fecal shedding. Thus, interferon-gamma, which may affect the level of macrophage activation, plays an essential role in the control of the persistent S. typhimurium infection in mice.

  View details for DOI 10.1084/jem.20031319

  View details for Web of Science ID 000188369700009

  View details for PubMedID 14734525

  View details for PubMedCentralID PMC2211772

• Detection of early secretory antigenic target-6 antibody for diagnosis of tuberculosis in non-human primates COMPARATIVE MEDICINE Kanaujia, G. V., Garcia, M. A., Bouley, D. M., Peters, R., Gennaro, M. L. 2003; 53 (6): 602-606

  Abstract

  Tuberculosis is one of the most economically devastating, zoonotic infections of captive non-human primates. The limitations of the tuberculin skin test, which is currently used to diagnose tuberculosis in living non-human primates, make it necessary to find new, simple, and economical diagnostic methods. We describe use of an enzyme-linked immunoassay to detect IgG antibodies against early secretory antigenic target (ESAT)-6, a small protein secreted by virulent tubercle bacilli, in paired (pre- and post-outbreak) sera from 57 non-human primates involved in an outbreak of Mycobacterium bovis infection in a research colony. Of 25 animals with tuberculosis lesions at necropsy, 22 (88%) had high serum levels of the ESAT-6 antibody. The ESAT-6 antibody was found in 16% (5/32) of post-outbreak sera from animals in which tuberculosis could not be confirmed at necropsy. The strong association between the ESAT-6 antibody and tuberculosis in non-human primates documented in this study, together with the robustness of the serologic assay, make the ESAT-6 ELISA a valuable tool for diagnosis of tuberculosis in captive non-human primates.

  View details for Web of Science ID 000188057800003

  View details for PubMedID 14727807

• Inhibition of delta-protein kinase C protects against reperfusion injury of the ischemic heart in vivo CIRCULATION Inagaki, K., Chen, L., Ikeno, F., Lee, F. H., Imahashi, K., Bouley, D. M., Rezaee, M., Yock, P. G., Murphy, E., Mochly-Rosen, D. 2003; 108 (19): 2304-2307

  Abstract

  Current treatment for acute myocardial infarction (AMI) focuses on reestablishing blood flow (reperfusion). Paradoxically, reperfusion itself may cause additional injury to the heart. We previously found that delta-protein kinase C (deltaPKC) inhibition during simulated ischemia/reperfusion in isolated rat hearts is cardioprotective. We focus here on the role for deltaPKC during reperfusion only, using an in vivo porcine model of AMI.An intracoronary application of a selective deltaPKC inhibitor to the heart at the time of reperfusion reduced infarct size, improved cardiac function, inhibited troponin T release, and reduced apoptosis. Using 31P NMR in isolated perfused mouse hearts, we found a faster recovery of ATP levels in hearts treated with the deltaPKC inhibitor during reperfusion only.Reperfusion injury after cardiac ischemia is mediated, at least in part, by deltaPKC activation. This study suggests that including a deltaPKC inhibitor at reperfusion may improve the outcome for patients with AMI.

  View details for DOI 10.1161/01.CIR.0000101682.24138.36

  View details for PubMedID 14597593

• Thermal shock in a colony of South African clawed frogs (Xenopus laevis) VETERINARY RECORD GREEN, S. L., Moorhead, R. C., Bouley, D. M. 2003; 152 (11): 336-?

  View details for Web of Science ID 000182199900015

  View details for PubMedID 12665149

• Cryptosporidiosis associated with emaciation and proliferative gastritis in a laboratory-reared South African clawed frog (Xenopus laevis) COMPARATIVE MEDICINE GREEN, S. L., Bouley, D. M., Josling, C. A., Fayer, R. 2003; 53 (1): 81-84

  Abstract

  A 2-year-old emaciated female South African clawed frog (Xenopus laevis) was euthanized because of chronic weight loss. At necropsy, there was no evidence of bacterial, fungal or viral disease; however, the histopathologic findings indicated a proliferative gastritis and the presence of numerous cryptosporidial stages throughout the intestinal tract. Crytosporidial oocysts were present in the water taken from the aquarium housing the infected frog and were likely shed by the sick frog; however, the exact source of the oocysts could not be identified. Water samples from other frog aquaria in the facility did not contain cryptosporidial oocysts. Some Cryptosporidium species are important zoonotic pathogens and, to our knowledge, this is the first report of disease associated with Cryptosporidium infection in a laboratory Xenopus laevis.

  View details for Web of Science ID 000181240800011

  View details for PubMedID 12625511

• Regional vs systemic antivenom administration in the treatment of snake venom poisoning in a rabbit model: A pilot study Research Forum of the American-College-of-Emergency-Physicians Norris, R. L., Dery, R., Johnson, C., Williams, S., Rose, K., Young, L., McDougal, L. R., Bouley, D., Oehlert, J., Thompson, R. C. ALLIANCE COMMUNICATIONS GROUP DIVISION ALLEN PRESS. 2003: 231–35

  Abstract

  To develop a model that compares 2 different routes of antivenom administration (standard intravenous [IV] administration vs regional administration below a tourniquet) to assess their ability to limit muscle necrosis in a rabbit model of rattlesnake venom poisoning.New Zealand white rabbits were randomly assigned to 4 groups. All animals underwent general anesthesia and were then injected intramuscularly (IM) with a sublethal dose of western diamond-back rattlesnake (Crotalus atrox) venom in the right thigh and a similar volume of normal saline (NS) control in the left thigh. Thirty minutes later, standard treatment group animals (n = 4) received 1 vial of reconstituted Antivenin (Crotalidae) Polyvalent (ACP) and 10 mL of NS through an ear vein. Experimental treatment group animals (n = 4) had their lower extremities exsanguinated and isolated by arterial tourniquets. One vial of ACP was then given through a distal IV in the envenomed extremity, and 10 mL of NS was given through an IV in the sham extremity. Tourniquets were removed 30 minutes later. Positive control group animals (n = 2) similarly had their lower extremities exsanguinated and isolated by tourniquets. They then received 10 mL of NS through distal IVs in each lower extremity. Tourniquets were again removed after 30 minutes. Negative control group animals (n = 2) received 2 doses of NS only (10 mL each) through an ear vein. Serum creatinine phosphokinase (CPK) levels were drawn at baseline and 48 hours following venom injection. At 48 hours, the animals were injected with technetium pyrophosphate. Two hours later, they were euthanized, and the lower extremities were scanned to determine levels of radionucleotide uptake in envenomed muscles compared to contralateral sham-injected muscles. The anterior thigh muscle groups were then removed, fixed, stained, sectioned, and analyzed in a blinded fashion by a veterinary pathologist for muscle necrosis grading.There was no evidence of statistically significant differences in changes in serum CPK levels (from baseline to 48 hours), technetium pyrophosphate uptake ratios (right leg/left leg), or muscle necrosis indices in any 2-group analysis.Results of this pilot study do not suggest any beneficial effect of ACP, in the dose and routes used, in limiting local muscle necrosis following IM rattlesnake venom poisoning in the rabbit model.

  View details for PubMedID 14719857

• Diffusion-weighted MRI after cryosurgery of the canine prostate JOURNAL OF MAGNETIC RESONANCE IMAGING Butts, K., Daniel, B. L., Chen, L. L., Bouley, D. M., Wansapura, J., Maier, S. E., Dumoulin, C., Watkins, R. 2003; 17 (1): 131-135

  Abstract

  To evaluate the acute lesion created by cryosurgery with diffusion-weighted magnetic resonance imaging (DWI).The appearance of the acute cryolesion was evaluated in four canine prostates DWI after they were warmed to original body temperature. The prostates were excised, stained with triphenyl tetrazolium chloride (TTC), photographed, prepared for hematoxylin and eosin (H&E) staining, and examined under a light microscope.A marked decrease in apparent diffusion coefficient of 38% was evident in the center of the previously frozen tissue, but not in all of the previously frozen tissue. Histologic results confirm differences between the iceball core and the periphery of the iceball, which have markedly different imaging characteristics on DWI.The core of the previously frozen tissue has a reduced apparent diffusion coefficient (ADC) compared to the periphery of the previously frozen tissue and previously unfrozen tissue.

  View details for DOI 10.1002/jmri.10227

  View details for Web of Science ID 000182657900014

• MR-guided conformal heating of canine prostate using interstitial applicators Conference on Thermal Treatment of Tissue: Energy Delivery and Assessment II Nau, W. H., Diederich, C. J., Ross, T., Butts, K., Rieke, V., Bouley, D., Gill, H., Daniel, B., Sommer, G. SPIE-INT SOC OPTICAL ENGINEERING. 2003: 220–226

  View details for Web of Science ID 000184241000021

• Magnetic resonance guided directional transurethral ultrasound thermal therapy Conference on Thermal Treatment of Tissue: Energy Delivery and Assessment II Ross, A. B., Diederich, C. J., Nau, W. H., Tyreus, P. D., Gill, H., Bouley, D., Butts, R. K., Rieke, V., Daniel, B., Sommer, G. SPIE-INT SOC OPTICAL ENGINEERING. 2003: 192–199

  View details for Web of Science ID 000184241000018

• Diffusion-weighted MRI after cryosurgery of the canine prostate. Magnetic resonance imaging. Journal of magnetic resonance imaging : JMRI Butts, K., Daniel, B. L., Chen, L., Bouley, D. M., Wansapura, J., Maier, S. E., Dumoulin, C., Watkins, R. 2003; 17 (1): 131-135

  Abstract

  To evaluate the acute lesion created by cryosurgery with diffusion-weighted magnetic resonance imaging (DWI).The appearance of the acute cryolesion was evaluated in four canine prostates DWI after they were warmed to original body temperature. The prostates were excised, stained with triphenyl tetrazolium chloride (TTC), photographed, prepared for hematoxylin and eosin (H&E) staining, and examined under a light microscope.A marked decrease in apparent diffusion coefficient of 38% was evident in the center of the previously frozen tissue, but not in all of the previously frozen tissue. Histologic results confirm differences between the iceball core and the periphery of the iceball, which have markedly different imaging characteristics on DWI.The core of the previously frozen tissue has a reduced apparent diffusion coefficient (ADC) compared to the periphery of the previously frozen tissue and previously unfrozen tissue.

  View details for PubMedID 12500282

• Noninvasive measurement of extraction fraction and single-kidney glomerular filtration rate with MR imaging in swine with surgically created renal artery stenoses RADIOLOGY Coulam, C. H., Lee, J. H., Wedding, K. L., Spielman, D. M., Pelc, N. J., Kee, S. T., Hill, B. B., Bouley, D. M., Derby, G. C., Myers, B. D., Sawyer-Glover, A. M., Sommer, F. G. 2002; 223 (1): 76-82

  Abstract

  To test whether magnetic resonance (MR) imaging enables accurate measurement of extraction fraction (EF) in swine with unilateral renal ischemia and to evaluate effects of renal arterial stenosis on EF and single-kidney glomerular filtration rate.High-grade unilateral renal arterial stenoses were surgically created in eight pigs. Direct measurements of renal venous and arterial inulin concentration provided reference standard estimates of single-kidney EF. Pigs were imaged with a 1.5-T imager to estimate EF, renal blood flow, and glomerular filtration rate. A breath-hold inversion-recovery spiral sequence was used to measure T1 of blood in the infrarenal inferior vena cava and renal veins after intravenous administration of gadopentetate dimeglumine, and these data were used to calculate EF. Cine-phase contrast material-enhanced imaging of the renal arteries provided quantitative renal blood flow measurements. Bilateral single-kidney glomerular filtration rate was then determined: glomerular filtration rate = renal blood flow x (1 - hematocrit level) x EF.A statistically significant linear correlation was found between EF, as determined with MR imaging, and inulin (r = 0.77). As compared with kidneys without renal arterial stenosis, kidneys with renal arterial stenosis showed 50% (0.14/0.28) EF reduction (P <.01) and 59% glomerular filtration rate reduction (P <.01).MR imaging shows promise for in vivo measurement of EF and glomerular filtration rate, which may be useful in assessing the clinical importance of renal arterial stenosis.

  View details for DOI 10.1148/radiol.2231010420

  View details for PubMedID 11930050

• A novel inhibitor of protein kinase-C as a therapeutic for ischemic reperfusion injury in acute coronary syndromes Lee, F., Inagaki, K., Chen, L., Bailey, L., Ikeno, F., Lyons, J., Bouley, D., Yock, P., Carter, A., Yeung, A., Mochly-Rosen, D. ELSEVIER SCIENCE INC. 2002: 445A–445A

  View details for Web of Science ID 000174106702000

• Measurement of renal volumes with contrast-enhanced MRI 9th Annual Meeting of the ISMRM Coulam, C. H., Bouley, D. M., Sommer, F. G. JOHN WILEY & SONS INC. 2002: 174–79

  Abstract

  To determine the accuracy of in vivo magnetic resonance imaging (MRI) measurement of total renal parenchymal volume and medullary fraction. MATERIALS andSixteen kidneys in eight pigs were imaged with a multiphasic contrast-enhanced fast three-dimensional sequence on a 1.5-T imager. Kidney segmentation, followed by a process of signal intensity thresholding for cortical and nephrographic phase datasets, allowed for MRI measurements of parenchymal volume and medullary fraction. Autopsy provided reference standards of renal volume, weight, and medullary fraction.An excellent correlation was found between MRI measurement of total renal parenchymal volume and autopsy volume (R2 = 0.86) and weight (R2 = 0.90). Medullary fraction (mean +/- SD) measured with MRI was 0.120 +/- 0.067, and with autopsy was 0.116 +/- 0.025 (t-test P = 0.84, F-test P = 0.001).MRI measurements of total renal volume are accurate. MRI measurements of medullary fraction show promise, but precision is limited when using a simple signal intensity thresholding algorithm.

  View details for DOI 10.1002/jmri.10058

  View details for Web of Science ID 000174756200008

  View details for PubMedID 11836773

• Dynamic nature of host-pathogen interactions in Mycobacterium marinum granulomas INFECTION AND IMMUNITY Bouley, D. M., Ghori, N., Mercer, K. L., FALKOW, S., Ramakrishnan, L. 2001; 69 (12): 7820-7831

  Abstract

  Mycobacterium marinum causes long-term subclinical granulomatous infection in immunocompetent leopard frogs (Rana pipiens). These granulomas, organized collections of activated macrophages, share many morphological features with persistent human tuberculous infection. We examined organs of frogs with chronic M. marinum infection using transmission electron microscopy in conjunction with immunohistochemistry and acid phosphatase cytochemistry to better define the bacterium-host interplay during persistent infection. Bacteria were always found within macrophage phagosomes. These phagosomes were often fused to lysosomes, in sharp contrast to those formed during in vitro infection of J774 macrophage-like cells by M. marinum. The infected macrophages in frog granulomas showed various levels of activation, as evidenced by morphological changes, including epithelioid transformation, recent phagocytic events, phagolysosomal fusion, and disintegration of bacteria. Our results demonstrate that even long-term granulomas are dynamic environments with regard to the level of host cell activation and bacterial turnover and suggest a continuum between constantly replicating bacteria and phagocytic killing that maintains relatively constant bacterial numbers despite an established immune response. Infection with a mutant bacterial strain with a reduced capacity for intracellular replication shifted the balance, leading to a greatly reduced bacterial burden and inflammatory foci that differed from typical granulomas.

  View details for Web of Science ID 000172297600077

  View details for PubMedID 11705964

• Murine cytomegalovirus CC chemokine homolog MCK-2 (m131-129) is a determinant of dissemination that increases inflammation at initial sites of infection JOURNAL OF VIROLOGY Saederup, N., Aguirre, S. A., Sparer, T. E., Bouley, D. M., Mocarski, E. S. 2001; 75 (20): 9966-9976

  Abstract

  The murine cytomegalovirus CC chemokine homolog MCK-2 (m131-129) is an important determinant of dissemination during primary infection. Reduced peak levels of viremia at day 5 were followed by reduced levels of virus in salivary glands starting at day 7 when mck insertion (RM461) and point (RM4511) mutants were compared to mck-expressing viruses. A dramatic MCK-2-enhanced inflammation occurred at the inoculation site over the first few days of infection, preceding viremia. The data further reinforce the role of MCK-2 as a proinflammatory signal that recruits leukocytes to increase the efficiency of viral dissemination in the host.

  View details for Web of Science ID 000171102900043

  View details for PubMedID 11559829

• Molecular and phenotypic analysis of Attractin mutant mice GENETICS Gunn, T. M., Inui, T., Kitada, K., Ito, S., Wakamatsu, K., He, L., Bouley, D. M., Serikawa, T., Barsh, G. S. 2001; 158 (4): 1683-1695

  Abstract

  Mutations of the mouse Attractin (Atrn; formerly mahogany) gene were originally recognized because they suppress Agouti pigment type switching. More recently, effects independent of Agouti have been recognized: mice homozygous for the Atrn(mg-3J) allele are resistant to diet-induced obesity and also develop abnormal myelination and vacuolation in the central nervous system. To better understand the pathophysiology and relationship of these pleiotropic effects, we further characterized the molecular abnormalities responsible for two additional Atrn alleles, Atrn(mg) and Atrn(mg-L), and examined in parallel the phenotypes of homozygous and compound heterozygous animals. We find that the three alleles have similar effects on pigmentation and neurodegeneration, with a relative severity of Atrn(mg-3J) > Atrn(mg) > Atrn(mg-L), which also corresponds to the effects of the three alleles on levels of normal Atrn mRNA. Animals homozygous for Atrn(mg-3J) or Atrn(mg), but not Atrn(mg-L), show reduced body weight, reduced adiposity, and increased locomotor activity, all in the presence of normal food intake. These results confirm that the mechanism responsible for the neuropathological alteration is a loss--rather than gain--of function, indicate that abnormal body weight in Atrn mutant mice is caused by a central process leading to increased energy expenditure, and demonstrate that pigmentation is more sensitive to levels of Atrn mRNA than are nonpigmentary phenotypes.

  View details for Web of Science ID 000170603700024

  View details for PubMedID 11514456

• Canine motor neuron disease: Clinicopathologic features and selected indicators of oxidative stress 17th Annual Meeting of the American-College-of-Veterinary-Internal-Medicine GREEN, S. L., Bouley, D. M., Pinter, M. J., CORK, L. C., Vatassery, G. T. WILEY-BLACKWELL PUBLISHING, INC. 2001: 112–19

  Abstract

  Hereditary canine spinal muscular atrophy (HCSMA) is an inherited motor neuron disease affecting a kindred of Brittanies. We have examined the clinicopathologic abnormalities in 57 animals with HCSMA, including 43 affected adult dogs and 14 homozygote pups. We also measured selected biochemical indices of oxidative stress: serum vitamin E (alpha-tocopherol) and Se concentrations; serum concentrations of Cu, Zn, Mg, and Fe; and total superoxide dismutase and glutathione peroxidase activities in red blood cells. Dogs with HCSMA had the following abnormalities: regenerative anemia, hypoglobulinemia, hypochloremia, and abnormally high creatine kinase and liver alkaline phosphatase activities. Serum Cu concentration was significantly (P = .01) increased in adult dogs with HCSMA compared to control dogs. Serum vitamin E concentrations tended to be lower in adult dogs with HCSMA compared to controls, and were significantly (P = .01) lower in homozygote pups compared to control pups.

  View details for Web of Science ID 000167403500006

  View details for PubMedID 11300593

• Suspected hypovitaminosis A in a colony of captive green anoles (Anolis carolinensis) CONTEMPORARY TOPICS IN LABORATORY ANIMAL SCIENCE Miller, E. A., GREEN, S. L., Otto, G. M., Bouley, D. M. 2001; 40 (2): 18-20

  Abstract

  In a colony of 18 green anoles (Anolis carolinensis), 3 animals experienced focally thickened lips, ulcerative cheilitis, lethargy, depression, and weight loss over a 5-month period. In addition to crickets fed fresh fruit and leafy green vegetables, the diet of the green anoles consisted of a supply of mealworms that had been dusted with a commercial liquid vitamin supplement. The history, clinical findings, and histopathologic lesions were suggestive of hypovitaminosis A, which is known to cause squamous metaplasia of the mucus secreting glands and epithelial surfaces in many species.

  View details for Web of Science ID 000167815600005

  View details for PubMedID 11300682

• MRI study of immediate cell viability in focused ultrasound lesions in the rabbit brain JOURNAL OF MAGNETIC RESONANCE IMAGING Chen, L., Bouley, D. M., Harris, B. T., Butts, K. 2001; 13 (1): 23-30

  Abstract

  The purpose of this study was to evaluate cell viability in MR imaged focused ultrasound (FUS) lesions using cell-viability staining with triphenyl tetrazolium chloride (TTC) and both light and electron microscopy. Ten paired ultrasonic lesions were created in 5 rabbit brains in vivo with an ultrasound beam of 1.5 MHz electrical power input to the transducer of 50 W and exposure duration of 15 seconds. T2-weighted fast spin-echo (FSE) MRI was performed to detect the FUS lesions in the brain 4 hours after treatment, after which the animals were immediately euthanized. Lesion sizes were measured on TTC-stained specimens, histological sections stained with hematoxylin and eosin (H&E), and T2-weighted MR images. The differences between the lesion diameters measured with the three methods were within the range of 0.1--0.7 mm. The lesion sizes measured from MRI correlated well with those seen from H&E sections. The measurements from MRI slightly overestimated lesion sizes on TTC-stained wet tissues by approximately one MRI pixel (0.31 mm). Electron microscopy demonstrated nuclear and cytoplasmic ultrastructural damage within the grey-white, non-TTC-stained lesion zone, whereas the TTC-stained normal tissue showed preservation of neuronal ultrastructure. Therefore, MR-imaged lesions represent a cell-death zone in rabbit brain 4 hours after FUS ablation, with slight overestimation by approximately one MRI pixel. J. Magn. Reson. Imaging 2001;13:23-30.

  View details for Web of Science ID 000171295800005

  View details for PubMedID 11169799

• A biochemical function for attractin in agouti-induced pigmentation and obesity NATURE GENETICS He, L., Gunn, T. M., Bouley, D. M., Lu, X. Y., Watson, S. J., Schlossman, S. F., Duke-Cohan, J. S., Barsh, G. S. 2001; 27 (1): 40-47

  Abstract

  Agouti protein, a paracrine signaling molecule normally limited to skin, is ectopically expressed in lethal yellow (A(y)) mice, and causes obesity by mimicking agouti-related protein (Agrp), found primarily in the hypothalamus. Mouse attractin (Atrn) is a widely expressed transmembrane protein whose loss of function in mahogany (Atrn(mg-3J)/ Atrn(mg-3J)) mutant mice blocks the pleiotropic effects of A(y). Here we demonstrate in transgenic, biochemical and genetic-interaction experiments that attractin is a low-affinity receptor for agouti protein, but not Agrp, in vitro and in vivo. Additional histopathologic abnormalities in Atrn(mg-3J)/Atrn(mg-3J) mice and cross-species genomic comparisons indicate that Atrn has multiple functions distinct from both a physiologic and an evolutionary perspective.

  View details for Web of Science ID 000166187900013

  View details for PubMedID 11137996

• Disease attributed to Mycobacterium chelonae in South African clawed frogs (Xenopus laevis) COMPARATIVE MEDICINE GREEN, S. L., Lifland, B. D., Bouley, D. M., Brown, B. A., Wallace, R. J., Ferrell, J. E. 2000; 50 (6): 675-679

  Abstract

  The fast-growing nontuberculous mycobacterial species Mycobacterium chelonae was isolated from six captive South African clawed frogs (Xenopus laevis) with chronic weight loss and nonhealing ulcerative skin lesions. Three of the M. chelonae isolates were evaluated to confirm the species identification using polymerase chain reaction restriction analysis. Disease associated with M. chelonae is reported mainly in people and in fish. To our knowledge, this is the first report of disease associated with M. chelonae in a colony of captive Xenopus sp.

  View details for Web of Science ID 000166363600020

  View details for PubMedID 11200577

• mig-14 is a horizontally acquired, host-induced gene required for Salmonella enterica lethal infection in the murine model of typhoid fever INFECTION AND IMMUNITY Valdivia, R. H., Cirillo, D. M., Lee, A. K., Bouley, D. M., FALKOW, S. 2000; 68 (12): 7126-7131

  Abstract

  We have characterized a host-induced virulence gene, mig-14, that is required for fatal infection in the mouse model of enteric fever. mig-14 is present in all Salmonella enterica subspecies I serovars and maps to a region of the chromosome that appears to have been acquired by horizontal transmission. A mig-14 mutant replicated in host tissues early after infection but was later cleared from the spleens and livers of infected animals. Bacterial clearance by the host occurred concomitantly with an increase in gamma interferon levels and recruitment of macrophages, but few neutrophils, to the infection foci. We hypothesize that the mig-14 gene product may repress immune system functions by interfering with normal cytokine expression in response to bacterial infections.

  View details for Web of Science ID 000167020000081

  View details for PubMedID 11083839

• Salmonella exploits caspase-1 to colonize Peyer's patches in a murine typhoid model JOURNAL OF EXPERIMENTAL MEDICINE Monack, D. M., Hersh, D., Ghori, N., Bouley, D., Zychlinsky, A., FALKOW, S. 2000; 192 (2): 249-258

  Abstract

  Salmonella typhimurium invades host macrophages and induces apoptosis and the release of mature proinflammatory cytokines. SipB, a protein translocated by Salmonella into the cytoplasm of macrophages, is required for activation of Caspase-1 (Casp-1, an interleukin [IL]-1beta-converting enzyme), which is a member of a family of cysteine proteases that induce apoptosis in mammalian cells. Casp-1 is unique among caspases because it also directly cleaves the proinflammatory cytokines IL-1beta and IL-18 to produce bioactive cytokines. We show here that mice lacking Casp-1 (casp-1(-/)- mice) had an oral S. typhimurium 50% lethal dose (LD(50)) that was 1,000-fold higher than that of wild-type mice. Salmonella breached the M cell barrier of casp-1(-/)- mice efficiently; however, there was a decrease in the number of apoptotic cells, intracellular bacteria, and the recruitment of polymorphonuclear lymphocytes in the Peyer's patches (PP) as compared with wild-type mice. Furthermore, Salmonella did not disseminate systemically in the majority of casp-1(-/)- mice, as demonstrated by significantly less colonization in the PP, mesenteric lymph nodes, and spleens of casp-1(-/)- mice after an oral dose of S. typhimurium that was 100-fold higher than the LD(50). The increased resistance in casp-1(-/)- animals appears specific for Salmonella infection since these mice were susceptible to colonization by another enteric pathogen, Yersinia pseudotuberculosis, which normally invades the PP. These results show that Casp-1, which is both proapoptotic and proinflammatory, is essential for S. typhimurium to efficiently colonize the cecum and PP and subsequently cause systemic typhoid-like disease in mice.

  View details for Web of Science ID 000088261100011

  View details for PubMedID 10899911

• Induction of angiogenesis by implantation of encapsulated primary myoblasts expressing vascular endothelial growth factor JOURNAL OF GENE MEDICINE Springer, M. L., Hortelano, G., Bouley, D. M., Wong, J., Kraft, P. E., Blau, H. M. 2000; 2 (4): 279-288

  Abstract

  We previously demonstrated that intramuscular implantation of primary myoblasts engineered to express vascular endothelial growth factor (VEGF) constitutively resulted in hemangioma formation and the appearance of VEGF in the circulation. To investigate the potential for using allogeneic myoblasts and the effects of delivery of VEGF-expressing myoblasts to non-muscle sites, we have enclosed them in microcapsules that protect allogeneic cells from rejection, yet allow the secretion of proteins produced by the cells.Encapsulated mouse primary myoblasts that constitutively expressed murine VEGF164, or encapsulated negative control cells, were implanted either subcutaneously or intraperitoneally into mice.Upon subcutaneous implantation, capsules containing VEGF-expressing myoblasts gave rise to large tissue masses at the implantation site that continued to grow and were composed primarily of endothelial and smooth muscle cells directly surrounding the capsules, and macrophages and capillaries further away from the capsules. Similarly, when injected intraperitoneally, VEGF-producing capsules caused significant localized inflammation and angiogenesis within the peritoneum, and ultimately led to fatal intraperitoneal hemorrhage. Notably, however, VEGF was not detected in the plasma of any mice.We conclude that encapsulated primary myoblasts persist and continue to secrete VEGF subcutaneously and intraperitoneally, but that the heparin-binding isoform VEGF164 exerts localized effects at the site of production. VEGF secreted from the capsules attracts endothelial and smooth muscle cells in a macrophage-independent manner. These results, along with our previous results, show that the mode and site of delivery of the same factor by the same engineered myoblasts can lead to markedly different outcomes. Moreover, the results confirm that constitutive delivery of high levels of VEGF is not desirable. In contrast, regulatable expression may lead to efficacious, safe, and localized VEGF delivery by encapsulated allogeneic primary myoblasts that can serve as universal donors.

  View details for Web of Science ID 000088563200007

  View details for PubMedID 10953919

• Quantitative comparison of MR imaged focused ultrasound lesions and cell death zones in the in vivo rabbit brain 22nd Annual International Conference of the IEEE-Engineering-in-Medicine-and-Biology-Society Chen, L., Bouley, D., Harris, B., Butts, K. IEEE. 2000: 805–808

  View details for Web of Science ID 000166896300224

• Cryptosporidiosis, coccidiosis, and total colonic mucosal collapse in an immunosuppressed puppy JOURNAL OF THE AMERICAN ANIMAL HOSPITAL ASSOCIATION Willard, M. D., Bouley, D. 1999; 35 (5): 405-409

  Abstract

  An eight-week-old puppy with chronic diarrhea was diagnosed with simultaneous opportunistic pathogens (i.e., cryptosporidiosis, coccidiosis) and total colonic mucosal collapse. Lack of lymphoid follicles in the spleen and lymph nodes suggested a primary underlying immunosuppression that most likely permitted infection with these pathogens. Intensive antibiotic therapy was most likely responsible for the severe colonic lesion, and bismuth subsalicylate administration in this severely dehydrated puppy may have contributed to renal failure as the ultimate cause of death.

  View details for Web of Science ID 000082405800009

  View details for PubMedID 10493416

• Study of focused ultrasound tissue damage using MRI and histology JOURNAL OF MAGNETIC RESONANCE IMAGING Chen, L., Bouley, D., Yuh, E., D'Arceuil, H., Butts, K. 1999; 10 (2): 146-153

  Abstract

  This paper reports on an experimental study of in vivo tissue damage in the rabbit brain with focused ultrasound (FUS) using magnetic resonance imaging (MRI) and histopathological analysis. Ten ultrasonic lesions (tissue damage) were created in five rabbits using a focused ultrasound beam of 1.5 MHz, electrical power input to the transducer of 70-85 W, and an exposure duration of 15-20 seconds. T1- and T2-weighted fast spin-echo (FSE) and Fluid attenuated inversion recovery (FLAIR) sequences were used to detect the ultrasonic lesions after treatment. Imaging was performed for 4-8 hours after treatment, after which the animals were immediately sacrificed. Ultrasonic lesion diameter was measured on MRI and histological sections after correction for tissue shrinkage during the histological processing. The T1-weighted images showed lesions poorly, whereas both T2-weighted and FLAIR images showed lesions clearly. The lesion diameters on both T2 and FLAIR imaging correlated well with measurements from histology. The time delay before lesions appeared on T2-weighted imaging was 15 minutes to 1 hour, depending on the exposure location in the brain. J. Magn. Reson. Imaging 1999;10:146-153.

  View details for Web of Science ID 000084567000006

  View details for PubMedID 10441017

• Identification and management of an outbreak of Flavobacterium meningosepticum infection in a colony of South African clawed frogs (Xenopus laevis) 1997 American-Association-of-Laboratory-Animal-Science Meeting GREEN, S. L., Bouley, D. M., Tolwani, R. J., Waggie, K. S., Lifland, B. D., Otto, G. M., Ferrell, J. E. AMER VETERINARY MEDICAL ASSOC. 1999: 1833-?

  Abstract

  During the summer of 1996, an outbreak of Flavobacterium meningosepticum infection developed in a colony of South African clawed frogs (Xenopus laevis). Clinical signs were consistent with septicemia: ascites, anasarca, dyspnea, extreme lethargy, congestion of web vessels, petechial hemorrhages, and sudden death. Mortality rate reached 35%, and all infections were fatal. The organism was resistant to most antibiotics but was susceptible to enrofloxacin, chloramphenicol, and trimethoprim-sulfadiazine. Treatment with trimethoprim-sulfadiazine was unsuccessful. Although the point source of the infection was not determined, several environmental reservoirs were identified, including a communal water barrel and various pieces of equipment. Molecular strain typing by pulsed-field gel electrophoresis and biochemical analyses revealed that frogs were infected with a single strain of F meningosepticum. Sanitation and management procedures were effective in controlling the outbreak.

  View details for Web of Science ID 000080933900027

  View details for PubMedID 10382028

• Partial tracheal obstruction due to chondromas in ball pythons (Python regius) JOURNAL OF ZOO AND WILDLIFE MEDICINE Drew, M. L., Phalen, D. N., Berridge, B. R., Johnson, T. L., Bouley, D., Weeks, B. R., Miller, H. A., Walker, M. A. 1999; 30 (1): 151-157

  Abstract

  Over a 9-mo period, three adult ball pythons (Python regius) (one male, two females) were evaluated for severe dyspnea. Partial obstructions of the tracheal lumen were identified radiographically and/or visualized with a 3.0-mm rigid laparoscope inserted into the tracheal lumen in all three snakes. Administration of systemic antibiotics and nebulization resulted in partial improvement of the dyspnea. In two snakes, the tracheal lesions were removed with a rigid laparoscope and a flexible biopsy instrument inserted into the tracheal lumen. The other snake died and was necropsied. Histologically, the lesions from two snakes were determined to be benign chondromas. The chondromas were composed of a variably disorganized chondroid matrix populated by quiescent, normal-appearing chondrocytes within lacunae, although the chondrocytes were increased in density compared with normal hyaline cartilage and contained rare mitotic figures. The tracheal masses in one snake grew by expansion, not invasion, and were focally continuous with a mineralized cartilage tracheal ring, suggesting a benign nature. This is the second report of tracheal chondroma in ball pythons. Tracheal chondromas are exceedingly rare in humans and domesticated animals, suggesting a possible predisposition of ball pythons for this neoplasm.

  View details for Web of Science ID 000080205700022

  View details for PubMedID 10367658

• Possible antibiotic-associated colitis in a dog JOURNAL OF THE AMERICAN VETERINARY MEDICAL ASSOCIATION Willard, M. D., Berridge, B., Braniecki, A., Bouley, D. 1998; 213 (12): 1775-?

  Abstract

  A Poodle referred for renal disease developed severe colonic disease characterized by total mucosal collapse and necrosis. The onset of colonic disease was temporally related to administration of antibiotics. On 3 occasions, bacterial culture of fecal samples yielded only Streptococcus pyogenes, Pseudomonas aeruginosa, and Candida albicans, suggesting that the colonic bacterial flora was severely disrupted. Findings, although not conclusive, were suggestive of antibiotic-associated colitis that ultimately proved fatal. Colonoscopy should be considered for dogs with unduly severe large-bowel diarrhea associated with antibiotic treatment and can be done with minimal restraint and bowel preparation, if necessary. Nonselective bacterial culture of fecal samples should be considered for dogs with unduly severe large-bowel diarrhea associated with antibiotic treatment.

  View details for Web of Science ID 000077464000025

  View details for PubMedID 9861973

• Yersinia-induced apoptosis in vivo aids in the establishment of a systemic infection of mice JOURNAL OF EXPERIMENTAL MEDICINE Monack, D. M., Mecsas, J., Bouley, D., FALKOW, S. 1998; 188 (11): 2127-2137

  Abstract

  Pathogenic Yersinia cause a systemic infection in mice that is dependent on the presence of a large plasmid encoding a number of secreted virulence proteins called Yops. We previously demonstrated that a plasmid-encoded Yop, YopJ, was essential for inducing apoptosis in cultured macrophages. Here we report that YopJ is a virulence factor in mice and is important for the establishment of a systemic infection. The oral LD50 for a yopJ mutant Yersinia pseudotuberculosis increases 64-fold compared with wild-type. Although the yopJ mutant strain is able to reach the spleen of infected mice, the mutant strain seldom reaches the same high bacterial load that is seen with wild-type Yersinia strain and begins to be cleared from infected spleens on day 4 after infection. Furthermore, when in competition with wild-type Yersinia in a mixed infection, the yopJ mutant strain is deficient for spread from the Peyer's patches to other lymphoid tissue. We also show that wild-type Yersinia induces apoptosis in vivo of Mac-1(+) cells from infected mesenteric lymph nodes or spleens, as measured by quantitative flow cytometry of TUNEL (Tdt-mediated dUTP-biotin nick-end labeling)-positive cells. The levels of Mac-1(+), TUNEL+ cells from tissue infected with the yopJ mutant strain were equivalent to the levels detected in cells from uninfected tissue. YopJ is necessary for the suppression of TNF-alpha production seen in macrophages infected with wild-type Yersinia, based on previous in vitro studies (Palmer, L.E., S. Hobbie, J.E. Galan, and J.B. Bliska. 1998. Mol. Microbiol. 27:953-965). We conclude here that YopJ plays a role in the establishment of a systemic infection by inducing apoptosis and that this is consistent with the ability to suppress the production of the proinflammatory cytokine tumor necrosis factor alpha.

  View details for Web of Science ID 000077484700016

  View details for PubMedID 9841926

• Progressive ataxia, myoclonic epilepsy and cerebellar apoptosis in cystatin B-deficient mice NATURE GENETICS Pennacchio, L. A., Bouley, D. M., Higgins, K. M., Scott, M. P., Noebels, J. L., Myers, R. M. 1998; 20 (3): 251-258

  Abstract

  Loss-of-function mutations in the gene (CSTB) encoding human cystatin B, a widely expressed cysteine protease inhibitor, are responsible for a severe neurological disorder known as Unverricht-Lundborg disease (EPM1). The primary cellular events and mechanisms underlying the disease are unknown. We found that mice lacking cystatin B develop myoclonic seizures and ataxia, similar to symptoms seen in the human disease. The principal cytopathology appears to be a loss of cerebellar granule cells, which frequently display condensed nuclei, fragmented DNA and other cellular changes characteristic of apoptosis. This mouse model of EPM1 provides evidence that cystatin B, a non-caspase cysteine protease inhibitor, has a role in preventing cerebellar apoptosis.

  View details for Web of Science ID 000076698100018

  View details for PubMedID 9806543

• The role of the innate immune system in the reconstituted SCID mouse model of herpetic stromal keratitis CLINICAL IMMUNOLOGY AND IMMUNOPATHOLOGY Bouley, D. M., Kanangat, S., Rouse, B. T. 1996; 80 (1): 23-30

  Abstract

  Herpetic stromal keratitis (HSK) has an immunopathological basis, thought primarily to involve a CD4+ T cell-mediated immune response to viral antigen. Other cell types, however, particularly those involved in nonspecific immunity, such as natural killer (NK) cells or neutrophils, may also contribute to tissue destruction in the cornea. The reconstituted SCID mouse model of HSK provides a powerful system in which to study the interactions of the innate and adaptive immune responses to herpes simplex virus type 1 corneal infection. In the present study, reconstituted SCID mice depleted of NK cells had a reduced incidence and severity of clinical and histopathological HSK. The levels of T cell cytokine protein and message in restimulated splenocytes and cytokine message in corneas did not differ between experimental groups. However, significantly fewer neutrophils were seen within the inflamed corneas of NK-depleted SCID mice. Therefore, endogenous NK cells may indirectly influence the severity of HSK in reconstituted SCID mice by affecting neutrophil migration into the cornea.

  View details for Web of Science ID A1996UU00700005

  View details for PubMedID 8674236

• CHARACTERIZATION OF HERPES-SIMPLEX VIRUS TYPE-1 INFECTION AND HERPETIC STROMAL KERATITIS DEVELOPMENT IN IFN-GAMMA KNOCKOUT MICE JOURNAL OF IMMUNOLOGY Bouley, D. M., Kanangat, S., WIRE, W., Rouse, B. T. 1995; 155 (8): 3964-3971

  Abstract

  Herpetic stromal keratitis (HSK) has an immune-mediated pathogenesis that involves T cells that have a type 1 cytokine profile. IFN-gamma is suspected to be the type 1 cytokine involved in ocular pathology, and to test this notion more directly the pathogenesis of HSK was compared in mice deficient in the IFN-gamma gene (gamma knockout or gko) and control mice (wild-type littermates or BALB/c mice). The clinical course of HSK in gko mice closely paralleled that in control mice, yet virus persisted in the corneas of gko mice for an extended period of time, severe periocular skin lesions developed, and gko mice were far more susceptible to encephalitis. Delayed-type hypersensitivity to viral Ag was present, though diminished, in knockout mice, and serum herpes simplex virus-specific IgG isotypes indicated a Th2 shift. No differences existed in proliferative responses to in vitro Ag stimulation in gko vs control mice nor in T cell or proinflammatory cytokine mRNA levels in the corneas of infected mice. However, up-regulation of Th2 cytokine mRNA did occur in in vitro Ag-stimulated gko immune splenocytes. Histopathologic lesions were not statistically different between any of the groups of mice analyzed. These observations indicate that although IFN-gamma plays an important role in the clearance of virus from the eye, the pathogenesis of HSK lesions most likely involves additional cytokines, inflammatory mechanisms, or immune responses to nonviral Ags.

  View details for Web of Science ID A1995RY58100034

  View details for PubMedID 7561104

• LYMPHOTOXIN-ALPHA-DEFICIENT MICE - EFFECTS ON SECONDARY LYMPHOID ORGAN DEVELOPMENT AND HUMORAL IMMUNE RESPONSIVENESS JOURNAL OF IMMUNOLOGY Banks, T. A., Rouse, B. T., Kerley, M. K., Blair, P. J., Godfrey, V. L., Kuklin, N. A., Bouley, D. M., Thomas, J., Kanangat, S., Mucenski, M. L. 1995; 155 (4): 1685-1693

  Abstract

  Targeted mutagenesis in embryonic stem cells was used to generate mice deficient in lymphotoxin-alpha (LT-alpha). Mice lacking LT-alpha -/- (LT-alpha -/- mice) exhibit a phenotype dominated by defects in secondary lymphoid organ development. LT-alpha -/- mice lack lymph nodes and Peyer's patches, and possess spleens in which the usual architecture is disrupted. However, in a few of the mutants, abnormal lymph node-like structures were observed, mainly within the mesenteric fat. Abnormal clusters of lymphocytes were also found to accumulate in the periportal and perivascular regions of the liver and lung of LT-alpha -/- mice. Yet, lymphocytes from LT-alpha -/- mice appeared phenotypically normal, expressing the expected ratios of B and T cell surface markers as well as the lymphocyte homing marker, L-selectin. In addition, bone marrow cells from LT-alpha -/- mice were able to successfully reconstitute the lymphoid organs of severe combined immunodeficient mice. However, LT-alpha -/- mutant mice examined for humoral immune responsiveness were found to be impaired in their ability to respond to different Ag. These data illustrate the utility of this mouse model as a system for understanding lymphoid organ development and its effects on immune responsiveness.

  View details for Web of Science ID A1995RN46400006

  View details for PubMedID 7636227

• RADIOGRAPHIC AND SCINTIGRAPHIC EVIDENCE OF FOCAL PULMONARY NEOPLASIA IN 3 CATS WITH HYPERTHYROIDISM - DIAGNOSTIC AND THERAPEUTIC CONSIDERATIONS JOURNAL OF VETERINARY INTERNAL MEDICINE Cook, S. M., Daniel, G. B., Walker, M. A., Maddux, J. M., Jenkins, C. C., Klebanow, E. R., Bouley, D. M., Dean, D. F., Petersen, M. G. 1993; 7 (5): 303-308

  Abstract

  Three cats were diagnosed as hyperthyroid based on clinical signs, historical findings, laboratory abnormalities, and basal serum thyroxine (T4) concentrations, and/or nuclear thyroid scans. Additionally, a presumptive diagnosis of thyroid carcinoma with pulmonary metastasis was made in each cat based on radiographic or scintigraphic evaluation. All three cats had solitary pulmonary nodules 1.5 to 2 cm in diameter on survey thoracic radiographs; one cat also had chylous pleural effusion and pulmonary lobar consolidation. Focal pulmonary accumulation of sodium pertechnetate (99mTcO4-) and/or radioiodine (131I) corresponding to radiographic lesions were seen in all cats. Two cats were treated with single ablative doses (1111 to 1480 MBq) of 131I; the remaining cat was euthanatized. One of the treated cats died 8 days later; the other cat was euthanatized 22 weeks following treatment. Histopathologic examination of tissue obtained at necropsy confirmed metastatic thyroid carcinoma in one cat and bronchogenic adenocarcinoma in two cats. Our findings indicate that increased radionuclide uptake in focal pulmonary lesions and cytologic evaluation of tissue obtained by fine-needle aspiration are not specific for thyroid tissue.

  View details for Web of Science ID A1993MD60100007

  View details for PubMedID 8263849

• HERPETIC STROMAL KERATITIS IN THE RECONSTITUTED SCID MOUSE MODEL JOURNAL OF VIROLOGY MERCADAL, C. M., Bouley, D. M., DESTEPHANO, D., Rouse, B. T. 1993; 67 (6): 3404-3408

  Abstract

  Infections of the cornea with herpes simplex virus type 1 cause inflammatory lesions which frequently lead to blindness. The disease is suspected to be immunopathological in nature. To establish this point and to study possible mechanisms involved, corneal infections in C.B-17 scid/scid and cell-reconstituted scid mice were investigated. Whereas unreconstituted scid mice failed to develop herpetic stromal keratitis (HSK) and died of encephalitis, mice reconstituted with T lymphocytes generated severe lesions. T cells of the CD4+ subset were found to be essential mediators of the HSK lesion, while T cells of the CD8+ subset protected mice from lethality. The results confirm that HSK is an immunopathological disease and that scid mice provide a convenient model that should prove valuable in establishing the biochemical mechanisms by which HSK is mediated.

  View details for Web of Science ID A1993LB79400053

  View details for PubMedID 8098778