- Pediatric Nephrology
Medical Director of Pediatric Kidney Transplant Program, LPCH at Stanford (2012 - Present)
Pediatric Nephrology Fellowship Director, Stanford (2007 - Present)
Honors & Awards
Member, American Board of Pediatrics-Pediatric Nephrology Subboard (2010-2018)
Residency:Winnipeg Children's Hospital (1986) Canada
Residency:Dalhousie University (1985) Canada
Fellowship:Dalhousie University (1988) Canada
Fellowship:UCLA Health Sciences (1991) CA
Internship:University of Saskatchewan (1982) Canada
Board Certification: Pediatric Nephrology, American Board of Pediatrics (1999)
Medical Education:University of Saskatchewan College of Medicine (1981) SKCanada
Fellowship, UCLA, Pediatric Transplant Immunology (1991)
Fellowship, Dalhousie University, Halifax, Pediatric Nephrology (1988)
Pediatric Residency, University of Manitoba, Pediatrics Yr III (1986)
Pediatric Residency, Dalhousie University, Halifax, Pediatrics I & II (1985)
Interneship, U. of Saskatchewan, Rotating Interne (1982)
MD with Distinction, University of Saskatchewan, Medicine (1981)
Current Research and Scholarly Interests
Computerized image analysis of kidney and liver biopsies to quantitate and diagnose subtle changes in tissue structure.
Renal Allograft Rejection
Renal Fibrosis in;
-Primary Kidney Disease
-Transplant Kidney Disease
- Independent Studies (5)
Quality of Life is Improved and Kidney Function Preserved in Patients with Nephropathic Cystinosis Treated for 2 Years with Delayed-Release Cysteamine Bitartrate.
journal of pediatrics
2014; 165 (3): 528-533 e1
To determine the long-term effects of delayed-release cysteamine bitartrate (DR-CYS) based on our previous work that established the short-term noninferiority of DR-CYS every 12 hours compared with immediate-release cysteamine bitartrate every 6 hours.We conducted a prospective, controlled, open label, single-arm study of DR-CYS for 2 years in 40 patients to assess efficacy in depletion of cystine in peripheral white blood cells, to assess the dose required to maintain white blood cell content of cystine <1 nmol ½ cystine/mg protein, to measure quality of life using the Pediatric Quality of Life Inventory, change in estimated glomerular filtration rate, and change in height Z-score.Through 24 months of study, the mean white blood cell content of cystine was always <1 nmol ½ cystine/mg protein, and the dose of DR-CYS decreased from 43.5-40.1 mg/kg/d (P = .05), and the significant improvement in social function, school function, and in total function scores on the Pediatric Quality of Life Inventory remained. The estimated glomerular filtration rate was maintained and growth velocity was maintained at 24 months compared with the baseline height Z-score.The use of a DR-CYS administered every 12 hours to patients with cystinosis is of great benefit to their quality of life and to important biomarkers of disease control, when studied in a prospective, controlled fashion. We suggest that DR-CYS should be considered for substrate depletion in patients with cystinosis.
View details for DOI 10.1016/j.jpeds.2014.05.013
View details for PubMedID 24948347
Immune cell function assay does not identify biopsy-proven pediatric renal allograft rejection or infection.
2014; 18 (5): 446-452
Management of pediatric renal transplant patients involves multifactorial monitoring modalities to ensure allograft survival and prevent opportunistic infection secondary to immunosuppression. An ICFA, which utilizes CD4 T-cell production of ATP to assess immune system status, has been used to monitor transplant recipients and predict susceptibility of patients to rejection or infection. However, the validity of this assay to reflect immune status remains unanswered. In a two-yr retrospective study that included 31 pediatric renal transplant recipients, 42 patient blood samples were analyzed for immune cell function levels, creatinine, WBC (white blood cell) count, immunosuppressive drug levels, and viremia, concurrent with renal biopsy. T-cell ATP production as assessed by ICFA levels did not correlate with allograft rejection or with the presence or absence of viremia. ICFA levels did not correlate with serum creatinine or immunosuppressive drug levels, but did correlate with WBC count. The ICFA is unreliable in its ability to reflect immune system status in pediatric renal transplantation. Further investigation is necessary to develop methods that will accurately predict susceptibility of pediatric renal transplant recipients to allograft rejection and infection.
View details for DOI 10.1111/petr.12295
View details for PubMedID 24930482
Use of eculizumab and plasma exchange in successful combined liver-kidney transplantation in a case of atypical HUS associated with complement factor H mutation
2014; 29 (3): 477-480
Atypical hemolytic uremic syndrome (aHUS) evolves into end-stage renal failure in nearly half of affected patients and is associated with defective regulation of the alternative complement pathway. Patients with a complement factor H (CFH) mutation have a 30-100% risk of graft loss due to aHUS recurrence or graft thrombosis. Since CFH is produced predominantly by the liver, combined liver-kidney transplant is a curative treatment option. One major unexpected risk includes liver failure secondary to uncontrolled complement activation. We report a successful combined liver-kidney transplantation with perioperative plasma exchange and use of the humanized anti-C5 monoclonal antibody eculizumab.An 11-month-old female presented with oliguric renal failure after 3 weeks of flu-like symptoms in the absence of diarrhea. Following the identification of Escherichia coli 0157:H7 in her stool, she was discharged home on peritoneal dialysis with a diagnosis of Shiga toxin-associated HUS. Three months later, she developed severe anemia, thrombocytopenia, and neurological involvement. aHUS was diagnosed and confirmed, and genetic testing revealed a mutation in CFH SCR20. Once donor organs became available, she received preoperative plasma exchange followed by eculizumab infusion with intra-operative fresh frozen plasma prior to combined liver-kidney transplant. At 19 months post-transplant, she continues to have excellent allograft and liver function without signs of disease recurrence.Perioperative use of eculizumab in conjunction with plasma exchange during simultaneous liver-kidney transplant can be used to inhibit terminal complement activity, thereby optimizing successful transplantation by reducing the risk of graft thrombosis.
View details for DOI 10.1007/s00467-013-2630-5
View details for Web of Science ID 000330976500019
Initial Experience Using Aminophylline to Improve Renal Dysfunction in the Pediatric Cardiovascular ICU
PEDIATRIC CRITICAL CARE MEDICINE
2014; 15 (1): 21-27
To determine if aminophylline administration is associated with improved creatinine clearance and greater urine output in children with acute kidney injury in the cardiovascular ICU.Single-center retrospective cohort study.Pediatric cardiovascular ICU, university-affiliated children's hospital.Children with congenital or acquired heart disease in the cardiovascular ICU who received aminophylline to treat oliguric acute kidney injury and fluid overload.Patients received aminophylline after consultation with a pediatric nephrologist. Data were collected retrospectively over 7 days to assess if aminophylline was associated with improvement in creatinine clearance, urine output, and fluid overload.Thirty-one patients received 52 aminophylline courses. Over the 7-day study period, serum creatinine decreased from a mean of 1.13 ± 0.91 to 0.87 ± 0.83 mg/dL (-0.05 mg/dL/d, p < 0.001). A concomitant increase was seen in estimated glomerular filtration rate from a mean of 50.0 ± 30.0 to 70.6 ± 58.1 mL/min/1.73 m (+3.66 mL/min/1.73 m/d, p < 0.001). Average daily urine output increased by 0.22 mL/kg/hr (p < 0.001), and fluid overload decreased on average by 0.42% per day in the 7-day study period (p = 0.005). Although mean furosemide dose increased slightly (0.12 mg/kg/d, p = 0.01), hydrochlorothiazide dosing did not significantly change over the study period. There were no complications related to aminophylline administration.Our study suggests that aminophylline therapy may be associated with significantly improved renal excretory function and may augment urine output in children who experience oliguric acute kidney injury in the cardiovascular ICU. Additionally, we did not identify any aminophylline-related side effects in this high-risk cardiac population. Future prospective studies are necessary to confirm the safety profile and to ensure that the beneficial effects are independent of other clinical interventions.
View details for DOI 10.1097/01.pcc.0000436473.12082.2f
View details for Web of Science ID 000329368400007
BK Polyomavirus Subtype III in a Pediatric Renal Transplant Patient with Nephropathy
JOURNAL OF CLINICAL MICROBIOLOGY
2013; 51 (12): 4255-4258
BK polyomavirus (BKV) is an emerging pathogen in immunocompromised individuals. BKV subtype III is rarely identified and has not previously been associated with disease. Here we provide the whole-genome sequence of a subtype III BKV from a pediatric kidney transplant patient with polyomavirus-associated nephropathy.
View details for DOI 10.1128/JCM.01801-13
View details for Web of Science ID 000327147100067
View details for PubMedID 24048534
Conversion From Tacrolimus/Mycophenolic Acid to Tacrolimus/Leflunomide to Treat Cutaneous Warts in a Series of Four Pediatric Renal Allograft Recipients
2012; 94 (5): 450-455
The challenge of immunosuppression in pediatric renal transplantation is to balance preventing rejection while avoiding infectious complications. A dermatological complication of immunosuppression is viral warts, which cause significant disfigurement and increase the risk of skin malignancy.We present three pediatric and adolescent renal allograft recipients with multiple, recalcitrant verrucae vulgares lesions and one patient with molluscum contagiosum who were switched from mycophenolate mofetil to leflunomide. Teriflunomide metabolite levels were carefully maintained between 50,000 and 100,000 ng/mL to balance its immunosuppressive and antiviral properties. No adverse events requiring discontinuation of leflunomide were encountered.Switching from mycophenolate mofetil to leflunomide successfully cleared verrucae vulgares and molluscum lesions in all four renal transplant patients.The ability to minimize and even resolve warts can improve quality of life by reducing risk of skin malignancies and emotional distress in solid organ transplant patients. Leflunomide is a potential therapeutic option for posttransplantation patients with skin warts because it serves both as an adjunct to the immunosuppressive regimen and an antiviral agent.
View details for DOI 10.1097/TP.0b013e318264351e
View details for Web of Science ID 000308668000012
View details for PubMedID 22960763
Rituximab treatment for recurrence of nephrotic syndrome in a pediatric patient after renal transplantation for congenital nephrotic syndrome of Finnish type
2012; 16 (5): E183-E187
Congenital nephrotic syndrome (CNS) of the Finnish type due to mutation in the NPHS-1 gene results in massive proteinuria due to structural abnormality in the glomerular slit diaphragm, and is usually refractory to immunosuppressive therapy. Patients eventually require bilateral nephrectomy and renal replacement therapy, with transplantation being the ultimate goal. Post-transplant recurrence of nephrotic syndrome occurs in about 25% of children and is thought to be immune-mediated secondary to antibodies formed against the nephrin protein in renal allograft. Conventional therapy with calcineurin inhibitors (CNI), cyclophosphamide and corticosteroids with or without plasmapheresis often fails to achieve remission resulting in graft loss in 12-16%. There is limited experience with use of rituximab (RTX) in pediatric organ transplant recipients. We report the first case of post-transplant recurrence of nephrotic syndrome in a 4-yr-old child with CNS due to NPHS-1 mutation in whom CNI, corticosteroid and cyclophosphamide therapy was unsuccessful, but who achieved remission after depletion of B cells with RTX, associated with a decrease in the level of anti-nephrin antibodies. The child remains in remission 5 yr following treatment. Our experience suggests that activated B cells may play a pivotal role in the recurrence of nephrosis after renal transplantation in children with CNS.
View details for DOI 10.1111/j.1399-3046.2011.01519.x
View details for Web of Science ID 000306131700011
View details for PubMedID 21672106
A Randomized Controlled Crossover Trial with Delayed-Release Cysteamine Bitartrate in Nephropathic Cystinosis: Effectiveness on White Blood Cell Cystine Levels and Comparison of Safety
CLINICAL JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY
2012; 7 (7): 1112-1120
Immediate-release cysteamine bitartrate (Cystagon; Mylan Pharmaceuticals, Canonsburg, PA) may prevent or delay kidney transplantation and other serious outcomes in patients with cystinosis, but has never been subjected to a prospective clinical trial. Cystagon efficacy requires strict lifelong dosing every 6 hours. Such a dosing schedule and Cystagon-associated side effects are often cited by patients as reasons for nonadherence.This open-label, randomized, controlled, crossover trial was powered to show that a new delayed-release formulation of cysteamine bitartrate, RP103, taken every 12 hours, was noninferior to Cystagon for maintenance of white blood cell (WBC) cystine at levels associated with optimal outcomes in the disease.Forty-three patients were randomized. Using a mixed-effects statistical analysis model, the least-squares mean peak value of WBC cystine level was 0.62±0.05 nmol 1/2 cystine/mg protein after 12 hours under RP103 and 0.54±0.05 nmol 1/2 cystine/mg protein after 6 hours under Cystagon, a difference of 0.08±0.04 nmol 1/2 cystine/mg protein (95.8% confidence interval, 0-0.16). The average steady-state total daily dose of RP103 was 82% of the incoming steady-state total daily dose of Cystagon. There were three-fold more gastrointestinal side effects compared with using Cystagon.A new delayed-release Q12H formulation of cysteamine bitartrate is not inferior to the Q6H formulation (Cystagon) in maintaining low WBC cystine levels in patients with cystinosis but at a lower total daily dose.
View details for DOI 10.2215/CJN.12321211
View details for Web of Science ID 000306148500010
View details for PubMedID 22554716
- Steroid-free immunosuppression in teenagers: Living without a safety net PEDIATRIC TRANSPLANTATION 2012; 16 (4): 305-307
Steroid avoidance in renal transplantation
CURRENT OPINION IN ORGAN TRANSPLANTATION
2011; 16 (5): 477-482
The recent surge in the use of steroid-avoidance protocols for pediatric renal transplant recipients has been fueled by the numerous adverse side effects of steroids and development of alternatives for successful immunosuppression. Steroid-avoidance protocols were first attempted in the adult population, and with positive outcomes, pediatrics soon followed. As more pediatric patients are placed on steroid-avoidance protocols, we must begin answering several important questions such as patient and graft outcome, safety profiles of various steroid-avoidance induction protocols, viral complications and incidence of transplant lymphoproliferative disease (PTLD), metabolic benefits, and the affect of steroid minimization on growth.Initial results from steroid-avoidance protocols show these protocols are safe and effective with improved graft survival, metabolic profiles, and linear growth without an increase in viremia or PTLD.Although initial results are promising, there is still a lack of long-term data from large, prospective randomized trials, and there is not enough data to determine the optimal steroid-avoidance protocol for pediatric renal transplant recipients.
View details for DOI 10.1097/MOT.0b013e32834a8c74
View details for Web of Science ID 000294676600006
View details for PubMedID 21844809
Screening for NPHS2 Mutations May Help Predict FSGS Recurrence after Transplantation
JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY
2011; 22 (3): 579-585
Steroid-resistant focal segmental glomerulosclerosis (FSGS) often recurs after renal transplantation. In this international survey, we sought to identify genotype-phenotype correlations of recurrent FSGS. We surveyed 83 patients with childhood-onset primary FSGS who received at least one renal allograft and analyzed 53 of these patients for NPHS2 mutations. The mean age at diagnosis was 6.7 years, and the mean age at first renal transplantation was 13 years. FSGS recurred in 30 patients (36%) after a median of 13 days (range, 1.5 to 152 days). Twenty-three patients received a second kidney transplant, and FSGS recurred in 11 (48%) after a median of 16 days (range, 2.7 to 66 days). None of the 11 patients with homozygous or compound heterozygous NPHS2 mutations developed recurrent FSGS compared with 45% of patients without mutations. These data suggest that genetic testing for pathogenic mutations may be important for prognosis and treatment of FSGS both before and after transplantation.
View details for DOI 10.1681/ASN.2010010029
View details for Web of Science ID 000288778800025
View details for PubMedID 21355056
Morphometric and Visual Evaluation of Fibrosis in Renal Biopsies
JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY
2011; 22 (1): 176-186
Interstitial fibrosis is an outcome measure of increasing importance in clinical trials of both renal transplantation and native disease, but data on the comparative advantages of fibrosis measurement methods are limited. We compared four morphometric techniques and contrasted these with two visual fibrosis-scoring methods on trichrome-stained slides. Two morphometric methods included whole-slide digital images: collagen III immunohistochemistry and a new technique using trichrome and periodic acid-Schiff subtraction morphometry; the other two methods included Sirius Red with and without polarization on multiple digital fields. We evaluated 10 serial sections from 15 renal biopsies with a range of fibrosis extent and diagnoses on duplicate sections with each method on separate days. Three pathologists performed visual scoring on whole-slide images. Visual and morphometric techniques had good to excellent interassay reproducibility (R(2) = 0.62 to 0.96) and interobserver reproducibility (R(2) = 0.75 to 0.99, all P < 0.001). Morphometry showed less variation between observers than visual assessment (mean of 1% to 5% versus 11% to 13%). Collagen III, Sirius Red unpolarized, and visual scores had the strongest correlations (R(2) = 0.78 to 0.89), the greatest dynamic range, and the best correlation with estimated GFR (R(2) = 0.38 to 0.50, P < 0.01 to 0.001). Considering efficiency, reproducibility, and functional correlation, two current techniques stand out as potentially the best for clinical trials: collagen III morphometry and visual assessment of trichrome-stained slides.
View details for DOI 10.1681/ASN.2009091005
View details for Web of Science ID 000288046500024
View details for PubMedID 21115619
Steroid-Free Immunosuppression Since 1999: 129 Pediatric Renal Transplants with Sustained Graft and Patient Benefits
AMERICAN JOURNAL OF TRANSPLANTATION
2009; 9 (6): 1362-1372
Despite early promising patient and graft outcomes with steroid-free (SF) immunosuppression in pediatric kidney transplant recipients, data on long-term safety and efficacy results are lacking. We present our single-center experience with 129 consecutive pediatric kidney transplant recipients on SF immunosuppression, with a mean follow-up of 5 years. Outcomes are compared against a matched cohort of 57 concurrent recipients treated with steroid-based (SB) immunosuppression. In the SF group, 87% of kidney recipients with functioning grafts remain corticosteroid-free. Actual intent-to-treat SF (ITT-SF) and still-on-protocol SF patient survivals are 96% and 96%, respectively, actual graft survivals for both groups are 93% and 96%, respectively and actual death-censored graft survivals for both groups are 97% and 99%, respectively. Unprecedented catch-up growth is observed in SF recipients below 12 years of age. Continued low rates of acute rejection, posttransplant diabetes mellitus (PTDM), hypertension and hyperlipidemia are seen in SF patients, with sustained benefits for graft function. In conclusion, extended enrollment and longer experience with SF immunosuppression for renal transplantation in low-risk children confirms protocol safety, continued benefits for growth and graft function, low acute rejection rates and reduced cardiovascular morbidity.
View details for DOI 10.1111/j.1600-6143.2009.02640.x
View details for Web of Science ID 000266448900017
View details for PubMedID 19459814
Melamine nephrotoxicity: an emerging epidemic in an era of globalization
2009; 75 (8): 774-779
Recent outbreaks of nephrolithiasis and acute kidney injury among children in China have been linked to ingestion of milk-based infant formula contaminated with melamine. These cases provide evidence in humans for the nephrotoxicity of melamine, which previously had been described only in animals. The consequences of this outbreak are already severe and will likely continue to worsen. Herein we summarize the global impact of the melamine milk contamination, the reemergence of melamine-tainted animal feed, and potential mechanisms of melamine nephrotoxicity. Large-scale epidemiologic studies are necessary to further characterize this disease and to assess its potential long-term sequelae. This epidemic of environmental kidney disease highlights the morbidity associated with adulterated food products available in today's global marketplace and reminds us of the unique vulnerability of the kidney to environmental insults. Melamine is the latest in a growing list of diverse potentially toxic compounds about which nephrologists and other health-care providers responsible for the diagnosis and management of kidney disease must now be aware.
View details for DOI 10.1038/ki.2009.16
View details for Web of Science ID 000264747900005
View details for PubMedID 19212415
Multicenter trial of everolimus in pediatric renal transplant recipients: Results at three year
2008; 12 (4): 456-463
There are few prospective clinical trials of mTOR inhibitors (or proliferation signal inhibitors) combined with CNI inhibitors in de novo pediatric renal transplantation. Results reported here are from a multicenter, open-label study in de novo pediatric renal transplant patients (
View details for DOI 10.1111/j.1399-3046.2007.00832.x
View details for Web of Science ID 000255551700015
View details for PubMedID 18466433
Incidence of PTLD in pediatric renal transplant recipients receiving basiliximab, calcineurin inhibitor, sirolimus and steroids
AMERICAN JOURNAL OF TRANSPLANTATION
2008; 8 (5): 984-989
Pediatric renal transplant recipients were enrolled in a multicenter, randomized, double-blind trial of steroid withdrawal. Subjects received basiliximab, calcineurin inhibitor, sirolimus and steroids. Of 274 subjects enrolled, 19 (6.9%) subjects developed posttransplant lymphoproliferative disorder (PTLD). The relative hazard (RH) for PTLD was 5.3-fold higher in children aged < or =5 versus those >12 years (p = 0.0017). EBV seronegative subjects had a 4.7-fold higher RH compared to EBV positive subjects (p = 0.02). Among EBV donor+/recipient- (D+/R-) subjects, the RH increased by 6.1-fold (p = 0.0001). In a multivariate model, risk factors included recipient age < or =5 years (RH 3.2, 95% CI: 1.1-9.6, p = 0.034) and EBV D+/R- status (RH 7.7, 95% CI: 1.6-35.9, p = 0.010). Of 19 patients with PTLD, 17 are alive with functioning grafts and 2 lost their grafts, 1 of whom subsequently died of recurrent PTLD. This 'robust' immunosuppression protocol was associated with low rejection rates but an unacceptably high incidence of PTLD. The combination of basiliximab, calcineurin inhibitor, sirolimus and steroids resulted in over-immunosuppression in a high-risk pediatric population and we do not recommend its use. Future studies must include routine viral monitoring to permit early identification of viral activity and a protocol driven reduction of immunosuppression aimed at avoiding complications.
View details for DOI 10.1111/j.1600-6143.2008.02167.x
View details for Web of Science ID 000254988500012
View details for PubMedID 18416737
Banff '05 Meeting report: Differential diagnosis of chronic allograft injury and elimination of chronic allograft nephropathy ('CAN')
AMERICAN JOURNAL OF TRANSPLANTATION
2007; 7 (3): 518-526
The 8th Banff Conference on Allograft Pathology was held in Edmonton, Canada, 15-21 July 2005. Major outcomes included the elimination of the non-specific term "chronic allograft nephropathy" (CAN) from the Banff classification for kidney allograft pathology, and the recognition of the entity of chronic antibody-mediated rejection. Participation of B cells in allograft rejection and genomics markers of rejection were also major subjects addressed by the conference.
View details for DOI 10.1111/j.1600-6143.2006.01688.x
View details for Web of Science ID 000244715800007
View details for PubMedID 17352710
Sirolimus pharmacokinetics in pediatric renal transplant recipients receiving calcineurin inhibitor co-therapy
2006; 10 (8): 914-919
We have previously reported sirolimus (SRL) pharmacokinetics (PK) in pediatric renal transplant recipients on a calcineurin inhibitor (CNI)-free protocol. We now report pediatric SRL PK in pediatric renal transplant patients receiving SRL + CNI. SRL was dosed to achieve target trough levels between 10 and 20 ng/mL. We performed 49 SRL PK profiles in pediatric renal transplant recipients receiving SRL in combination with either cyclosporine (CsA; 25 profiles), or tacrolimus (TCL; 24 profiles). Ten of the SRL + TCL profiles were obtained from children receiving SRL on a b.i.d. dosing regimen. All other SRL profiles were q.d. regimens. We calculated, the maximum concentration (C(max)), AUC, apparent clearance (aCL; dose/AUC) for dose in mg/m(2), and mean residence time (MRT). SRL levels were measured at 6 and 7 time points for b.i.d. and q.d. dosing, respectively. Regression analysis of SRL trough values vs. AUC showed good correlation in the SRL q.d. + CsA, SRL q.d. + TCL, and SRL b.i.d. + TCL groups (r(2) = 0.95, 0.68, and 0.44, respectively). SRL aCL corrected for body surface area was higher in children aged 0-5 yr receiving SRL with either CsA or TCL. SRL dosing schedule should be tailored to each patient. Higher SRL aCL may be present in younger children when administered with CNI.
View details for DOI 10.1111/j.1399-3046.2006.00541.x
View details for Web of Science ID 000241678100007
View details for PubMedID 17096757
- Use of an immune function assay to monitor immunosuppression PEDIATRIC TRANSPLANTATION 2006; 10 (5): 533-535
Transcriptional analysis of the molecular basis of human kidney aging using cDNA microarray profiling
2005; 68 (6): 2667-2679
The molecular basis of renal aging is not completely understood.We used global gene expression monitoring by cDNA microarrays to identify age associated genes in human kidney samples. Our samples included young (8 weeks-8 years, N= 4), adult (31-46 years, N= 7), and old kidneys (71-88 years, N= 9).Old kidneys had more glomerulosclerosis, tubular atrophy, interstitial fibrosis, and fibrous intimal thickening in small arteries. We identified approximately 500 genes that were differentially expressed among the three age groups. Old kidneys appeared to have increased extracellular matrix turnover and a nonspecific inflammatory response, combined with a reduction in processes dependent on energy metabolism and mitochondrial function. Quantitative supervised bioinformatics analyses of adult and old kidney expression data correlated the expression of 255 gene profiles with renal pathology scores. Microarray class prediction analysis (PAM) identified 50 unique genes that segregated old kidneys into two distinct clusters: those more similar within age class (OO, N= 5) versus old kidneys more similar to adult kidneys (OA, N= 4). The expression of six functionally significant genes was further validated by quantitative reverse transcription-polymerase chain reaction (RT-PCR) (FN1, MMP7, TNC, SERPIN3A, BPHL, CSPG2) in the experiment group and, subsequently, confirmed independently in 17 additional old and adult age-stratified test kidney samples. The p53 inducible gene, CSPG2, performed best in separating OO kidneys from adults and OA samples in this analysis.The method described in this study using independent validation samples can be envisioned to test utility of the identified genes in assessing age-related changes that contribute to decline in renal function.
View details for Web of Science ID 000233204300022
View details for PubMedID 16316342
Expression and role of the hyaluronan receptor RHAMM in inflammation after bleomycin injury
AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY
2005; 33 (5): 447-454
Lung injury is associated with increased concentrations of hyaluronan (hyaluronic acid, HA). HA modifies cell behavior through interaction with cell-associated receptors such as receptor for HA-mediated motility (RHAMM, CD168). Using a function blocking anti-RHAMM antibody (R36), we investigated the expression and role of RHAMM in the inflammatory response to intratracheal bleomycin in rats. Immunostaining showed increased expression of RHAMM in macrophages 4-7 d after injury. Surface biotin labeling of cells isolated by lavage confirmed increased surface expression of a 70-kD RHAMM after lung injury, and in situ hybridization demonstrated increased RHAMM mRNA in macrophages responding to injury. Time-lapse cinemicrography demonstrated a 5-fold increase in motility of alveolar macrophages from bleomycin-treated animals that was completely blocked by R36 in vitro. Further, HA-stimulated macrophage chemotaxis was also inhibited by R36. Daily administration of R36 to injured animals resulted in a 40% decrease in macrophage accumulation 7 d after injury. Further, H&E staining of tissue sections showed that bleomycin-mediated changes in lung architecture were improved with R36 treatment. Taken together with previous results showing the inhibitory effects of HA-binding peptide on inflammation and fibrosis, we conclude that the interaction of RHAMM with HA is a critical component of the recruitment of inflammatory cells to the lung after injury.
View details for DOI 10.1165/rcmb.2004-0333OC
View details for Web of Science ID 000233181900005
View details for PubMedID 16037485
- Novel methods of prediction of long-term renal allograft outcome PEDIATRIC TRANSPLANTATION 2004; 8 (6): 531-532
Computerized image analysis vs semiquantitative scoring in evaluation of kidney allograft fibrosis and prognosis
NEPHROLOGY DIALYSIS TRANSPLANTATION
2004; 19 (11): 2838-2845
Chronic morphological changes in the kidney allograft predict long-term graft function, but there are few studies comparing different methods in assessing chronic lesions. In the present study, we evaluated allograft cortical interstitial fibrosis, and compared semiquantitative assessment with computerized image analysis of Sirius red-stained collagen in prediction of graft prognosis.Sections were obtained from a series of 1-year protocol living donor kidney graft biopsies (n = 33) and their corresponding baseline specimens (n = 32). At light microscopy, the biopsies were scored for interstitial fibrosis as a percentage of involved tubulointerstitium according to the Banff schema. Quantitation of cortical fractional interstitial fibrosis volume (Vint) was performed with computerized image analysis on coded sections stained with Sirius red. The results were correlated with kidney function at 8-10 years after transplantation, and with late graft loss.There was a significant correlation between the semiquantitative and quantitative methods for measuring cortical interstitial fibrosis in all the biopsies (n = 65, percentage area vs Vint: R = 0.439, P = 0.0003). The correlation further improved when analysing the baseline specimens separately (n = 32, R = 0.704, P<0.0001) and was still significant, but less precise for the 1-year biopsies (n = 33, R = 0.384, P = 0.0274). One-year semiquantitative fibrosis (percentage area) was correlated to serum creatinine at 8-10 years (P = 0.010) and to late graft loss (P = 0.0445). The 1-year Vint values for interstitial fibrosis showed a similar trend but did not reach statistical significance in prediction of long-term graft function.Image analysis quantitation of interstitial collagen with Sirius red corresponded well to light microscopic semiquantitative assessment of interstitial fibrosis. In prediction of long-term graft function, the semiquantitative method was superior, indicating that accumulation of matrix molecules other than fibrillary collagens, oedema and inflammation are also important in graft prognosis.
View details for Web of Science ID 000225115400024
View details for PubMedID 15385637
Dietary phosphorus reduction by pretreatment of human breast milk with sevelamer
2004; 19 (7): 775-779
Hyperphosphatemia leading to hyperparathyroidism and ultimately renal osteodystrophy is a well-known complication of chronic renal failure. A new hydrogel binder, sevelamer, has recently become available for use in hyperphosphatemic patients with renal failure. We had previously mixed the capsule with pumped breast milk and formula, but discovered that the hydrogel formed a viscous solution that infants were unable or unwilling to swallow. We therefore evaluated the phosphorus content of fresh and frozen breast milk before and after treating with different doses of sevelamer at different temperatures and for varying lengths of time. The hydrogel bound promptly to phosphorus, reducing the phosphorus content 78% within 5 min. The viscous hydrogel settled to the bottom of the container within 10 min allowing the supernatant to be easily decanted. We also evaluated the breast milk for changes in other electrolytes, osmolality, pH, and macronutrient content. These results show that fresh or frozen breast milk can be safely pretreated with sevelamer without significantly changing its macronutrient or ionic content, with the exception of calcium and protein. The supernatant can be fed to infants or instilled through a gastrostomy tube without difficulty since the viscous hydrogel settles rapidly to the bottom of the container.
View details for DOI 10.1007/s00467-004-1448-6
View details for Web of Science ID 000221752000014
View details for PubMedID 15103549
- The protocol renal allograft biopsy: Has its time come? PEDIATRIC TRANSPLANTATION 2004; 8 (1): 3-5
Mycobacterial peritonitis in pediatric peritoneal dialysis patients
2004; 19 (1): 114-117
Peritonitis is the most common complication and the leading cause of death in pediatric peritoneal dialysis (PD) patients. According to the most recent data available from the North American Pediatric Renal Transplant Cooperative Study (NAPRTCS), approximately 25% of pediatric PD patients who die succumb to infection. There are no reported cases of Mycobacterium tuberculosis (MTB) or Mycobacterium avium-intracellulare peritonitis in the NAPRTCS registry. With an increasing incidence of MTB worldwide and the impairment of cellular immunity in chronic renal failure patients, it is not surprising that mycobacterium peritonitis can occur in PD patients. We report two pediatric PD patients with mycobacterial peritoneal infection diagnosed over an 11-year period at our institution. One patient presented with a malfunctioning Tenckhoff catheter and again 3 years later with hyponatremia and ascites. The other presented with recurrent culture-negative peritonitis. These cases illustrate the importance of more extensive evaluation of PD complications, to include evaluation for mycobacterium with special media or peritoneal biopsy, in the above clinical settings if the routine work-up is unrevealing.
View details for DOI 10.1007/s00467-003-1348-1
View details for Web of Science ID 000188455400021
View details for PubMedID 14648331
Computer-assisted quantification of fibrosis in chronic allograft nephropaty by picosirius red-staining: A new tool for predicting long-term graft function
2003; 76 (6): 955-958
Chronic allograft nephropathy (CAN) has become the predominant limiting factor for long-term transplant survival. A cardinal histomorphologic correlate for CAN is interstitial fibrosis. Currently, no method has been established in routine use that reliably quantifies the extent of interstitial fibrosis in renal grafts. We have used staining with picrosirius red followed by computerized image analysis to study the correlation between graft fibrosis and future development of glomerular filtration rate (GFR) in a group of children with advanced CAN.Renal biopsies were performed in 56 children (mean age, 13.7+/-3.6 years) after a mean period of 4.6+/-3.1 years after transplantation because of significant increases in serum creatinine. All biopsy specimens were stained with picrosirius red. The magnitude of fibrotic tissue was calculated by computerized image analysis. Linear regression analysis was performed correlating the intensity of graft fibrosis and the changes in the GFR at the time points of renal biopsy and 2 years later.There was a significant positive correlation (r=0.62, P<0.001) between the picrosirius red-stained cortical fractional interstitial fibrosis volume (V(intFib)) and the decrease of GFR within 2 years postrenal biopsy. When V(intFib) was below 5%, 82% of the patients had an increase in GFR within 2 years. Ninety-three percent of the patients with greater than 10% of fibrosis experienced a worsening renal function after 2 years. When comparing patients with stable GFR with patients having a decrease in GFR, a highly significant difference in V(intFib) was found (P=0.008).The quantitative measurement of fibrosis by picrosirius red staining appears to be a useful prognostic indicator for estimating long-term graft function in CAN and may provide an easy, fast, and inexpensive tool helpful for treatment decisions in patients developing CAN.
View details for DOI 10.1097/01.TP.0000078899.62040.E5
View details for Web of Science ID 000185637900013
View details for PubMedID 14508360
Cell senescence in rat kidneys in vivo increases with growth and age despite lack of telomere shortening
2003; 63 (6): 2134-2143
Somatic cells in vitro have a finite life expectancy before entering a state of senescence, but it is unclear whether this state occurs in vivo in kidney development, growth, and aging. We previously showed that human kidney cortex displays telomere shortening with age. In this study, we compared the structural and functional changes in rat kidney with age to phenomena associated with cellular senescence in vitro.We assessed the changes in Fischer 344 rat kidneys from age 1 to 9 months to define growth and development and from age 9 to 24 months to define aging.Rat kidney telomeres were approximately 35 to 40 kb long and did not shorten significantly. Expression of mRNA for p16INK4a, a characteristic senescence gene in vitro, was undetectable in most young rats but rose 27 fold during growth and a further 72-fold during aging. p16INK4a protein was localized to the nucleus and increased with age. p16INK4a mRNA also increased in other tissues. Lipofuscin and senescence-associated beta-galactosidase increased in epithelium with growth and aging and their occurrence was significantly associated with each other. Lipofuscin was particularly found in atrophic nephrons.We conclude that cell senescence occurs in both growth and aging in rat kidney and may contribute to the age-related pathology. These changes are not due to telomere shortening, but may reflect cumulative environmental stress.
View details for Web of Science ID 000182781900017
View details for PubMedID 12753300
Computerized image analysis of sirius red-stained renal allograft biopsies as a surrogate marker to predict long-term allograft function
JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY
2003; 14 (6): 1662-1668
Chronic allograft nephropathy (CAN) is a major problem in posttransplant management. The lack of a reliable and early surrogate marker of CAN has hampered patient care and research. In this study, the Cortical Fractional Interstitial Fibrosis Volume (V(IntFib)), quantitated with computerized image analysis of Sirius Red-stained protocol biopsies, was examined as a potential surrogate for time to graft failure (TTGF) in 68 renal allograft recipients. At 6 mo posttransplant, V(IntFib) was highly correlated with TTGF (r = 0.64, P < 0.001). Both the Banff Chronic Sum and the Acute Sum Scores were also correlated with TTGF, but less strongly (r = 0.28, P < 0.02; r = 0.35, P < 0.003, respectively). As V(IntFib) was not correlated with the Banff Chronic Score, a multivariate model was created that incorporated V(IntFib) and both Acute and Chronic Banff pathology. This model was highly correlated with TTGF (r = 0.7, P < 0.0001). These findings suggest that V(IntFib) determined by computerized image analysis of Sirius Red-stained protocol biopsies at 6 mo posttransplant, with or without incorporation of Banff acute and chronic scoring, may provide an early surrogate for time to graft failure in renal allograft recipients.
View details for DOI 10.1097/01.ASN.0000066143.02832.5E
View details for Web of Science ID 000183095600029
View details for PubMedID 12761269
Neointimal and tubulointerstitial infiltration by recipient mesenchymal cells in chronic renal-allograft rejection.
NEW ENGLAND JOURNAL OF MEDICINE
2001; 345 (2): 93-97
Tissue remodeling depends on mesenchymal cells (fibroblasts and myofibroblasts) and is a prominent feature of chronic renal-transplant rejection. It is not known whether the mesenchymal cells that participate in remodeling originate locally or from circulating precursor cells.We obtained biopsy specimens of renal allografts from six male recipients of an allograft from a female donor, four female recipients of an allograft from a male donor, two male recipients of an allograft from a male donor, and two female recipients of an allograft from a female donor. All the allografts were undergoing chronic rejection. All but two specimens were obtained within six months after transplantation. We used immunohistochemical methods to identify mesenchymal cells with smooth-muscle alpha-actin and in situ hybridization to identify mesenchymal cells with Y-chromosome DNA.No Y-chromosome bodies were identified in the case of the two renal-allograft specimens in which both the donor and the recipient were female. In the case of the two renal-allograft specimens in which both the donor and the recipient were male, approximately 40 percent of mesenchymal cells contained a Y-chromosome body. In the case of the six specimens in which the donor was female and the recipient was male, a mean (+/-SD) of 34+/-16 percent of mesenchymal cells in the neointima, 38+/-12 percent of such cells in the adventitia, and 30+/-7 percent of such cells in the interstitium contained the Y-chromosomal marker, indicating that they originated from the recipient rather than the donor. In the case of the four renal-allograft specimens in which the donor was male and the recipient was female, the respective values were 24+/-15 percent, 33+/-9 percent, and 23+/-8 percent, indicating a persistent population of donor mesenchymal cells.The presence of mesenchymal cells of host origin in the vascular and interstitial compartments of renal allografts undergoing chronic rejection provides evidence that a circulating mesenchymal precursor cell has the potential to migrate to areas of inflammation.
View details for Web of Science ID 000169776900003
View details for PubMedID 11450677
Transjugular intrahepatic portosystemic shunt prior to renal transplantation in a child with autosomal-recessive polycystic kidney disease and portal hypertension: A case report
2001; 5 (3): 210-214
Autosomal-recessive polycystic kidney disease (ARPKD) can cause renal failure and portal hypertension in children. Portal hypertension may complicate the course of renal transplantation (Tx). We report the successful outcome of a patient with end-stage renal disease (ESRD) and portal hypertension treated with transjugular intrahepatic portosystemic shunt (TIPS), a minimally invasive endovascular technique of portosystemic shunt, prior to renal Tx.
View details for Web of Science ID 000169342500012
View details for PubMedID 11422825
Neoral pharmacokinetics in Latino and Caucasian pediatric renal transplant recipients
2001; 16 (4): 311-314
Interpopulation variability of drug pharmacokinetics (PK) has been described. For example, the systemic clearance of nifedipine is higher in Mexicans than Caucasians. African-Americans have a lower cyclosporine bioavailability than Caucasians. Limited data are available in the Latino population. Under identical conditions, we compared the PK profile of Neoral (cyclosporine for microemulsion) obtained in stable pediatric renal transplant recipients of Latino and Caucasian origin. A slightly lower area under the curve (AUC) when corrected for dose per body surface area or per kilogram of body weight was observed in Caucasians compared with Latinos. This difference was more pronounced in the younger age group (< 12 years) with a higher peak-to-trough ratio. However, the Caucasians required a higher dosage of Neoral than the Latinos to achieve that same AUC. There was no difference between the groups in the time (tmax) to reach maximal concentration (Cmax) of Neoral. A higher apparent clearance of the drug was observed in the Caucasians compared with the Latinos, especially in the younger age group. No differences in pre- and post-dose levels were observed between the two groups. These differences might affect dose adjustment between the two subpopulations.
View details for Web of Science ID 000168399200001
View details for PubMedID 11354772
A role for hyaluronan in macrophage accumulation and collagen deposition after bleomycin-induced lung injury
AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY
2000; 23 (4): 475-484
Elevated concentrations of hyaluronan (HA) are associated with the accumulation of macrophages in the lung after injury. We have investigated the role of HA in the inflammatory and fibrotic responses to lung injury using the intratracheal instillation of bleomycin in rats as a model. After bleomycin-induced lung injury, both HA content in bronchoalveolar lavage (BAL) and staining for HA in macrophages accumulating in injured areas of the lung were maximal at 4 d. Increased HA in BAL correlated with increased locomotion of isolated alveolar macrophages. HA-binding peptide was able to specifically block macrophage motility in vitro. Importantly, systemic administration of HA-binding peptide to rats before injury not only decreased alveolar macrophage motility and accumulation in the lung, but also reduced lung collagen alpha (I) messenger RNA and hydroxyproline contents. We propose a model in which HA plays a critical role in the inflammatory response and fibrotic consequences of acute lung injury.
View details for Web of Science ID 000089888300009
View details for PubMedID 11017912
- Interstitial nephritis in children with Crohn's disease CLINICAL PEDIATRICS 2000; 39 (4): 253-254
Quantitation of allograft fibrosis and chronic allograft nephropathy.
1999; 3 (4): 257-270
Despite improvements in the prevention and treatment of acute renal allograft rejection, the long-term survival of renal transplants has not increased. Immunologic and non-immunologic factors contribute to the gradual deterioration of graft function and to the histologic lesion characterized by vascular and interstitial fibrosis ('chronic rejection'). Quantitation of this process has been attempted using various invasive and non-invasive methods. These methods, performed at different times post-transplant, are reviewed in this article. In particular, pathology scoring systems and the potential of using computerized image analysis of biopsy material are discussed.
View details for PubMedID 10562970
Effect of increasing baseline immunosuppression on the prevalence of clinical and subclinical rejection: A pilot study
JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY
1999; 10 (8): 1801-1805
This group has reported that treatment of subclinical rejection in the first 3 mo posttransplant with corticosteroids decreases late clinical rejections and improves graft function at 2 yr in renal transplant recipients. The current study was performed to determine whether an increase in baseline immunosuppression would decrease the prevalence of early subclinical rejections, as well as the incidence of early and late clinical rejections. Patients received mycophenolate mofetil (MMF) and Neoral cyclosporin A (CsA) posttransplant (n = 29), of which 17 underwent protocol biopsies at months 1, 2, 3, and 6 (Neoral + MMF Protocol Biopsy [Bx]), while 12 declined protocol biopsies (Neoral + MMF Control). These individuals were compared with 72 historical control patients treated with Sandimmune CsA and Imuran, of which 36 had undergone protocol biopsies at months 1, 2, 3, and 6 (Sandimmune + Azathioprine [AZA] Protocol Bx), and 36 had a protocol biopsy at month 6 (Sandimmune + AZA Control). Baseline immunosuppression with Neoral + MMF decreased the incidence of early clinical rejections (0 to 3 mo) and cumulative corticosteroid exposure, but had no impact on the prevalence of early subclinical rejection. Moreover, to maximally decrease the risk of developing late clinical rejections (months 7 to 12) in Neoral + MMF patients required that protocol biopsies be done and that subclinical rejection be treated. The paradoxical finding of recent clinical trials that a reduction in acute clinical rejection has not improved long-term graft outcome may be explained in part by the failure to control subclinical rejection.
View details for Web of Science ID 000081721300020
View details for PubMedID 10446949
Immune-activation gene expression in clinically stable renal allograft biopsies - Molecular evidence for subclinical rejection
1998; 66 (12): 1673-1681
A significant percentage of biopsies from stable, well-functioning renal allografts have histologic findings consistent with acute rejection or borderline rejection. The implication of this finding is not yet fully understood. We analyzed immune-activation gene transcripts in stable protocol biopsies to determine the extent of immunologic activity of graft-infiltrating cells in this setting. Histologic classification of the biopsies was based on the Banff criteria. To emphasize that the tissue samples were procured from grafts with no clinical evidence of impaired function, we interjected the term "subclinical" into the Banff terminology. This produced three histologic categories: normal, borderline subclinical rejection, and acute subclinical rejection.We used competitive template polymerase chain reaction techniques to quantify transcript amounts for the constant region of the T-cell receptor beta chain; the cytokines, tumor necrosis factor alpha, interleukin (IL)-1beta, transforming growth factor beta, interferon gamma, IL-2, IL-4, IL-10, and IL-15; and the cytotoxic T lymphocyte effector molecules, granzyme B, perforin, and Fas ligand.We found that histologically normal biopsies were typically devoid of gene transcripts or had very low amounts. Conversely, biopsies with acute subclinical rejection by histologic examination had heightened amounts of transcripts for many of the genes assayed. Borderline subclinical rejection samples showed an intermediate amount of expression.These results demonstrate that histologic features of rejection are often accompanied by enhanced expression of pro-inflammatory gene transcripts, despite the absence of clinically overt graft dysfunction. As this state of subclinical rejection could prove detrimental to long-term graft function, a role for surveillance biopsies of stable grafts with intent to treat subclinical rejection should be considered.
View details for Web of Science ID 000077958500018
View details for PubMedID 9884258
Protocol biopsies in renal transplantation: research tool or clinically useful?
CURRENT OPINION IN NEPHROLOGY AND HYPERTENSION
1998; 7 (6): 691-694
Early protocol biopsies of stable, well functioning renal allografts reveal a high prevalence of clinically unsuspected acute and chronic pathology. It is becoming increasingly apparent that these histopathological findings are both pathogenic and predictive of long-term allograft outcome. Therefore, protocol biopsies may be required for optimal post-transplant surveillance until non-invasive methods to detect allograft inflammation are developed.
View details for Web of Science ID 000077477000012
View details for PubMedID 9864667
Beneficial effects of treatment of early subclinical rejection: A randomized study
JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY
1998; 9 (11): 2129-2134
The prevalence of subclinical rejection, by the Banff criteria, is approximately 30% in the first 3 mo in renal transplant recipients. A randomized study was performed to determine whether the treatment of subclinical rejection with corticosteroids was associated with improved outcomes in these patients. Seventy-two patients, stratified by donor source, were randomized to biopsies at 1, 2, 3, 6, and 12 mo (Biopsy group), or to 6- and 12-mo biopsies only (Control group). Patients were analyzed by "intent to treat" and were followed for a minimum of 2 yr. Patients in the Biopsy arm of the study had a significant decrease in early (months 2 and 3) and late (months 7 to 12) acute rejection episodes, a reduced chronic tubulointerstitial score at 6 mo, and a lower serum creatinine at 24 mo than did patients in the Control arm. There was a trend toward an increase in infectious morbidity, but no increase in mortality, in the patients randomized to the Biopsy group. The results of this study suggest that early protocol biopsies and the treatment of subclinical rejection with corticosteroids may lead to better histologic and functional outcomes in renal transplant recipients.
View details for Web of Science ID 000076547400020
View details for PubMedID 9808101
Matching for private or public HLA epitopes reduces acute rejection episodes and improves two-year renal allograft function
1998; 66 (1): 38-43
The current role of HLA matching in renal transplantation is controversial. Public HLA epitope matching has been suggested to be as advantageous as private HLA matching, with the added benefit of increasing recipients' access to well-matched grafts.In this single-center study of 105 renal transplant recipients, we examined the association of HLA matching with early (0-3 months) and late (4-6 months) rejection episodes (RE), as well as renal allograft function up to 2 years after transplant.Poor HLA-DR, but not HLA-A or -B, matching was associated with early RE (0 DR matches, RE=2.7+/-0.19, 1 DR match, RE=2.37+/-0.18, vs. 2 DR matches, RE=1.5+/-0.38; P < 0.01). In contrast, poor HLA-B, but not HLA-A or -DR, matching was associated with late rejections (0 HLA-B matches, RE=1.1+/-0.51 vs. 1-2 HLA-B matches, RE=0.51+/-0.1; P < 0.004). HLA-B matching was also associated with a significantly lower serum creatinine (SCr) level at 24 months (0 HLA-B matches, SCr=178+/-20 micromol/L vs. SCr=132+/-6 micromol/L for 1-2 HLA-B matches; P < 0.025). Matching for 10 supertypic HLA-A and -B cross-reactive groups was associated with reduced late graft rejection (0-2 residue matches, RE=1.15+/-0.18 vs. RE=0.62+/-0.12 for 3 to 7 residue matches; P < 0.013) as well as a significantly lower SCr level at 24 months (0-2 residue matches, SCr=205+/-29 micromol/L vs SCr=131+/-6 micromol/L for 3 to 7 residue matches; P < 0.001) after transplantation.HLA-DR matching was associated with a reduced frequency of early rejection episodes, whereas HLA-B or residue/cross-reactive group matching was associated with a reduced frequency of late rejection episodes and improved graft function at 2 years.
View details for Web of Science ID 000074889300006
View details for PubMedID 9679819
Nonradioactive Northern blotting with biotinylated and digoxigenin-labeled RNA probes
1998; 19 (8-9): 1351-1355
The application of nonradioactive RNA probes for Northern blotting offers the advantage of a rapid turn-around time for results without the loss of sensitivity for target mRNA detection. However, a problem that has impeded the widespread use of nonradioactive RNA probes for use in Northern blotting is the difficulty in stripping these probes from nylon membranes after hybridization. In this report we describe two protocols for stripping digoxigenin (Dig)-labeled RNA probes from nylon membranes. One protocol utilizes a phosphate-buffered formamide stripping solution to remove nonchemically modified (regular) RNA probes while the other method utilizes strippable probes that were produced with a chemically modified nucleotide (CTP) and removed by a specific stripping solution. This latter method was developed by Ambion Inc. and is called Strip-EZ. We also describe a protocol for the detection of two separate rat mRNAs using both biotin and digoxigenin-labeled RNA probes that does not require stripping the membrane after hybridization. Finally, we describe the use of another new labeling technology, called Chem-Link, that quickly and conveniently labels RNA for use in Northern blotting.
View details for Web of Science ID 000074652100025
View details for PubMedID 9694280
Production of digoxigenin-labelled RNA probes and the detection of cytokine mRNA in rat spleen and brain by in situ hybridization
BRAIN RESEARCH PROTOCOLS
1998; 2 (4): 339-351
Non-radioactive in situ hybridization is a sensitive method for determining the site of production for secretory molecules such as cytokines. We report here on the central and peripheral induction of proinflammatory cytokines by endotoxin, and outline procedures for the generation and application of rat-specific digoxigenin (Dig)-labelled RNA probes for the localization of mRNA by in situ hybridization. Rats were injected either intravenously (i.v.) or intracerebroventricularly (i.c.v.) with vehicle or lipopolysaccharide (LPS) and sacrificed at various time intervals post-injection. Rats were then perfused with 4% paraformaldehyde and the spleens and brains were removed and cryoprotected in 30% sucrose. Dig-labelled, rat-specific, antisense and sense RNA probes were generated by in vitro transcription from PCR-derived templates. Positive staining with all the antisense probes was cytoplasmic, whereas the sense probes showed no staining. Numerous tumor necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1beta) mRNA positive cells were observed in the marginal zone and in the red pulp of the spleen after iv LPS injections, whereas sections from saline-treated animals showed minimal cytokine mRNA expression. Cells positive for TNF-alpha and IL-1beta mRNA were detectable in the brain after i.c.v. injections of LPS, but not after icv injection of vehicle. An antisense probe for c-fos was utilized in these studies as a positive control for our procedure due to its anatomically specific expression in the rat brain after LPS. In conclusion we have demonstrated that in situ hybridization with Dig-labelled RNA probes is an efficient, sensitive and reliable tool to localize cytokine mRNA production in rat tissue.
View details for Web of Science ID 000074668100013
View details for PubMedID 9630715
Pediatric renal transplantation: indications and special considerations. A position paper from the Pediatric Committee of the American Society of Transplant Physicians.
1998; 2 (2): 117-129
Renal transplantation of children with chronic renal insufficiency (CRI) and end-stage renal disease (ESRD) appears to be the optimal form of renal replacement therapy. This report, which expresses the opinions of the nephrology members of the Pediatric Committee of the American Society of Transplant Physicians, discusses the indications for pediatric renal transplantation and identifies the unique aspects of caring for children with CRI and ESRD. Indications for pediatric renal transplantation include: 1) symptoms of uremia not responsive to standard therapy; 2) failure to thrive due to limitations in total caloric intake; 3) delayed psychomotor development; 4) hypervolemia; 5) hyperkalemia; and 6) metabolic bone disease due to renal osteodystrophy. The urgency and timing of renal transplantation in children must be considered in the context of a number of issues unique to children with CRI and ESRD such as delayed cognitive and educational performance, growth retardation, delayed puberty, etiology of ESRD, and timing of immunizations. In addition, these children frequently display various inherited and sporadic syndromes with multiorgan involvement requiring the expertise of a variety of pediatric subspecialists including the pediatric urologist, who plays a critical role in the evaluation of children with obstructive uropathy and other anomalies of the genito-urinary system. The advantages of a living-related donor are also delineated. The importance of adequate immunosuppression on graft function, early recognition of the signs and symptoms acute rejection, preventive strategies for minimizing the morbidity and mortality from viral infections in the post-transplant period, and the impact of transplantation on cognitive function, educational status, and catch-up growth are also discussed. To address these complex issues, transplant care of pediatric patients must be provided by a multidisciplinary team of pediatric health care professionals.
View details for PubMedID 10082443
Identification of clinical and histopathologic risk factors for diminished renal function 2 years posttransplant
JOURNAL OF THE AMERICAN SOCIETY OF NEPHROLOGY
1998; 9 (3): 482-487
The aim of this study was to identify early clinical and pathologic variates that independently predict diminished renal allograft function at 24 mo posttransplant. A clinical pathologic data base was prospectively derived from 71 patients in whom protocol renal biopsies were performed at 1, 2, 3, 6, and 12 mo posttransplant. The major end point was the 24-mo serum creatinine. Variates correlating independently (r2 = 0.67) with the 24-mo serum creatinine were the chronic biopsy scores (months 3 and 6), late rejections (months 4 to 6), cyclosporin A (CsA) levels (months 1 to 2), and delayed graft function. The adjusted odds ratio (OR) and 95% confidence interval (CI) for having a serum creatinine > or = 130 mumol/L at 24 mo increased for every year the donor age increased (OR = 1.07; 95% CI, 1.02 to 1.13; range, 9 to 55) or for each late rejection episode (OR = 5.9; 95% CI, 1.7 to 20.1), whereas a mean CsA level > 300 micrograms/L from months 1 to 3 was protective (OR = 0.07; 95% CI, 0.01 to 0.43). Variates correlating independently (r2 = 0.53) with the change in serum creatinine from 6 to 24 mo (delta Cr6-24) were the chronic biopsy scores at months 3 and 6. The adjusted OR of the delta Cr6-24 rising > or = +20 mumol/L increased for every year the donor age increased (OR = 1.09; 95% CI, 1.02 to 1.16; range 9 to 56) or when the 6-mo chronic biopsy score was > or = 2 (OR = 6.6; 95% CI, 1.2 to 36.4). An estimate of the relative risk for diminished renal function at 2 yr can be assigned within 6 mo of transplant based on chronic pathology, late acute rejections, CsA levels, and donor age.
View details for Web of Science ID 000072214900018
View details for PubMedID 9513912
Enhanced immunohistochemical detection of autonomic nerve fibers, cytokines and inducible nitric oxide synthase by light and fluorescent microscopy in rat spleen
JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY
1997; 45 (4): 599-610
We have developed enhanced immunohistochemical protocols for detecting autonomic nerve fibers and splenocyte-associated proteins in rat spleen. This includes norepinephrine-synthesizing enzymes (dopamine-beta hydroxylase (DBH) and tyrosine hydroxylase (TH)), neuropeptide Y (NPY), tumor necrosis factor-alpha (TNF-alpha), interferon-gamma (IFN-gamma), c-fos protein, inducible nitric oxide synthase (iNOS), and the macrophage cell marker ED1. Animals were divided into sham-operated and splenic nerve-sectioned groups for detection of DBH, TH, and NPY. For immunodetection of TNF-alpha, iNOS, IFN-gamma and c-fos, animals were injected IV with saline or 100 microg of lipopolysaccharide (LPS) and were sacrificed at various time intervals post injection. Rats were perfused with 4% paraformaldehyde, spleens removed and cryoprotected, and 50-microm floating sections were cut on a freezing microtome. Immunodetection was performed with various detection systems and substrate/chromogen solutions, and in some cases using pretreatment with proteinase K (PK) for antigen unmasking. PK pretreatment increased immunostaining for DBH, TH, NPY, IFN-gamma, iNOS, and ED1, and the improvement was concentration-dependent. Using NPY immunostaining to index the signal-to-noise ratio for various substrates and detection systems, we found that an alkaline phosphatase detection system with NBT/BCIP as a substrate was the best procedure for light microscopy, whereas the CY3-labeled secondary antibody technique proved optimal for fluorescent microscopy. Surgical transection of the splenic nerve eliminated all nerve fiber staining for DBH, TH, and NPY. TNF-alpha, IFN-gamma, c-fos, and iNOS proteins were observed in the spleen in a time-dependent manner after LPS stimulation. Fluorescent double labeling, visualized with fluorescent confocal scanning laser microscopy, revealed many NPY fibers distributed among the ED1-labeled macrophages. These results demonstrate that immunohistochemistry can be used to index the activational effects of an immune challenge on splenocytes in situ and verifies that splenic immune cells are innervated by the sympathetic nervous system.
View details for Web of Science ID A1997WU66200012
View details for PubMedID 9111238
- Histamine H-1-receptors in the nasal mucosa: A mystery solved? CLINICAL AND EXPERIMENTAL ALLERGY 1996; 26 (4): 368-370
- HEMOLYTIC-UREMIC SYNDROME - THE MOST COMMON-CAUSE OF ACUTE-RENAL-FAILURE IN CHILDHOOD PEDIATRIC ANNALS 1994; 23 (9): 505-511
RAPID NONRADIOACTIVE IN-SITU HYBRIDIZATION FOR INTERLEUKIN-2 MESSENGER-RNA WITH RIBOPROBES GENERATED USING THE POLYMERASE CHAIN-REACTION
JOURNAL OF IMMUNOLOGICAL METHODS
1994; 167 (1-2): 83-89
In situ hybridization is a technique with widespread application. However, its usefulness has been limited by the need for radioactive materials and the requirement for the DNA to be cloned onto an appropriate vector. We have utilized the polymerase chain reaction to directly incorporate a T7 RNA polymerase promoter sequence onto the cDNA for interleukin-2. Digoxigenin-labelled riboprobes were then synthesized using this PCR product as a template. The digoxigenin-labelled riboprobes were then used in non-radioactive in situ hybridization to detect messenger RNA for interleukin-2 in mitogen stimulated peripheral blood mononuclear cells. This methodology has the potential for widespread application in immunology and cytokine research.
View details for Web of Science ID A1994MU31400009
View details for PubMedID 8308289
RENAL TUBULE NA,K-ATPASE POLARITY IN GLUCOCORTICOID-INDUCED POLYCYSTIC KIDNEY-DISEASE
JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY
1993; 41 (4): 555-558
Cyst formation in polycystic kidney disease (PKD) involves proliferation of cyst lining epithelial and changes in trans-epithelial fluid and electrolyte transport. In vitro studies have suggested that mislocation of Na,K-ATPase to the apical tubular surface may be an important component of cyst fluid transport. We undertook in vivo studies of Na,K-ATPase location using the "threshold" murine model of glucocorticoid-induced PKD (GIPKD). Using histological, immunohistochemical, and densitometric techniques, we compared cyst formation and the cellular location of Na,K-ATPase in suckling C3H (low threshold for GIPKD) and DBA (high threshold) mice given an inducing dose of 200 mg/kg methylprednisolone acetate. As expected, C3H mice demonstrated greater cyst formation as measured by proportion of section area occupied by the tubule lumen (26.7% vs 15.5%; p < 0.001). Cyst formation was associated with increased Na,K-ATPase staining and increased apical Na,K-ATPase location. MPA treatment in C3H mice resulted in apical staining that exceeded basolateral staining (35.3% of reference window vs 29.8%; p < 0.001). The relatively GIPKD-resistant DBA mice did not show such change in Na,K-ATPase location. These immunohistochemical studies suggest a role for Na,K-ATPase in renal cyst formation.
View details for Web of Science ID A1993KT97300009
View details for PubMedID 8383715
PEDIATRIC RENAL BIOPSY IN THE AMBULATORY CARE ENVIRONMENT
1992; 6 (3): 311-312
The use of pediatric ambulatory care facilities to perform invasive procedures that have low morbidity is increasingly popular. Over a 2-year period, 46 pediatric renal biopsies were performed in an ambulatory care setting at the Winnipeg Children's Hospital, with the patient discharged the same day. There was no serious complications and adequate tissue was obtained in 45 cases. Renal biopsy may be safely performed on an outpatient basis on carefully selected patients by experienced operators in properly equipped facilities.
View details for Web of Science ID A1992HU49200022
View details for PubMedID 1616846
Pediatric renal transplantation.
Advances in pediatrics
1992; 39: 441-493
View details for PubMedID 1442319
- RENAL-TRANSPLANTATION IN CHILDREN PEDIATRIC ANNALS 1991; 20 (12): 657-667
THE IMPACT OF RECOMBINANT-HUMAN-ERYTHROPOIETIN THERAPY ON RENAL-TRANSPLANTATION
AMERICAN JOURNAL OF KIDNEY DISEASES
1991; 18 (4): 57-61
This report describes the potential and actual effects that recombinant human erythropoietin (rHuEpo) may have on the practice of renal transplantation. Three aspects are highlighted. The first is the effects in the dialysis patient transplanted after treatment with rHuEpo. These include the potential risks of graft thrombosis and prolonged initial nonfunction (for which there is little supportive evidence), and the impact on pretransplant immune-modulating regimens, which take advantage of the so-called transfusion effect. As the importance of this effect to overall graft survival has diminished strikingly, this may be of little consequence. The second aspect relates to the highly presensitized dialysis patient. The literature and our own data are presented, showing the beneficial effects of rHuEpo therapy on reducing the level of humoral anti-HLA sensitization. This may lead to benefits that include reduced time on the waiting list for a cadaveric renal transplant, and possibly improved allograft survival. Finally, our data on the use of rHuEpo in 13 patients with anemia (usually due to chronic graft failure) after transplantation is discussed. rHuEpo therapy was effective in all patients, leading to reversal of anemia. Side effects, including hypertension and hypertensive seizures, occurred in the subgroup of patients with significant renal dysfunction (serum creatinine greater than 2.5 mg/DL).
View details for Web of Science ID A1991GK19700011
View details for PubMedID 1928081
SENSITIZED LYMPHOCYTES-B CONTRIBUTE TO ACUTE ALLOGRAFT-REJECTION
JOURNAL OF SURGICAL RESEARCH
1991; 51 (3): 204-209
The contribution of sensitized B lymphocytes to second-set allograft rejection has been relatively ignored despite their regular appearance in rejecting allografts. This study presents evidence that adoptively transferred sensitized B lymphocytes accelerate the rate of acute allograft rejection in a sublethally irradiated rat cardiac allograft model. Donors of reconstituting B lymphocytes were sensitized with three consecutive ACI skin grafts. Transplantation of a heart from an ACI strain donor into a Lewis strain recipient (complete RT1 mismatch) results in rejection in 6.8 +/- 0.3 days. When the allograft donor and recipient are irradiated with 650 cGy prior to transplantation, rejection occurs at 31.5 +/- 3.0 days. Irradiated recipients reconstituted with 10(6) syngeneic sensitized splenic B cells reject their grafts in 20.1 +/- 2.0 days, while reconstitution with 10(6) unsensitized syngeneic B cells has no effect on the rate of rejection (P = 0.0007). These data strongly suggest that sensitized B lymphocytes have a marked accelerating effect on the tempo of allograft rejection.
View details for Web of Science ID A1991GD58000005
View details for PubMedID 1881134
- LONG-TERM RESULTS WITH CYCLOSPORINE IMMUNE SUPPRESSION IN PEDIATRIC CADAVER RENAL-TRANSPLANTATION TRANSPLANTATION PROCEEDINGS 1991; 23 (1): 1011-1012
EFFECTS OF RECOMBINANT-HUMAN-ERYTHROPOIETIN ON HLA SENSITIZATION AND CELL-MEDIATED-IMMUNITY
1990; 38 (1): 12-18
Highly presensitized patients wait longer for a renal allograft than unsensitized patients and have a poorer allograft survival rate. Repeated blood transfusions have been implicated in the induction and maintenance of sensitization. To determine the effect of recombinant human erythropoietin (rHuEPO) therapy on five transfusion dependent, highly sensitized adolescents on dialysis, we serially measured percentage panel reactive antibody (%PRA) levels, titers of identifiable discrete anti-HLA Class I antibody specificities, and non-specific indices of cellular immunity before and following initiation of rHuEPO therapy. Although four of the five patients had previously rejected at least one renal allograft, the removal of chronic antigenic stimulation from blood transfusions led to a marked reduction in anti-HLA antibody titers to recognizable private and public specificities (P less than 0.001) and a reduction of mean %PRA from 80% to 56% (P less than 0.05). Each patient demonstrated a reduction of two or more dilutions to at least two anti-HLA antibody specificities. A control group of five patients matched for age, transfusion dependence and sensitization status demonstrated no change during a comparable time interval. PHA responsiveness decreased significantly in the rHuEPO group whereas autologous and allogenic mixed lymphocyte response, spontaneous blastogenesis and T-cell subsets did not. These data indicate that in highly sensitized dialysis patients rHuEPO may lead to decreased sensitization, shorter waiting time on dialysis and possibly improved allograft survival rates.
View details for Web of Science ID A1990DK40000003
View details for PubMedID 2385080
THE MICROANATOMY OF THE INTRAHEPATIC BILE-DUCT IN POLYCYSTIC DISEASE - COMPARISON OF THE CPK MOUSE AND HUMAN
JOURNAL OF EXPERIMENTAL PATHOLOGY
1990; 71 (1): 119-131
The cpk mutation in mice produces a lethal recessive form of polycystic kidney disease (PKD) that, like human forms of the condition, is associated with an age-related incidence of hepatic cysts. Injection of plastic into the biliary tree of affected animals revealed that these cysts arise from focal dilatations of the epithelial lining that may enlarge to the point that they obstruct the bile ducts. This concept was supported by histological and scanning and electron microscopic studies. No evidence could be found of primary obstruction of the biliary tree. The same techniques were then employed in specimens of human liver from patients with both recessive (ARPKD) and dominantly inherited PKD (ADPKD). Similar abnormalities of the biliary tree were identified. These abnormalities were not found in control liver samples from patients without PKD. The liver of the patient with ADPKD also demonstrated many von Meyenburg complexes. These were related to some cyst development, but these complexes freely communicated with bile ducts, contrary to currently held opinion. We conclude that hepatic abnormalities in the cpk mouse and human PKD arise from changes in bile ducts that are analogous to the renal lesions.
View details for Web of Science ID A1990CQ84600014
View details for PubMedID 2310613
NIFEDIPINE, VERAPAMIL AND CYCLOSPORINE-A PHARMACOKINETICS IN CHILDREN
1989; 3 (3): 314-316
We report two paediatric renal transplant patients in whom interaction with the calcium channel blocking agent verapamil resulted in reduced cyclosporin A (CyA) elimination. Prior therapy with another calcium channel blocking agent, nifedipine, did not affect CyA pharmacokinetics. We speculate that verapamil reduced CyA elimination in children via an effect upon the microsomal oxidase system that is independent of calcium channel activity.
View details for Web of Science ID A1989AE67500017
View details for PubMedID 2702114
- INTERSTITIAL NEPHRITIS INDUCED BY CLOXACILLIN NEPHRON 1989; 51 (2): 285-286
Role of 24 Hour Ambulatory Blood Pressure Monitoring after Pediatric Renal Transplantation.
WILEY-BLACKWELL. 2013: 151-151
View details for Web of Science ID 000318240300387
Ethnic Differences in Rates of Peritubular Capillary Dropout in Post-Transplant Fibrosis Observed in Protocol Renal Allograft Biopsies.
WILEY-BLACKWELL. 2010: 341-341
View details for Web of Science ID 000275921702457
Clinical rejection is distinguished from subclinical rejection by increased infiltration by a population of activated macrophages
AMER SOC NEPHROLOGY. 1999: 1582-1589
It has been reported previously that one-third of protocol renal biopsies in asymptomatic, biochemically stable renal transplant recipients in the first 6 mo show unsuspected subclinical graft rejection (both infiltrate and tubulitis) and that subclinical rejection is a risk factor for chronic renal dysfunction. This study was performed to determine whether differences in phenotype or activation status of graft-infiltrating cells underlie these different manifestations of acute rejection. Biopsies with normal histology (n = 10), subclinical rejection (n = 13), and clinical rejection (n = 9) were studied using immunohistochemistry and computerized image analysis. Subclinical and clinical rejections had similar histologic Banff scores. Univariate analysis showed a trend for a higher infiltration with CD8+ (P = 0.053) and CD68+(P = 0.06) cells in clinical rejection. Of the activation markers studied (CD25, perforin, tumor necrosis factor-alpha), only allograft inflammatory factor-1+-activated macrophages were significantly (P = 0.014) increased in the infiltrate of clinical rejection biopsies. These data suggest that activated macrophages or their products are responsible for acute renal dysfunction associated with clinical rejection episodes.
View details for Web of Science ID 000081143900020
View details for PubMedID 10405215
- The receptor for hyaluronan-mediated motility is expressed in human renal allografts and is correlated with Banff chronic rejection scores ELSEVIER SCIENCE INC. 1997: 2603-2604
LOW-FREQUENCY OF INFILTRATING CELLS INTENSELY EXPRESSING T-CELL CYTOKINE MESSENGER-RNA IN HUMAN RENAL-ALLOGRAFT REJECTION
WILLIAMS & WILKINS. 1995: 579-584
Immunosuppressive drugs used in clinical transplantation block cytokine mRNA transcription in vitro, but clinical rejection episodes are common. An understanding of what cytokine message is transcribed would be helpful in determining what contributes to the success of immunosuppression and provide directions for further research aimed at targeting specific cytokines. Previous studies have examined cytokine mRNA in rejecting solid organ biopsies by the reverse transcriptase polymerase chain reaction (RT-PCR) with variable results. We used nonradioactive in situ hybridization with cytokine-specific riboprobes to determine the frequency of cells expressing cytokine mRNA in the allograft infiltrate. Kidney biopsies were obtained from patients receiving protocol biopsies and with clinical evidence of rejection. Fourteen biopsies with a pathologic diagnosis of rejection were studied. Eight showed no cytokine staining, 2 expressed IL-2, and 3 expressed IL-4 and IFN-gamma. The positive cells were present at a low frequency (mean 2, range 1-5 per 10 high-power fields). The proportion of kidney biopsies expressing detectable message for interleukin-2 (IL-2), interleukin-4 (IL-4), and interferon-gamma (IFN-gamma) by in situ hybridization were similar to those reported using RT-PCR. The novel finding is that these cytokines are expressed in a few strongly positive cells in the allograft infiltrate. The vast majority of infiltrating cells are negative. This suggests that either the biopsies were performed when cytokine message was not expressed at a high level or that in human allograft recipients the sustained expression of the cytokines IL-2, IL-4, and IFN-gamma may not be necessary for graft rejection.
View details for Web of Science ID A1995QK22500024
View details for PubMedID 7533348
CADAVER RENAL-TRANSPLANTATION IN CHILDREN - LONG-TERM IMPACT OF NEW IMMUNOSUPPRESSIVE STRATEGIES
MUNKSGAARD INT PUBL LTD. 1991: 197-203
View details for Web of Science ID A1991FH34400013
- RECOMBINANT HUMAN ERYTHROPOETIN DECREASES ANTI-HLA SENSITIZATION AND MAY IMPROVE RENAL-ALLOGRAFT OUTCOME - INVOLVEMENT OF ANTIIDIOTYPIC ANTIBODY ELSEVIER SCIENCE INC. 1991: 407-408
- HIGH CELLULAR ANTI-HLA IMMUNE RESPONSIVENESS ACCOMPANIES THE HIGHLY SENSITIZED (HIGH PRA) STATE IN DIALYSIS PATIENTS ELSEVIER SCIENCE INC. 1991: 405-406
CADAVER RENAL-TRANSPLANTATION IN CHILDREN - RESULTS WITH LONG-TERM CYCLOSPORINE IMMUNOSUPPRESSION
MUNKSGAARD INT PUBL LTD. 1990: 329-336
View details for Web of Science ID A1990EG10800004