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  • Soluble FLT1 Gene Therapy Alleviates Brain Arteriovenous Malformation Severity. Stroke Zhu, W., Shen, F., Mao, L., Zhan, L., Kang, S., Sun, Z., Nelson, J., Zhang, R., Zou, D., McDougall, C. M., Lawton, M. T., Vu, T. H., Wu, Z., Scaria, A., Colosi, P., Forsayeth, J., Su, H. 2017; 48 (5): 1420–23

    Abstract

    Brain arteriovenous malformation (bAVM) is an important risk factor for intracranial hemorrhage. Current therapies are associated with high morbidities. Excessive vascular endothelial growth factor has been implicated in bAVM pathophysiology. Because soluble FLT1 binds to vascular endothelial growth factor with high affinity, we tested intravenous delivery of an adeno-associated viral vector serotype-9 expressing soluble FLT1 (AAV9-sFLT1) to alleviate the bAVM phenotype.Two mouse models were used. In model 1, bAVM was induced in R26CreER;Eng(2f/2f) mice through global Eng gene deletion and brain focal angiogenic stimulation; AAV2-sFLT02 (an AAV expressing a shorter form of sFLT1) was injected into the brain at the time of model induction, and AAV9-sFLT1, intravenously injected 8 weeks after. In model 2, SM22αCre;Eng(2f/2f) mice had a 90% occurrence of spontaneous bAVM at 5 weeks of age and 50% mortality at 6 weeks; AAV9-sFLT1 was intravenously delivered into 4- to 5-week-old mice. Tissue samples were collected 4 weeks after AAV9-sFLT1 delivery.AAV2-sFLT02 inhibited bAVM formation, and AAV9-sFLT1 reduced abnormal vessels in model 1 (GFP versus sFLT1: 3.66±1.58/200 vessels versus 1.98±1.29, P<0.05). AAV9-sFLT1 reduced the occurrence of bAVM (GFP versus sFLT1: 100% versus 36%) and mortality (GFP versus sFLT1: 57% [12/22 mice] versus 24% [4/19 mice], P<0.05) in model 2. Kidney and liver function did not change significantly. Minor liver inflammation was found in 56% of AAV9-sFLT1-treated model 1 mice.By applying a regulated mechanism to restrict sFLT1 expression to bAVM, AAV9-sFLT1 can potentially be developed into a safer therapy to reduce the bAVM severity.

    View details for DOI 10.1161/STROKEAHA.116.015713

    View details for PubMedID 28325846

    View details for PubMedCentralID PMC5404983

  • Activation of Alpha-7 Nicotinic Acetylcholine Receptor Reduces Brain Edema in Mice with Ischemic Stroke and Bone Fracture. Molecular neurobiology Zou, D., Luo, M., Han, Z., Zhan, L., Zhu, W., Kang, S., Bao, C., Li, Z., Nelson, J., Zhang, R., Su, H. 2016: -?

    Abstract

    Stroke is an important risk factor for bone fracture. We showed previously that bone fracture at the acute stage of ischemic stroke worsens, and activation of α-7 nicotinic acetylcholine receptor (α-7 nAchR) improves, stroke recovery by attenuating inflammation. We hypothesized that activation of α-7 nAchR also improves the blood-brain barrier (BBB) integrity. Permanent distal middle cerebral artery occlusion (pMCAO) was performed on C57BL/6J mice followed by tibia fracture 1 day later. Mice were treated with 0.8 mg/kg PHA 568487 (PHA, α-7 nAchR-specific agonist), 6 mg/kg methyllycaconitine (MLA, α-7 nAchR antagonist), or saline 1 and 2 days after pMCAO. Brain water content, the expression of monoamine oxidase B (MAO-B), and tight junction protein (claudin-5) were assessed. We found that tibia fracture increased water content in the ischemic stroke brain (p = 0.006) and MAO-B-positive astrocytes (p < 0.001). PHA treatment reduced water content and MAO-B-positive astrocytes and increased claudin-5 expression in stroke and stroke + tibia fracture mice (p < 0.05), while MLA had the opposite effect. Our findings suggest that in addition to inhibiting inflammation, activation of α-7 nAchR also reduces brain edema, possibly through diminished astrocyte oxidative stress and improved BBB integrity. Thus, the α-7 nAchR-specific agonist could be developed into a new therapy for improving recovery of patients with stroke or stroke + bone fracture.

    View details for PubMedID 27914011

    View details for PubMedCentralID PMC5457363

  • BECN1/Beclin 1 sorts cell-surface APP/amyloid ß precursor protein for lysosomal degradation. Autophagy Swaminathan, G., Zhu, W., Plowey, E. D. 2016: 1-16

    Abstract

    The regulation of plasma membrane (PM)-localized transmembrane protein/receptor trafficking has critical implications for cell signaling, metabolism and survival. In this study, we investigated the role of BECN1 (Beclin 1) in the degradative trafficking of PM-associated APP (amyloid β precursor protein), whose metabolism to amyloid-β, an essential event in Alzheimer disease, is dependent on divergent PM trafficking pathways. We report a novel interaction between PM-associated APP and BECN1 that recruits macroautophagy/endosomal regulatory proteins PIK3C3 and UVRAG. We found that BECN1 promotes surface APP internalization and sorting predominantly to endosomes and endolysosomes. BECN1 also promotes the targeting of a smaller fraction of internalized APP to LC3-positive phagophores, suggesting a role for BECN1-dependent PM macroautophagy in APP degradation. Furthermore, BECN1 facilitates lysosomal degradation of surface APP and reduces the secretion of APP metabolites (soluble ectodomains, sAPP). The association between APP and BECN1 is dependent on the evolutionarily conserved domain (ECD) of BECN1 (amino acids 267-337). Deletion of a BECN1 ECD subregion (amino acids 285-299) did not impair BECN1- PIK3C3 interaction, PtdIns3K function or macroautophagy, but was sufficient to impair the APP-BECN1 interaction and BECN1's effects on surface APP internalization and degradation, resulting in increased secretion of sAPPs. Interestingly, both the BECN1-APP association and BECN1-dependent APP endocytosis and degradative trafficking were negatively regulated by active AKT. Our results further implicate phosphorylation of the BECN1 Ser295 residue in the inhibition of APP degradation by AKT. Our studies reveal a novel function for BECN1 in the sorting of a plasma membrane protein for endolysosomal and macroautophagic degradation.

    View details for PubMedID 27715386

    View details for PubMedCentralID PMC5173276

  • Persistent infiltration and pro-inflammatory differentiation of monocytes cause unresolved inflammation in brain arteriovenous malformation ANGIOGENESIS Zhang, R., Han, Z., Degos, V., Shen, F., Choi, E., Sun, Z., Kang, S., Wong, M., Zhu, W., Zhan, L., Arthur, H. M., Oh, S. P., Faughnan, M. E., Su, H. 2016; 19 (4): 451-461

    Abstract

    An abnormally high number of macrophages are present in human brain arteriovenous malformations (bAVM) with or without evidence of prior hemorrhage, causing unresolved inflammation that may enhance abnormal vascular remodeling and exacerbate the bAVM phenotype. The reasons for macrophage accumulation at the bAVM sites are not known. We tested the hypothesis that persistent infiltration and pro-inflammatory differentiation of monocytes in angiogenic tissues increase the macrophage burden in bAVM using two mouse models and human monocytes. Mouse bAVM was induced through deletion of AVM causative genes, Endoglin (Eng) globally or Alk1 focally, plus brain focal angiogenic stimulation. An endothelial cell and vascular smooth muscle cell co-culture system was used to analyze monocyte differentiation in the angiogenic niche. After angiogenic stimulation, the Eng-deleted mice had fewer CD68(+) cells at 2 weeks (P = 0.02), similar numbers at 4 weeks (P = 0.97), and more at 8 weeks (P = 0.01) in the brain angiogenic region compared with wild-type (WT) mice. Alk1-deficient mice also had a trend toward more macrophages/microglia 8 weeks (P = 0.064) after angiogenic stimulation and more RFP(+) bone marrow-derived macrophages than WT mice (P = 0.01). More CD34(+) cells isolated from peripheral blood of patients with ENG or ALK1 gene mutation differentiated into macrophages than those from healthy controls (P < 0.001). These data indicate that persistent infiltration and pro-inflammatory differentiation of monocytes might contribute to macrophage accumulation in bAVM. Blocking macrophage homing to bAVM lesions should be tested as a strategy to reduce the severity of bAVM.

    View details for DOI 10.1007/s10456-016-9519-4

    View details for Web of Science ID 000384411900001

    View details for PubMedID 27325285

    View details for PubMedCentralID PMC5029790

  • Bone Fracture Pre-Ischemic Stroke Exacerbates Ischemic Cerebral Injury in Mice PLOS ONE Wang, L., Kang, S., Zou, D., Zhan, L., Li, Z., Zhu, W., Su, H. 2016; 11 (4)

    Abstract

    Ischemic stroke is a devastating complication of bone fracture. Bone fracture shortly after stroke enhances stroke injury by augmenting inflammation. We hypothesize that bone fracture shortly before ischemic stroke also exacerbates ischemic cerebral injury. Tibia fracture was performed 6 or 24 hours before permanent middle cerebral artery occlusion (pMCAO) on C57BL/6J mice or Ccr2RFP/+Cx3cr1GFP/+ mice that have the RFP gene knocked into one allele of Ccr2 gene and GFP gene knocked into one allele of Cx3cr1 gene. Behavior was tested 3 days after pMCAO. Infarct volume, the number of CD68+ cells, apoptotic neurons, bone marrow-derived macrophages (RFP+), and microgila (GFP+) in the peri-infarct region were quantified. Compared to mice subjected to pMCAO only, bone fracture 6 or 24 hours before pMCAO increased behavioral deficits, the infarct volume, and the number of CD68+ cells and apoptotic neurons in the peri-infarct area. Both bone marrow-derived macrophages (CCR2+) and microglia (CX3CR1+) increased in the peri-infarct regions of mice subjected to bone fracture before pMCAO compared to stroke-only mice. The mice subjected to bone fracture 6 hours before pMCAO had more severe injury than mice that had bone fracture 24 hours before pMCAO. Our data showed that bone fracture shortly before stroke also increases neuroinflammation and exacerbates ischemic cerebral injury. Our findings suggest that inhibition of neuroinflammation or management of stroke risk factors before major bone surgery would be beneficial for patients who are likely to suffer from stroke.

    View details for DOI 10.1371/journal.pone.0153835

    View details for Web of Science ID 000374527000054

    View details for PubMedID 27089041

    View details for PubMedCentralID PMC4835054

  • Vascular Integrity in the Pathogenesis of Brain Arteriovenous Malformation. Acta neurochirurgica. Supplement Zhang, R., Zhu, W., Su, H. 2016; 121: 29-35

    Abstract

    Brain arteriovenous malformation (bAVM) is an important cause of intracranial hemorrhage (ICH), particularly in the young population. ICH is the first clinical symptom in about 50 % of bAVM patients. The vessels in bAVM are fragile and prone to rupture, causing bleeding into the brain. About 30 % of unruptured and non-hemorrhagic bAVMs demonstrate microscopic evidence of hemosiderin in the vascular wall. In bAVM mouse models, vascular mural cell coverage is reduced in the AVM lesion, accompanied by vascular leakage and microhemorrhage. In this review, we discuss possible signaling pathways involved in abnormal vascular development in bAVM.

    View details for DOI 10.1007/978-3-319-18497-5_6

    View details for PubMedID 26463919

    View details for PubMedCentralID PMC4654926

  • Neuropathologic analysis of Tyr69His TTR variant meningovascular amyloidosis with dementia. Acta neuropathologica communications Ziskin, J. L., Greicius, M. D., Zhu, W., Okumu, A. N., Adams, C. M., Plowey, E. D. 2015; 3 (1): 43-?

    Abstract

    Transthyretin/TTR gene mutations usually cause systemic amyloidotic diseases. Few TTR variants preferentially affect the central nervous system, manifesting as oculoleptomeningeal amyloidosis. Patients with TTR meningovascular amyloidosis often show dementia, however the neuropathologic features of dementia in these cases have not been elucidated. We report the neuropathologic findings from a brain autopsy of a 72-year-old man with the rare Tyr69His (Y69H) TTR gene variant, dementia and ataxia. Severe amyloid deposits were observed in the leptomeninges and in a subpial and subependymal distribution. Mass spectrometry analysis demonstrated that the amyloid deposits were comprised of over 80 % of the variant TTR. TTR was undetectable by mass spectrometry in the neocortex subjacent to the subpial amyloid deposits. Subpial TTR amyloid deposits were associated with brisk superficial reactive gliosis and siderosis in the neocortex and cerebellar cortex. Subependymal TTR amyloid deposits were associated with subjacent myelin pallor in the hippocampal outflow tract structures including the alveus, fimbria and fornix. Phospho-tau immunostains demonstrated transentorhinal-stage neurofibrillary degeneration (Braak stage II) which, in the absence of neocortical amyloid-beta and neuritic plaques, was indicative of primary age-related tauopathy (PART). However, distinctive phospho-tau aggregates were observed subjacent to the subpial TTR amyloid deposits in all regions of the neocortex, including the primary motor and striate cortices, suggesting a potential link between TTR amyloid and neocortical tauopathy. Our report reveals novel insights into the potential neuropathologic substrates of dementia in variant TTR amyloidosis that need to be investigated in larger autopsy series.

    View details for DOI 10.1186/s40478-015-0216-0

    View details for PubMedID 26156087

    View details for PubMedCentralID PMC4496870

  • GA binding protein augments autophagy via transcriptional activation of BECN1-PIK3C3 complex genes AUTOPHAGY Zhu, W., Swaminathan, G., Plowey, E. D. 2014; 10 (9): 1622-1636

    View details for DOI 10.4161/auto.29454

    View details for Web of Science ID 000341647200012

  • Mutant LRRK2 overexpression in cultured cortical neurons is associated with increased glutamatergic synapse activity and excitotoxic dendrite degeneration Biochim Biophys Acta Plowey, E. D., Johnson, J. W., Steer , E., Zhu, W., Eisenberg, D. A., Valentino, N. M., Liu, Y., Chu, C. T. 2014
  • GA-binding protein augments autophagy via transcriptional activation of BECN1-PIK3C3 complex genes Autophagy Zhu, W., Swaminathan, G., Plowey, E. 2014
  • Transcriptional and Post-Transcriptional Regulation of SPAST, the Gene Most Frequently Mutated in Hereditary Spastic Paraplegia PLOS ONE Henson, B. J., Zhu, W., Hardaway, K., Wetzel, J. L., Stefan, M., Albers, K. M., Nicholls, R. D. 2012; 7 (5)

    Abstract

    Hereditary spastic paraplegias (HSPs) comprise a group of neurodegenerative disorders that are characterized by progressive spasticity of the lower extremities, due to axonal degeneration in the corticospinal motor tracts. HSPs are genetically heterogeneous and show autosomal dominant inheritance in ∼70-80% of cases, with additional cases being recessive or X-linked. The most common type of HSP is SPG4 with mutations in the SPAST gene, encoding spastin, which occurs in 40% of dominantly inherited cases and in ∼10% of sporadic cases. Both loss-of-function and dominant-negative mutation mechanisms have been described for SPG4, suggesting that precise or stoichiometric levels of spastin are necessary for biological function. Therefore, we hypothesized that regulatory mechanisms controlling expression of SPAST are important determinants of spastin biology, and if altered, could contribute to the development and progression of the disease. To examine the transcriptional and post-transcriptional regulation of SPAST, we used molecular phylogenetic methods to identify conserved sequences for putative transcription factor binding sites and miRNA targeting motifs in the SPAST promoter and 3'-UTR, respectively. By a variety of molecular methods, we demonstrate that SPAST transcription is positively regulated by NRF1 and SOX11. Furthermore, we show that miR-96 and miR-182 negatively regulate SPAST by effects on mRNA stability and protein level. These transcriptional and miRNA regulatory mechanisms provide new functional targets for mutation screening and therapeutic targeting in HSP.

    View details for DOI 10.1371/journal.pone.0036505

    View details for Web of Science ID 000305349800070

    View details for PubMedID 22574173