Autosomal dominant polycystic kidney disease (ADPKD) is typified by the accumulation of fluid-filled cysts and abnormalities in renal epithelial cell function. The disease is principally caused by mutations in the gene encoding polycystin-1, a large basolateral plasma membrane protein expressed in kidney epithelial cells. Our studies reveal that, in normal kidney cells, polycystin-1 forms a complex with the adherens junction protein E-cadherin and its associated catenins, suggesting a role in cell adhesion or polarity. In primary cells from ADPKD patients, the polycystin-1/polycystin-2/E-cadherin/beta-catenin complex was disrupted and both polycystin-1 and E-cadherin were depleted from the plasma membrane as a result of the increased phosphorylation of polycystin-1. The loss of E-cadherin was compensated by the transcriptional upregulation of the normally mesenchymal N-cadherin. Increased cell surface N-cadherin in the disease cells in turn stabilized the continued plasma membrane localization of beta-catenin in the absence of E-cadherin. The results suggest that enhanced phosphorylation of polycystin-1 in ADPKD cells precipitates changes in its localization and its ability to form protein complexes that are critical for the stabilization of adherens junctions and the maintenance of a fully differentiated polarized renal epithelium.

Changes in extracellular space (ECS) diffusion parameters in astrogliotic tissue around a unilateral cortical stab wound were determined from concentrationtime profiles of tetramethylammonium (TMA ϩ ) using TMA ϩ -selective microelectrodes. Three diffusion parameters-ECS volume fraction ␣ (␣ ϭ ECS volume/ total tissue volume), tortuosity ( 2 ϭ D/ADC; where D is the free and ADC is the apparent diffusion coefficient of TMA ϩ in the brain), and nonspecific TMA ϩ uptake kЈ-were determined at 3, 7, 21, and 35 days postwounding (dpw), in the hemispheres ipsilateral and contralateral to the lesion. Following diffusion experiments, tissue sections were immunostained for glial fibrillary acidic protein (GFAP) and chondroitin-sulphate proteoglycans (CSPG). In the area 300-1000 µm around the wound, ␣ was increased at 3, 7, and 21 dpw by about 20% but returned to control values at 35 dpw; was increased at all four intervals, reaching a maximum at 7 dpw. kЈ was lower than in the contralateral hemisphere at 7, 21, and 35 dpw. Measurements 1,500-2,000 µm from the wound revealed only an increase in at 7 dpw. The time course of changes in ECS diffusion parameters closely correlated with increased staining for GFAP and CSPG. Our results show that astrogliosis significantly changes the diffusion properties of nervous tissue, making it less permissive. Both hypertrophied astrocytic processes and an enhanced formation of some extracellular matrix molecules could affect, through changes in the diffusion of molecules in the ECS, neuron-glia communication, ''cross-talk'' between synapses, extrasynaptic transmission, and regenerative processes.

A multifunctional microRNA, miR-155, has been recently recognized as an important modulator of numerous biological processes. In our previous in vitro studies, miR-155 was identified as a potential regulator of the endothelial morphogenesis. The present study demonstrates that in vivo inhibition of miR-155 supports cerebral vasculature after experimental stroke. Intravenous injections of a specific miR-155 inhibitor were initiated at 48 h after mouse distal middle cerebral artery occlusion (dMCAO). Microvasculature in peri-infarct area, infarct size, and animal functional recovery were assessed at 1, 2, and 3 weeks after dMCAO. Using in vivo two-photon microscopy, we detected improved blood flow and microvascular integrity in the peri-infarct area of miR-155 inhibitor-injected mice. Electron microscopy revealed that, in contrast to the control group, these animals demonstrated well preserved capillary tight junctions (TJs). Western blot analysis data indicate that improved TJ integrity in the inhibitor-injected animals could be associated with stabilization of the TJ protein ZO-1 and mediated by the miR-155 target protein Rheb. MRI analysis showed significant (34%) reduction of infarct size in miR-155 inhibitor-injected animals at 21 d after dMCAO. Reduced brain injury was confirmed by electron microscopy demonstrating decreased neuronal damage in the peri-infarct area of stroke. Preservation of brain tissue was reflected in efficient functional recovery of inhibitor-injected animals. Based on our findings, we propose that in vivo miR-155 inhibition after ischemia supports brain microvasculature, reduces brain tissue damage, and improves the animal functional recovery.

Vascular cells provide a neural stem/progenitor cell (NSPC) niche that regulates expansion and differentiation of NSPCs within the germinal zones of the embryonic and adult brain under both physiologic and pathologic conditions. Here, we examined the NSPC-endothelial cell (NSPC/EC) interaction under conditions of ischemia, both in vitro and after intracerebral transplantation. In culture, embryonic mouse NSPCs supported capillary morphogenesis and protected ECs from cell death induced by serum starvation or by transient oxygen and glucose deprivation (OGD). Neural stem/progenitor cells constitutively expressed hypoxia-inducible factor 1alpha (HIF-1alpha) transcription factor and vascular endothelial growth factor (VEGF), both of which were increased approximately twofold after the exposure of NSPCs to OGD. The protective effects of NSPCs on ECs under conditions of serum starvation and hypoxia were blocked by pharmacological inhibitors of VEGF signaling, SU1498 and Flt-1-Fc. After intracerebral transplantation, NSPCs continued to express HIF-1alpha and VEGF, and promoted microvascular density after focal ischemia. These studies support a role for NSPCs in stabilization of vasculature during ischemia, mediated via HIF-1alpha-VEGF signaling pathways, and suggest therapeutic application of NSPCs to promote revascularization and repair after brain injury.