In response to fluctuations in environmental oxygen in the cells of the blood brain barrier (BBB) undergo a number of complex adaptive measures in order to maintain tissue homeostasis and hemostasis. These adaptive responses are particularly important when the balance of oxygen availability and utilization is altered as a result of the pathophysiology of CNS disease. At the microvascular level oxygen responsive signaling mechanisms involving both the endothelial cell and the pericyte regulate angiogenesis, vascular permeability and metabolism. We propose to investigate early responses of the CNS pericyte following in vitro exposure to hypoxia using the GaSPak 100 hypoxia system (Becton Dickinson and Company Sparks, MD.) Within 15 minutes of exposure to low oxygen (1%) pericytes synthesize and release cyclopentenone prostaglandins. Increased PGD2 and the dehydration product delta12pGJ2 were detected by HPLC and by immune techniques. Delta12pGD2 was not synthesized. Pericytes constitutively express three of the five alternate splice variants of vascular endothelial cell growth factor (VEGF) mRNA, but little to no protein. Exposure to low oxygen increased mRNA levels as well as the synthesis and release of VEGF protein. Addition of either Delta12pGJ2 or 15-deoxyDelta12/14PGJ2 to pericyte under normoxic conditions increased the synthesis and release of VEGF protein in a dose dependent manner. Results suggest that PGD produced by the CNS pericyte is an early signaling molecule in regulation of the angiogenic response to hypoxia. We will further investigate the pericyte VEGF response:
Aim # l: Determine the mechanism of hypoxia induced upregulation of VEGF gene expression. We will also evaluate the role of translation initially and inhibiting factor elF-4E and 4EBprincipal investigator.
Aim#2 : To determine the role of COX-1 in pericyte VEGF response to hypoxia.
Aim#3 : To determine the role of the COX-1 mediated release of pericyte PGD/PGJ in hypoxia. We propose that the CNS microvascular pericyte is a regulatory cell important in the vascular response to hypoxic stress and that cyclopentenone prostagladins are involved in the pericyte stress response.
