The thrust of the application is the same as for the original application: to investigate the physiological interactions between microvascular endothelial cells and luteal steroidogenic cells in the development and function of the primate (rhesus monkey) corpus luteum. The overall hypothesis is that luteal cell-microvascular endothelial cell interactions occur via vascular endothelial growth factor (VEGF) and its receptor and that such interactions are critical for the development and function of the corpus luteum. The 4 Specific Aims are the same as before: 1) identify VEGF producing cells and VEGF target cells (with Flt-1/KDR receptors) in the periovulatory follicle and corpus luteum; 2) investigate the actions of VEGF on endothelial cells isolated from the primate corpus luteum; 3) investigate the factors controlling VEGF expression; 4) determine whether paracrine communication between luteal cells and endothelial cells via VEGF-Flt-1/KDR receptors regulates the development and function of the corpus luteum. To do these studies, endothelial cells are isolated using magnetic beads coated with a lectin. VEGF and VEGF receptor or their mRNAs will be analyzed by immunocytochemistry and Western blotting or by RT-PCR and in situ hybridization, respectively. Interactions between luteal cells and endothelial cells in co-culture or in vivo will be analyzed in the presence of antisense oligonucleotides or neutralizing antibodies that disrupt the VEGF-receptor system. Overall, the proposed research may have relevance to an understanding of certain ovarian disorders such as ovarian hyperstimulation syndrome and luteal phase defects where microvasculature growth may be disrupted.
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