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Article Title

HISTAMINERGIC REGULATION OF ANGIOGENIC POTENTIAL IN HUMAN UMBILICAL VEIN ENDOTHELIAL CELLS

Abstract

Histamine has been shown to be a pro-angiogenic mediator, and may contribute to physiological angiogenesis associated with exercise training. Endothelial cells are a key effector of angiogenesis, and express histamine H1 and H2 receptors. Within the skeletal muscle vasculature, combined histamine H1 and H2 receptor blockade blunts the sustained post-exercise vasodilation after an acute bout of dynamic exercise. However, whether histamine receptor activation with exercise contributes to angiogenesis is unknown. The ability of endothelial cells to form capillary-like tubules when seeded on a basement membrane-like matrix can be used to investigate angiogenic signaling pathways in vitro. PURPOSE: To test the hypothesis that histamine receptor activation contributes to angiogenic potential in human umbilical vein endothelial cells (HUVEC). METHODS: HUVEC from passage numbers 2 - 3 were used for this study. After a 16 hour serum starvation to induce cellular quiescence, HUVEC were then plated in Matrigel-coated wells on a 96-well cell culture plate at a density of 1.0x105 cells/mL. Each well was then treated with a single dose (range: 10-9 to10-3M) of one of the following: histamine, fexofenadine or pyrilamine (H1-receptor blockers), and ranitidine or cimetidine (H2-receptor blockers). HUVEC plated in cell growth media served as controls. The HUVEC were then incubated at 37°C and 5% CO2 for 8hr. After 8hr, images of each well were acquired using phase-contrast microscopy. Total tubule length was quantified as a measure of endothelial tube formation using Image J. RESULTS: Compared to control conditions, histamine had no effect on endothelial tube formation (p=0.71). Additionally, preliminary data suggests neither H1 nor H2 receptor blockade alone have any effect on endothelial tube formation compared to control conditions. However, preliminary data indicates that combination of H1 and H2 receptor blockade with 10-3M pyrilamine plus cimetidine reduces total endothelial tubule length by 24%. CONCLUSION: Histamine does not increase angiogenic potential, and blocking either H1 or H2 receptors alone does not diminish angiogenic potential in vitro. However, combining H1 and H2 receptor blockade at 10-3M concentration appears to blunt angiogenic potential in the HUVEC endothelial tube formation assay.

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