Expression of GPR80/99 Receptor and Leukotriene E4-induced Intracellular Ca2+ Response in Human Vascular Endothelial Cells
Sunday, March 4, 2018
South Hall A2 (Convention Center)
Hideaki Shirasaki, MD PhD, Etsuko Saikawa, MD,PhD, Tetsuo Himi, MD, PhD
RATIONALE: GPR99 (also called GPR80) has emerged as a potential novel receptor with LTE4.Recently,we have shown that the nasal GPR80/99 receptor localized exclusively in blood vessels (Shirasaki et al. ANL 2017). The purpose of this study was to investigate the expression of GPR80/99 protein in human umbilical vein endothelial cells (HUVECs). Ca2+ fluxes caused by LTE4 in HUVECs was also studied.

METHODS: Western blot and immunohistochemical analysis were performed using anti-human GPR80/99 antibody. HUVECs in 96-well plates were loaded with the fluorescent Ca2+-sensitive dye Fluo-4 AM, and LTE4-induced Ca2+ influx was measured by using fluorescence plate reader. Ca2+ images were acquired sequentially with an exposure period of 30m second as full-flame image.

RESULTS: Western blot analysis revealved that a single band of approximately 38kDa was detected in HUVECs. The immunohistochemical analysis showed that anti-GPR80/99 antibody densely labeled the cell membrane of HUVECs. LTE4 caused a rapid increase of the intracellular Ca2+ concentration. CysLT1 receptor antagonist, montelukast did not affect the LTE4-induced intracellular Ca2+ response.

CONCLUSIONS: These results suggest that a potential third CysLT receptor such as GPR80/99 receptor may contribute LTE4-induced activation of vascular endothelial cells.