RGR modulates angiogenic sprouting under mild heat stress in vitro

Summary: Abstract Body: Objective: To elucidate the impact of the retinal G protein-coupled receptor (RGR) on angiogenic sprouting under MHS conditions. Methods: The expression profile of RGR in human umbilical vein endothelial cells (HUVECs) was examined utilizing immunofluorescent staining techniques. Quantification of RGR mRNA and protein levels was conducted through real-time quantitative PCR and Western blot analysis under varying MHS conditions (37°C, 39°C, 41°C and 43°C). Additionally, the expression and subcellular localization of RGR in HUVECs were detected under laser scanning confocal microscopy. The pro-angiogenic properties under different MHS conditions (37°C and 41°C) were identified using spheroid-based angiogenesis and tube formation assays. Lentiviral transfection techniques were employed to achieve overexpression or suppression of RGR in HUVECs. The effects of RGR overexpression or suppression on angiogenic sprouting in HUVECs under different MHS conditions (37°C and 41°C) were assessed through spheroid and tube formation assays. Results：Co-immunostaining analysis of RGR and CD31 revealed that RGR was localized to the cell membrane and nucleus of HUVECs, with a predominant expression in the cell membrane. Upon exposure to heat stress at temperatures of 37°C, 39°C, 41°C, and 43°C for one hour, a progressive increase in RGR mRNA and protein levels was detected. Significantly, cellular damage was observed in the group exposed to 43°C, whereas no cellular damage was detected in the other groups. Significantly enhanced angiogenic sprouting and tube formation were observed at 41°C for one hour, in comparison to the 37°C group. This enhancement effect was modulated by the overexpression and knockdown of RGR, respectively. Conclusions: RGR is capable of modulating angiogenic sprouting under various MHS conditions in vitro. Yulin Yang¹, Huanhuan Luo¹, Ning Han¹, Dechao Jia¹, Caixia Zhang¹, Jingdong Zhang¹, Yinghua Lan¹, Yu Wang¹, Hongguang Lu¹ 1. Department of Dermatology, Guizhou Medical University, Guiyang, Guizhou, China. Stem Cell Biology, Tissue Regeneration and Wound Healing