A new look at angiogenesis inhibition of geniposide in experimental arthritis by blocking angiopoietin‐2 exocytosis

Abstract Angiogenesis is a key player in the pathogenesis of rheumatoid arthritis. Exocytosis from Weibel–Palade bodies is a prerequisite for angiopoietin‐2 (Ang‐2) to activate endothelial cells and initiate angiogenesis. Geniposide (GE) was previously reported to exert anti‐angiogenic effects. The aim of this study was to shed light on whether and how GE regulates Ang‐2 exocytosis. A rat model of adjuvant arthritis (AA) was established to evaluate the therapeutic effect of GE (60 and 120 mg/kg) especially in synovial angiogenesis. In addition, the Matrigel plug assay was used to detect the effect of GE (120 and 240 mg/kg) on angiogenesis in AA mice. In vitro, sphingosine‐1‐phosphate (S1P)‐stimulated human umbilical vein endothelial cells (HUVECs) were used to investigate the effect and mechanism of GE on Ang‐2 exocytosis. It was found that GE improved the symptoms of AA rats and inhibited angiogenesis in AA, which may be related to the down‐regulation of S1P receptors 1, 3 (S1PR1, S1PR3), phospholipase Cβ3 (PLCβ3), inositol 1,4,5‐trisphosphate receptor (IP 3 R) and Ang‐2 expression. The results of in vitro experiments showed that S1P induced rapid release of Ang‐2 from HUVECs with multigranular exocytosis. Suppression of the S1P/S1PR1/3/PLCβ3/Ca 2+ signal axis by the S1PR1/3 inhibitor VPC23019 and the IP 3 R inhibitor 2‐APB blocked Ang‐2 exocytosis, accompanied by diminished angiogenesis in vitro. GE dose‐dependently weakened S1P/S1PR1/3/PLCβ3/Ca 2+ signal axis activation, Ang‐2 exocytosis and angiogenesis in HUVECs ( p < 0.05, p < 0.01). Overall, these findings revealed that angiogenesis inhibition of GE was partly attributed to the intervention of Ang‐2 exocytosis through negatively modulating the S1P/S1PR1/3/PLCβ3/Ca 2+ signal axis, providing a novel strategy for rheumatoid arthritis anti‐angiogenic therapy.