SKF38393 prevents high glucose (HG)-induced endothelial dysfunction by inhibiting the effects of HG on cystathionine γ-lyase/hydrogen sulfide activity and via a RhoA/ROCK1 pathway.

BACKGROUND: Endothelial dysfunction plays a crucial role in diabetic vascular complications. A decrease in hydrogen sulfide (H2S) levels is increasingly becoming a vital factor contributing to high glucose (HG)-induced endothelial dysfunction. Dopamine D1-like receptors (DR1) activation has important physiological functions in the cardiovascular system. H2S decreases the dysfunction of vascular endothelial cells. However, no studies have reported whether DR1 protects the function of vascular endothelial cells by regulating H2S levels. AIM: The present study aimed to determine whether DR1 regulates the levels of endogenous H2S, which exerts protective effects against HG-induced injury of human umbilical vein endothelial cells (HUVECs) via Ras homolog gene family member A (RhoA)/Rho-associated coiled-coil containing kinase 1 (ROCK1) signalling. METHODS: HUVECs were exposed to HG (30 mM) or normal glucose (5.5 mM) after different treatments. Cell viability, proliferation and migration were measured by Cell Counting Kit-8, EdU cell proliferation assay, transwell assay and wound healing assay, respectively. H2S probe (7-Azido-4-Methylcoumarin) was used to detect levels of H2S. The intracellular calcium concentration ([Ca2+]i) were measured using Fluo-4 AM. The protein expressions were quantified by Western blot. RESULTS: We found that HG decreased the expression of DR1 and cystathionine γ-lyase (CSE) and H2S production. The DR1 agonist SKF38393 significantly increased DR1 and CSE expression and H2S production, whereas NaHS (a H2S donor) only increased CSE expression and H2S production but had no effect on DR1 expression. Meanwhile, SKF38393 further increased the [Ca2+]i induced by HG. In addition, HG reduced cell viability and the expression of Cyclin D1 and proliferating cell nuclear antigen and increased the expression of p21C⁢i⁢p/W⁢A⁢F-1, collagen I, collagen III, matrix metalloproteinase 9, osteopontin and α-smooth muscle actin and the activity of phosphorylated RhoA and ROCK1. SKF38393 and NaHS reversed these effects of HG. PPG (a CSE inhibitor) abolished the beneficial effect of SKF38393. These effects of SKF38393 were similar to those of Y-27632 (a ROCK inhibitor). CONCLUSION: Taken together, our results suggest that DR1 activation upregulates the CSE/H2S pathway by increasing the [Ca2+]i, which protects endothelial cells from HG-induced injury by inhibiting the RhoA/ROCK1 pathway.

[Effects of exogenous H2S on hepatic fibrosis in diabetic mice and its mechanism].

OBJECTIVE: To investigate the effects of exogenous hydrogen sulfide (H2S) on the hepatic fibrosis in diabetic mice and its mechanism. METHODS: Twenty-four C57 male mice (weight 22±2 g) were randomly divided into three groups (n=8): ① Normal control group (Control): Mice were intraperitoneally injected equal amount of normal saline, the injection time was the same as that of the experimental groups; ② Diabetes model groups (HG): Streptozotocin (STZ) was injected intraperitoneally once according to body weight (150 mg/kg) to establish diabetes model; ③ NaHS treatment groups (HG + NaHS): Mice were intraperitoneally injected with NaHS (100 µmol/L·kg·d) once a day for 12 consecutive weeks. The hepatocyte injury was detected by HE staining; the hepatic fibrosis was observed through Masson staining; the protein expressions of cystathionine - ß - synthetase (CBS), collagen-I (CoL-I), collagen-III (CoL-III) and matrix metalloproteinase-9 (MMP-9) were detected by Western blot. RESULTS: Compared with the control group, the damage and fibrosis of hepatocyte were significantly aggravated, the expression of CBS proteins was decreased (P＜0.01), and the expression levels of CoL-I, CoL-III and MMP-9 proteins were increased (P＜0.01) in the diabetic model group. Compared with the diabetic model group, the damage and fibrosis of hepatocyte were significantly lightened, the expression of CBS proteins was obviously increased (P＜0.01), and the expression levels of CoL-I, CoL-III and MMP-9 proteins were markedly decreased (P＜ 0.01). CONCLUSION: H2S inhibits the hepatic fibrosis in diabetic mice, and its mechanism is related to the decrease of collagen and matrix metalloproteinase-9.