Effects Of H ₂ S On Oxidative Stress Induced By High Glucose In Human Peritoneal Mesothelial Cells

OBJECTIVETo investigate the impaired effect of high glucose on the human peritonealmesothelial cells (HPMCs) line HMrSV5cells and to observate the effect of highglucose on the secretion of endogenous hydrogen sulfide (H2S) and explore thepossible mechanisms in HMrSV5cells, to further investigate the protective effect ofextrogenous H2S on the injury induced by high glucose and explore the possiblemechanisms in HMrSV5cells.METHODSThe human peritoneal mesothelial cells (HPMCs) line HMrSV5cells werecultivated in dulbecco’s modification of eagle’s medium (DMEM). HMrSV5cellswere incubated by4.25%D-glucose (high glucose) to induce injury and the cells weretreated with the different concentration (5×10-5,10-4,5×10-4,10-3and5×10-3mol/L) of extrogenous H2S donor sodium bisulfide (NaHS) for24h. MTT assay wasused to detect the cell viability. The levels of lactate dehydrogenase (LDH) and H2Sin the culture medium were measured by spectrophotography. The level of totalantioxidant capacity (T-AOC) was tested by double antibody sandwich method assay.The level of reactive oxygen species (ROS) in the cells was measured by flowcytometry. The level of malondialdehyde (MDA) in the cells and the activity ofsuperoxide dismutase (SOD) in the cells were measured by spectrophotography. Thelevel of reduced glutathione hormon (GSH) in the cells was measured by ELISAassay. The mRNA and protein expressions of cystathionine gamma lyase (CSE) andcystathionine beta synthase (CBS) in the cells were measured by Real-time PCR andWestern-Blot respectively.RESULTS(1) Compared with the control group, the cell viability was significantlydecreased in the high glucose group (P﹤0.05) and the level of LDH in the culture medium was significantly increased in the high glucose group (P﹤0.05).(2) Compared with the control group, the concentration of H2S in the culturemedium of HMrSV5cells was significantly decreased in the high glucose group(4.25%D-glucose)(P﹤0.05). The mRNA and protein expressions of CSE wassignificantly down-regulated compared with the control group (all P﹤0.05). Therewas no significant difference in the mRNA and protein expressions of CBS betweenthe control group and the high glucose group.(3) Compared with the control group, the levels of ROS and MDA in the cellswere significantly increased in the high glucose group (all P﹤0.05). The level ofT-AOC, the acivity of SOD and the level of GSH were significantly decreased in thehigh glucose group compared with the control group (all P﹤0.05).(4) Compared with the high glucose group, the cell viabilities were significantlyincreased in extrogenous H2S donor NaHS (5×10-4,10-3and5×10-3mol/L) group (allP﹤0.05) and the levels of LDH in the culture medium were significantly decreased inNaHS (5×10-4,10-3and5×10-3mol/L) group.(5) Compared with the high glucose group, the levels of ROS and MDA in thecells were significantly decreased in the extrogenous H2S donor NaHS (10-3mol/L)group (all P﹤0.05). The level of T-AOC, the acivity of SOD and the level of GSHwere significantly increased in the extrogenous H2S donor NaHS (10-3mol/L) groupcompared with the control group (all P﹤0.05).CONCLUSIONS(1) The high glucose circumstance induces the injury in the human peritonealmesothelial cells line HMrSV5cells, which the mechanisms may be related to thedown-reguration of CSE expression, the decrease of endogenous H2S and oxidativestress induced by the high glucose.(2) Extrogenous H2S prevents the injury induced by high glucose in humanperitoneal mesothelial cells line HMrSV5cells, which the mechanisms may be relatedwith the inhibition of oxidative stress induced by extrogenous H2S.