Hydrogen Sulfide Inhibits Epithelial-mesenchymal Transition In Peritoneal Mesothelial Cells

Objective:To investigate the molecular mechanism of hydrogen sulfide(H2S)in inhibiting epithelial-mesenchymal transition in peritoneal mesothelial cells.Methods:1.In vivo study,Twenty male Sprague-Dawley rats(male)were randomly divided into 4 groups:(1)control group: normal saline 20 m L/d intraperitoneally;(2)H2S group: 1 μmol/kg*d Na HS was intraperitoneally injected with normal saline;(3)Modeling group: 4.25% peritoneal dialysate 20 m L/d intraperitoneally,with 0.6mg/kg lipopolysaccharide(LPS)intraperitoneally on day 1,3,5,and 7;(4)H2S treatment group: 4.25% peritoneal dialysis solution was injected intraperitoneally at 20 m L/d,and 0.6mg/kg lipopolysaccharide(LPS)was intraperitoneally injected on day 1,3,5,and 7,and injected intraperitoneally with 1μmol/kg*d Na HS.After 28 days,the peritoneal tissue of the parietal layer was stained with Masson’s staining to observe the thickness of the peritoneum and collagen fibers.2.In vitro study,rat peritoneum was obtained under aseptic conditions.Rat primary peritoneal cells were obtained after digestion and cultured in vitro.After the cells were grown to 2-3 passages,the synchronized rat primary peritoneal mesothelial cells were randomly divided into 6 groups:(1)control group: no treatment;(2)model group: 4.25% peritoneal dialysate : 15% FBS complete culture solution(V:V)= 1:1;(3)50μmol/L H2 S treatment group: 4.25% peritoneal dialysate: 15% FBS complete culture solution(V:V)= 1:1 with 50μmol/L Na HS;(4)100μmol/L H2 S treatment group: 4.25% peritoneal dialysate:15% FBS complete culture solution(V:V)= 1:1 with 100μmol/L Na HS;(5)300μmol/L H2 S treatment Group: 4.25% peritoneal dialysate:15% FBS complete culture solution(V:V)= 1:1 with 300 μmol/L Na HS;(6)H2S group: non-treated cells with 300 μmol/L Na HS.Na HS is pretreated for 30 minutes in advance.The expressions of IL-6 and MCP-1 intracellular were detected by RT-PCR.The expressions of the phosphorylation of Smad2/3 and Smad3 in TGF-β1 signal pathway and related proteins were detected by Western Blot.Results:1.In vivo study,Masson staining showed that compared with the control group,the deposition of collagen fibers in the peritoneal mesothelial cells of the model group increased significantly,the dense layer thickened,and the peritoneal thickness increased significantly(P<0.001).After Na HS treatment,collagen fiber deposition,the volume and peritoneal thickness decreased significantly(P<0.01).2.In vitro study,Compared with the control group,the expression of inflammatory cytokines IL-6 and MCP-1 in the peritoneal mesothelial cells increased obviously in the model group,and the expression of the tight junction proteins ZO-1 and cytokeratin decreased meaningfully,while α-SMA,PAI-1,FN and phosphorylation Smad3 and Smad2/3 proteins expressions were markedly increased;after intervention with Na HS,the expression of inflammatory factors was significantly reduced compared with the model group,and the expressions of ZO-1 and cytokeratin proteins were increased,α-SMA,PAI-1,FN,and the phosphorylated Smad3,Smad2/3 proteins expression were downregulated,the difference was statistically significant.Conclusions:1.Exogenous H2 S can inhibit peritoneal fibrosis in rats with chronic peritonitis caused by high glucose peritoneal dialysis fluid and LPS.2.H2 S can ameliorate peritoneal fibrosis by affecting EMT of peritoneal mesothelial cells.3.The anti-inflammatory mechanism of H2 S is related to its regulation of TGF-β1-Smad signaling pathway and the anti-inflammation effect.In summary,H2 S can inhibit peritoneal fibrosis by inhibiting EMT of rat peritoneal mesothelial cells.H2 S anti-peritoneal mesothelial cell EMT mechanism is related to its regulation of TGF-β1-Smad signaling pathway and the anti-inflammation effect.