Exogenous Hydrogen Sulfide Inhibites NIH3T3 Cells Proliferation By Down Regulating TGF β-PI3K/AKT-HIF-1α Signal Pathway

Objective:To investigate whether exogenous hydrogen sulfide can inhibit the proliferation of NIH3T3 cells through the TGFβ-PI3K/AKT-HIF-1αamplification pathway and its mechanism.Methods:The mouse embryonic lung fibroblasts（NIH3T3 cells）cultured in vitro were randomly divided into five groups:group A（blank control group）,group B（TGF-β1）,group C（TGF-β1+NaHS）,group D（TGF-β1+MK-2206）,and group E（TGF-β1+NaHS+MK-2206）.Group A did not add TGF-β1 and incubated it in A normal cell incubator（37℃,5%CO₂ and 95%air）for 24 hours.Group B joined TGF-β1（5μg/L）cultivation for 24 hours;Group C added NaHS（40μmol/L）for 1 hour pretreatment and then added TGF-β1（5μg/L）for 24 hours.Group D was added to MK-2206（50 nmol/L）for 1 hour and then added to TGF-β1（5μg/L）for 24 hours.Group E added MK-2206（50 nmol/L）for 1 hour,then added NaHS（40μmol/L）for 1 hour,and finally added TGF-β1（5μg/L）for 24 hours.The morphological changes of cells in the five groups were observed at 12,24 and 48 hours respectively,and the expressions of the five proteins ofα-SMA,Collagen I,AKT,ρ-AKT,and HIF-1α,respectively,were detected.Results:1.The results of inverted phase contrast microscopy showed that the fibroblasts in group A were spindle-shaped or stellate structures with multiple protuberances.However,there were more myofibroblasts in other groups,especially in group B.2.MTT results:the cell survival rate of group B,C,D and E in 12,24and 48 hours was from the highest to the lowest at B>D>C>E>A（P<0.05）,and the cell survival rate at 24 hours was the highest at three time points.3.WB results:The protein levels ofα-SMA and collagen I were B>D>C>E>A（P<0.05）.The protein levels of AKT,ρ-AKT and HIF-1αwere B>C>D>E>A（P<0.05）.Conclusion:Exogenous hydrogen sulfide can inhibit the expression ofρ-AKT and HIF-1αby down-regulating the TGFβ-PI3K/AKT-HIF-1αsignaling pathway,and inhibit the proliferation of NIH3T3 cells.