Effects Of Exogenous H₂S On Autophagy In Rats With Pulmonary Fibrosis Through PI3K/AKT/mTOR Signaling Pathway

Objective::To investigate the effect of exogenous hydrogen sulfide on the expression of PI3k/AKT/mTOR mRNA and autophagy-associated protein:LC3Ⅱ,P62 protein in lung tissue of rats with bleomycin-induced pulmonary fibrosis.To investigate the effect of exogenous hydrogen sulfide on autophagy in rats with pulmonary fibrosis through PI3k/AKT/mTOR signaling pathway.Methods:Forty-five adult male SD rats were randomly divided into two groups:control group（n=15）,BLM NaHS group（n=15）.The model of pulmonary fibrosis was established by intratracheal injection of bleomycin（5mg/Kg）in,BLM NaHS group in BLM group.The rats in Control group were treated with single intratracheal injection of equal volume saline,and the first injection date was set as 0 day.Group C received intraperitoneal injection of NaHS（donor of exogenous hydrogen sulfide）at 28μmol·kg^-1·d^-1,while Contro was injected intraperitoneally on the first day after the establishment of the model.L group,BLM group was injected intraperitoneally with normal saline(1mg·kg^-1·d^-1)every day.On the 7th day,14th day,28th day after the establishment of the model,5rats in three groups were killed at random.The left lung of the rats was placed in 10%formalin solution for HE staining and Masson staining.The histopathological changes of the lung were observed under light microscope.The right lung of rats was stored in-80℃refrigerator after liquid nitrogen freezing,and the expression level of TGF-β1,α-SMA,LC3Ⅱ,P62 protein was determined by Western-blot.Results:HE staining and Masson staining:the lung tissue structure of Control group was clear,and no obvious changes of alveolar wall rupture,tissue edema,interstitial thickening and fibrous hyperplasia were observed in Control group.On the 7th day and 14th day in,BLM NaHS group,the lung tissue structure was clear and clear,and there were no obvious changes in pulmonary alveolar wall rupture,tissue edema,interstitial thickening and fibrous hyperplasia.On the 28th day,the lung tissue was disordered,interstitial thickening,inflammatory exudation,fibrous hyperplasia and so on,which increased with the time.At the same time,the changes of lung tissue in BLM group were more severe than those in BLM NaHS group.Comparison of RT-qPCR results:Compared with Control group,the expression level of PI3K,AKT,mTOR mRNA in,BLM NaHS group of BLM group was relatively higher than that of BLM group;The expression level of PI3K,AKT,mTOR mRNA in BLM group was lower than that in,BLM NaHS group at the same time（P<0.05）.WB results:1)Protein expression of LC3Ⅱ:compared with Control group and BLM group,the expression level of LC3Ⅱ protein in,BLM NaHS group decreased significantly,especially in BLM group,the difference was statistically significant（P62）,TGF-β1,TGF-β1.The protein expression level ofα-SMA:compared with Control group,the expression level of,BLM NaHS group in BLM group was higher than that in BLM NaHS group,and the expression level in BLM group was significantly higher than that in BLM NaHS group（P<0.05）.Conclusion:1.The mechanism of pulmonary fibrosis may be related to the lack of autophagy;2.Exogenous hydrogen sulfide may increase cell autophagy by inhibiting the PI3k/AKT/mTOR signaling pathway to inhibit the progression of pulmonary fibrosis.