SIRT3在砷暴露诱导HELF细胞纤维化中的作用和机制OA

[Background]Arsenic exposure has been implicated as an important environmental contributor to pulmonary fibrosis,the pathogenesis of which is closely associated with abnormal activation of pulmonary fibroblasts.However,the specific mechanism remain incompletely understood. [Objective]To examine sirtuin 3(SIRT3)expression during arsenite-induced fibrotic responses in lung fibroblasts,and to investigate its effects on aerobic glycolysis and profibrotic progression,as well as its underlying mechanisms. [Methods]Human embryonic lung fibroblasts(HELF)were treated with sodium arsenite(NaAsO2)at different concentrations(0,2.5,5,10,20,40,and 80 μmol·L-1)for 48 h.Cell viability was assessed,and lactate and hydroxyproline(HYP)levels were measured as indicator of glycolytic activity and fibrotic response.Western blot and quantitative polymerase chain reaction(q-PCR)were used to determine the protein and mRNA expression levels of SIRT3,hypoxia-inducible factor 1-alpha(HIF-1α),fibrosis-related genes[Collagen type Ⅰ(Collagen-Ⅰ),and α-smooth muscle actin(α-SMA)],and glycolysis-related genes[pyruvate kinase M2(PKM2),hexokinase 2(HK2),and lactate dehydrogenase A(LDHA)]. [Results]Based on the cell viability results,the NaAsO2 concentrations of 2.5,5,and 10 μmol·L-1 for 48 h were selected for subsequent experiments.Compared with the control group,HYP levels were significantly increased in the 2.5,5,and 10 μmol·L-1 NaAsO2 exposure groups(P<0.05).The protein and mRNA expression levels of HIF-1α,glycolysis-related genes(PKM2,HK2,and LDHA)and fibrosis-related genes(Collagen-Ⅰ,and α-SMA)were significantly upregulated,whereas SIRT3 protein and mRNA expression levels were significantly down-regulated,in the NaAsO2 treated groups compared with the control group(P<0.05).SIRT3 overexpression significantly reduced HYP levels and downregulated the protein and mRNA expression levels of HIF-1α,glycolysis-related genes(PKM2,HK2,and LDHA),fibrosis-related genes(Collagen-Ⅰ,and α-SMA)compared with the NaAsO2+negative control group(P<0.05).Similarly,HIF-1α knockdown significantly decreased HYP levels and downregulated the protein and mRNA expression levels of glycolysis-related genes(PKM2,HK2,and LDHA)and fibrosis-related genes(Collagen-Ⅰ,and α-SMA)compared with the NaAsO2+negative control group(P<0.05).In con-trast,simultaneous overexpression of SIRT3 and HIF-1α abolished the inhibitory effects of SIRT3 overexporession,as HYP levels and the expression of glycolysis-and fibrosis-related markers showed no statistically significant differences compared with the NaAsO2+negative control group(P>0.05). [Conclusion]NaAsO2 exposure could induce profibrotic and glycolysis-related responses in HELF cells,accompanied by SIRT3 downregu-lation and HIF-1α activation.These findings suggest that the SIRT3/HIF-1α regulatory axis may be a potential target for preventing or at-tenuating arsenic exposure-related pulmonary fibrotic responses.