In the present study,we aimed to confirm whether NF-κB inhibitor pyrrolidine dithiocarbamate(PDTC)could induce apoptosis of human hepatic stellate cells(HSCs)LX-2 and explore the potential pharmacological mechanism u...In the present study,we aimed to confirm whether NF-κB inhibitor pyrrolidine dithiocarbamate(PDTC)could induce apoptosis of human hepatic stellate cells(HSCs)LX-2 and explore the potential pharmacological mechanism underlying these effects.In this study,LX-2 cells were cultured in vitro,and the experiment was divided into two groups,including the control and PDTC groups.The viability of LX-2 cells was measured by CCK8 assay after the cells were exposed to PDTC.The anti-apoptotic effect of PDTC was detected by AO/EB double assay staining kit.Additionally,the activities of NF-κB,Fas/FasL,apoptosis-related proteins,as well as the cellular localization of AIF,were determined by Western blotting analysis and immunofluorescence staining respectively.After PDTC treatment for 12 and 24 h,AO/EB dual staining showed typical apoptotic changes,such as cell volume reduction,cell shrinkage,nuclear fragmentation,and so on.PDTC at 60μmol/L significantly increased the proliferation inhibition rate and decreased the secretion of collagen I,collagen III,andα-SMA in LX-2 cells.The Western blotting analysis and RT-PCR showed no significant difference in the expression of AIF between the control group and PDTC group,and the expressions of Fas and FasL were not observed in all groups(P>0.05).Further results showed that PDTC could promote the displacement of AIF from mitochondria to the nucleus,activate the apoptotic signaling in the cell nucleus,and possibly participate in the apoptosis process of LX-2 cells.In conclusion,the pharmacological mechanism of PDTC against hepatic fibrosis might be to promote the displacement of AIF from mitochondria to the nucleus,then activate the apoptotic signaling in the cell nucleus,and finally induce the apoptosis of LX-2 cells.Meanwhile,these results also revealed that the Fas/FasL-mediated apoptosis pathway was not involved in the PDTC-induced apoptosis process of LX-2 cells.展开更多
基金supported by the Fujian Provincial Natural Science Foundation,China(Grant No.2019J01611)the Youth Project of Fujian Provincial Health Commission,Fujian Province,China(Grant No.2017-1-94).
文摘In the present study,we aimed to confirm whether NF-κB inhibitor pyrrolidine dithiocarbamate(PDTC)could induce apoptosis of human hepatic stellate cells(HSCs)LX-2 and explore the potential pharmacological mechanism underlying these effects.In this study,LX-2 cells were cultured in vitro,and the experiment was divided into two groups,including the control and PDTC groups.The viability of LX-2 cells was measured by CCK8 assay after the cells were exposed to PDTC.The anti-apoptotic effect of PDTC was detected by AO/EB double assay staining kit.Additionally,the activities of NF-κB,Fas/FasL,apoptosis-related proteins,as well as the cellular localization of AIF,were determined by Western blotting analysis and immunofluorescence staining respectively.After PDTC treatment for 12 and 24 h,AO/EB dual staining showed typical apoptotic changes,such as cell volume reduction,cell shrinkage,nuclear fragmentation,and so on.PDTC at 60μmol/L significantly increased the proliferation inhibition rate and decreased the secretion of collagen I,collagen III,andα-SMA in LX-2 cells.The Western blotting analysis and RT-PCR showed no significant difference in the expression of AIF between the control group and PDTC group,and the expressions of Fas and FasL were not observed in all groups(P>0.05).Further results showed that PDTC could promote the displacement of AIF from mitochondria to the nucleus,activate the apoptotic signaling in the cell nucleus,and possibly participate in the apoptosis process of LX-2 cells.In conclusion,the pharmacological mechanism of PDTC against hepatic fibrosis might be to promote the displacement of AIF from mitochondria to the nucleus,then activate the apoptotic signaling in the cell nucleus,and finally induce the apoptosis of LX-2 cells.Meanwhile,these results also revealed that the Fas/FasL-mediated apoptosis pathway was not involved in the PDTC-induced apoptosis process of LX-2 cells.
