The Role Of Endoplasmic Reticulum Stress In Lead-induced Mitophagy Of HEK 293 Cells

Lead?Pb?is one of the harmful heavy metals,and has been used in industrial production for a long time.With the development of industrial revolution,the rela-tionship between Pb and human life has become more and more close.As a mul-ti-organ toxicant,Pb can be absorbed into various tissues and organs,thus possessing a potential toxic effect.The kidney is one of the most sensitive target organs to Pb toxicity.Pb accumulates mainly in renal tubule after entering kidney through metabo-lism,which leads to proximal tubular injury and finally induces failure.Organelles are necessary to maintain normal life activity of live cell.After entering the organism,all kinds of toxins will destroy the function of organelles,leading to the abnormal syn-thesis and metabolism of various biological macromolecules.Endoplasmic reticulum is closely networked with mitochondria via membrane structure,and plays a co-regulating role in life activities.Recently,the study has found that Pb can lead to dysfunction of endoplasmic reticulum,accumulation of misfolded proteins,and final-ly induce endoplasmic reticulum stress.Simultaneously,it is found that Pb-induced oxidative stress could not only destroy the dynamic balance of mitochondria,lead to abnormal morphology and dysfunction,but also trigger selective mitophagy.Under stress conditions,autophagy can be mediated by activation of endoplasmic reticulum stress pathway.At present,researchers have found that Pb can induce mitophagy and endoplasmic reticulum stress.However,the role and mechanism of endoplasmic re-ticulum stress in Pb induced mitophagy have not been reported.In the present research,human embryonic kidney cells HEK293 were applied to investigate the role and mechanism of endoplasmic reticulum stress in lead acetate?Pb?Ac?₂?induced mitophagy.Firstly,the effects of different concentrations of Pb?Ac?₂?0?0.5?1?2?4 mM?on cell viability and cell damage were detected by CCK8kit and LDH kit.Secondly,the cells were exposed to 1 mM and 2 mM Pb?Ac?₂ for 24h,respectively,and effects of different concentrations of Pb?Ac?₂ on mitophagy and endoplasmic reticulum stress were detected by Western blot.Finally,the mitophagy was detected by Western blot under the condition that endoplasmic reticulum stress was inhibited by inhibitor 4-phenylbutyric acid?4-PBA?.The role and mechanism of endoplasmic reticulum stress in Pb?Ac?₂ induced mitophagy was discussed,which provide theoretical reference for further understanding the Pb cytotoxicity and the molecular mechanism of mitophagy induced by Pb.The results showed that:1.Pb?Ac?₂?0,0.5,1,2,4 mM?decreased the cell viability and induced cell damage after exposure to HEK293 cells for 24 h in a dose-dependent manner.2.After Pb?Ac?₂?1 mM and 2 mM?exposure to HEK293 cells for 24 h,the ex-pression of LC3-?were significantly increased in all treatment groups?p<0.05?,and the expression of HSP60 and COX IV were significantly decreased in 2 mM Pb?Ac?₂treatment group?p<0.01?.However,there was no significant change in the expression level of p62 protein.The results indicated that Pb?Ac?₂ exposure could induce mi-tophagy in HEK293 cells.3.After Pb?Ac?₂?1 mM and 2 mM?exposure to HEK293 cells for 24 h,the ex-pression levels of ATF6,IRE1?,CHOP and GRP78 were gradually increased in a dose-dependent manner.In 2 mM treatment group,the expression levels of IRE1?,CHOP and GRP78 were significantly higher than control group?p<0.05?.The results indicated that Pb?Ac?₂ exposure could induce endoplasmic reticulum stress in HEK293 cells.4.When the endoplasmic reticulum stress was inhibited by 0.5 mM 4-PBA,the expression levels of HSP60 and COX IV in the 4-PBA+2 mM Pb?Ac?₂ treatment group were significantly higher than 2 mM Pb?Ac?₂ treatment group?p<0.05?,and the expression levels of LC3-?were obviously decreased.The results suggested that Pb?Ac?₂ induced mitophagy through activating the endoplasmic reticulum stress in HEK293 cells.In summary,the results showed that Pb exposure could reduce cell viability and induce cell damage.Moreever,Pb could not only induce mitophagy,but also induce endoplasmic reticulum stress.Endoplasmic reticulum stress may play a regulatory role in the upstream of mitophagy and Pb may induce mitophagy by activating endo-plasmic reticulum stress.