The Effect of Astemizole on Autophagy of HeLa Cells

Abstract: To investigate the effects of Astemizole (AST) on regulating autophagy in HeLa cells， we used Western blot， mCherry-EGFP tandem fluorescent-tagged LC3B (microtubule-associated proteins 1A/1B light chain 3B， LC3) stable expression system， mt-Keima based living cell fluorescence imaging method in this study. The results from Western blot showed that AST resulted in dose- and time-dependent accumulation of LC3-II and SQSTM1/p62 in HeLa cells. And AST could not facilitate the further accumulation of LC3-II in combination with Bafilomycin A1 (Baf-A1)， a blocker of autophagic flux. MTS assay presented that AST promoted cell death. The mCherry-EGFP-LC3B fluorescence system indicated that HeLa cells exposed to AST displayed greatly accumulation of autophagosomes as well as autolysosomes. Furthermore， the generic mitochondrial membrane depolarizer CCCP (carbonyl cyanide 3-chlorophenylhydrazone) could induce mitochondrial autophagy， or mitophagy， which accompanied by the rapid degradation of mitochondrial outer membrane marker TOMM20 (translocase of the outer membrane 20). The result from Western blot showed that AST inhibited CCCPinduced TOMM20 degradation as chloroquine (CQ) and Baf-A1 did. In addition， the mitochondrial autophagy was obviously reduced in cells treated with AST detected by mt-Keima. These results implied that AST could significantly inhibit autophagic flux and the clearance of damaged mitochondria in HeLa cells.

CSTR: 32200.14.cjcb.2018.04.0008