CdCl2 induces autophagy and apoptosis in HEK293 cells / 中国药理学与毒理学杂志

ABSTRACT

OBJECTIVE To evaluate the possibility that CdCl2 induces autophagy and apoptosis in HEK293 cells,and the role of extracellular regulated protein kinases (ERK1/2) and AKT proteins in autophagy. METHODS Green fluorescence protein(GFP)-light chain 3B(LC3B)expression plasmid was transfected into HEK293 cells. After 24 h,HEK293 cells were induced with CdCl2 2,4,8 and 10μmol·L-1 for 12 h. The expression of GFP-LC3B was detected by fluorescent microscopy. HEK293 cells were induced with CdCl2 2,4,8 and 10μmol · L-1 without transfection of GFP-LC3B for 12 h while autophagic vacuoles were observed by transmission electron microscopy. The expression of LC3B-Ⅱ/Ⅰproteins and the phosphorylation levels of ERK1/2 and AKT were analyzed by Western blotting. Apoptosis was detected by flow cytometry microscopy. HEK293 cells were treated with 3-MA 20μmol · L-1+CdCl2 10 μmol · L-1 for 12 h before cleaved caspase 3 protein was detected by Western blotting. RESULTS When HEK293 cells were exposed to CdCl2(≤10μmol · L-1)for 12 h,cytoplasmic GFP-LC3B punctuates were observed under the fluorescence microscope,and autophagic vacuoles were observed under an electron microscope. The expression of LC3B-Ⅱ/Ⅰ,p-ERK1/2 and p-AKT proteins was significantly increased in CdCl2-induced cells(P<0.05,P<0.01). Moreover,apoptosis was observed. The addition of 3-MA 20μmol · L-1+CdCl2 10μmol · L-1 enhanced apoptosis. Cleaved capase 3 protein expression was significantly increased(P<0.01). CONCLUSION CdCl2(≤10μmol·L-1)can induce autophagy in HEK293 cells. ERK1/2 and AKT proteins might be associated with the activation of autophagy that is accompanied by apoptosis,suggesting that autophagy can inhibit apoptosis at certain concentrations of CdCl2.