Effect of Endo G over-expression on cadmium induced apoptosis of HEK-293 cells

ObjectiveTo examine the effects of over-expression of endonuclease G(Endo G)gene on cadmiuminduced apoptosis of embryonic kidney cell HEK-293 in vitro.MethodsTotal RNA was obtained from human hepatocarcinoma cell line(HepG 2)and cDNA was obtained by reverse transcription.The Endo G gene was amplified by PCR.The Endo G was inserted reversely into pcDNA 3.0,and the recombinant plasmid pcDNA 3.0-Endo G was selected and then transferred into DH5αstrain.The pcDNA 3.0-Endo G was transfected into HEK-293 cells.Western blot was used to confirm the expression of Endo G gene in HEK-293 cells.Acridine orange/ethidium bromide(AO/EB)and flow cytometry were used to determine the effect of Endo G gene on cadmium-induced apoptosis of HEK-293 cells.The cell proliferation was detected by methyl thiazolyl tetrazolium(MTT)method.Results The size of Endo G gene fragment is about 1100 bp.The eukaryotic expressing plasmid of pcDNA 3.0-Endo G was constructed successfully.The expression of Endo G was first increased and then decreased in HEK-293 cells with the increase of concentration of cadmium chloride.The study with AO/EB and flow cytometry showed that the apoptotic rate of HEK-293 cells over-expressing Endo G gene was over 37% and significantly increased.ConclusionEndo G induces apoptosis of HEK-293 cells with the main form of early and late apoptosis.