mda-7/IL-24 induces apoptosis of hepatic carcinoma cells through endoplasmic reticulum stress pathway / 第二军医大学学报

Objective: To study the effects of mda-7/IL-24 on the growth, proliferation, apoptosis of different hepatic carcinoma cell lines and the related mechanisms. Methods: A recombinant adenovirus Ad-mda-7 was constructed and was used to transfect human hepatic carcinoma cell lines (HepG2, Hep3B and PLC/ PRF/5) and normal liver cell line L02. MTT assay and FACS were employed to assess the growth and apoptosis of cells; the expression of related protein expression was examined by Western blotting. The cells were treated with calpastatin I (ALLN,25 μmol/L) for 30 min to block the endoplasmic reticulum stress (ER-stress) and the above indices were examined again. Results: Treatment with Ad-mda-7 resulted in selective inhibition of cell proliferation and induced apoptosis, especially in HepG2 cells; Ad-mda-7 showed no influence on normal cells. Pretreatment with ALLN partially inhibited the above effects of Ad-mda-7. Western blotting revealed that Ad-mda-7 induced up-regulation of BiP/GRP7B and Bax protein, activation of caspase-12, caspase-3 and phosphorylation of p38 MAPK in HepG2 cells. Blocking ER-stress with ALLN down-regulated Bax, caspase-12 expression and inhibited activation of caspase-3 and caspase-12, but showed no effect on the expression of BiP/GRP78 or phosphorylation of p38 MAPK. Conclusion: mda-7/IL-24 can cause growth inhibition and promote apoptosis of hepatic carcinoma cells through the ER-stress pathway.

In vitro Efficacy of mda-7 Gene for Hepatocellular Carcinoma Gene Therapy Mediated by Human Ribosomal DNA Targeting Vector / 生物化学与生物物理进展

Human ribosomal DNA (hrDNA) targeting vector pHr is a homologous recombinant plasmid for human genome which developed in the State Key Laboratory of Medical Genetics. pHr was used to construct a recombinant plasmid pHr-CMG expressing mda-7/GFP fusion gene and its efficacy in the hepatocellular carcinoma cell line Bel-7402 was investigated. The expression of mda-7/GFP fusion gene was detected by fluorescent microscope, RT-PCR and Western blotting, and its function was detected by cell-cycle analyses, MTT assay and Hoechst33258 staining. The results demonstrated that pHr-CMG vector could express MDA-7/GFP fusion protein effectually and the mda-7 gene could induce cell apoptosis and proliferation suppression in Bel-7402 cell line, which might be caused by the G2/M cell cycle arrest. These results also suggested that human ribosomal DNA targeting vector system and the pHr-CMG vector may be applied in further gene therapy researches for hepatocellular carcinoma.