Melanoma differentiation-associated gene-7, MDA-7/IL-24, selectively induces growth suppression, apoptosis in human hepatocellular carcinoma cell line HepG2 by replication-incompetent adenovirus vector.
OBJECTIVE
To investigate the effect of replication-incompetent adenovirus vector expressing MDA-7/IL-24 on tumor growth and apoptosis in human hepatocellular carcinoma (HCC) cell line HepG2 and normal liver cell line L02.


METHODS
We constructed the recombinant replication-incompetent Ad.mda-7 virus vector and infected it into the human HCC cell line HepG2 and normal liver cell line L02. RT-PCR was performed to detect the mRNA expressing in cells. by ELISA was used to detect MDA-7/IL-24 protein expression in the culture supernatant. The effect of apoptosis induced by Ad.mda-7 was confirmed by Hoechst staining and flow cytometry assay with Annexin-V and PI staining. MTT assay was used to determine growth inhibition of HepG2 cells, and cell-cycle and hypodiploidy analyses were performed by flow cytometry.


RESULTS
Recombinant replication-defective virus expressing MDA-7/IL-24 was constructed successfully. RT-PCR showed that the Ad.mda-7 could mediate the expression of the exogenous gene MDA-7/IL-24 into HepG2 and L02. The concentration of MDA-7/IL-24 protein in supernatant was 130 pg/mL and 110 pg/mL in Ad.mda-7-infected L02 and HepG2 cells, respectively. Ad.mda-7 infection obviously induced apoptosis (from 2.60%+/-0.72% to 33.6%+/-13.2%, P=0.00012) and growth suppression in HepG2 (inhibition ratio IR=68%) and an increase in the percentage of specific cancer cell types at the G2/M phase of the cell cycle (from 6.44% to 32.29%, P<0.01), but not in L02 cells.


CONCLUSIONS
These results confirm selectively induction of apoptosis and growth suppression by the mda-7/IL-24 gene with replication-incompetent adenovirus vector in human hepatocellular carcinoma cell line HepG2.
Department of Biliary and Pancreatic Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, Hubei Province, China.
Cong-Jun
CJ
Wang
Xin-Bo
XB
Xue
Ji-Lin
JL
Yi
Kun
K
Chen
Jian-Wei
JW
Zheng
Jian
J
Wang
Jian-Ping
JP
Zeng
Rong-Hua
RH
Xu
D000256:Adenoviridae / Q000235:genetics; D017209:Apoptosis / Q000235:genetics*; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000473:pathology* / Q000628:therapy; D002453:Cell Cycle / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002470:Cell Survival / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic; D015967:Gene Expression Regulation, Viral; D015316:Genetic Therapy; D005822:Genetic Vectors; D022781:Hepatocytes; D006801:Humans; D007378:Interleukins / Q000235:genetics*; D008113:Liver Neoplasms / Q000235:genetics / Q000473:pathology* / Q000628:therapy; D008856:Microscopy, Fluorescence; D012333:RNA, Messenger; D012334:RNA, Neoplasm / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014779:Virus Replication; D000099180:Interleukin-24
D007378:Interleukins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D012367:RNA, Viral; D000099180:Interleukin-24