Mitochondrial DNA alterations analysis in human hepatocellular carcinoma cells

Abstract

Objective To compare mitochondrial DNA alterations in hepatocellular carcinoma cells and the relationship between mitochondrial DNA alterations and tumorigenesis,serum starvation tolerance and chemotherapy sensitivity.Methods Direct DNA sequencing methods were used to detect the variants of the mtDNA in human hepatocellular carcinoma cells.Cell viability and cell growth inhibition rate of cisplatin were tested by methyl thiazol tetrazolium (MTT).Tumorigenesis was examined by subcutaneously inoculating hepatocellular carcinoma cells in nude mice.Results Numbers of single variant in the mitochondrial DNA D-Loop region were:hepatic stellate cells (HSC,16),BEL-7402 (11),HepG2 (11) Numbers of single variant in the COX Ⅱ gene were:HSC (2),BEL-7402 (1),HepG2 (3) ; a 8-bp fragment deletion were detected in COX Ⅱ gene of HepG2 cells.The level of mtDNA was significantly decreased in HSC and HepG2 cells as compared to BEL-7402 cells.Cell viability was significantly higher in HSC and BEL-7402 cells as compared to HepG2 cells.The serum starvation tolerance ability was significantly decreased in BEL-7402 cells as compared to HepG2 and HSC cells.The cell growth inhibition rate of cisplatin was significantly increased in BEL-7402 cells as compared to HepG2 and HSC cells.Higher tumorigenesis ability was detected in HSC and BEL-7402 cells,no tumorigenesis was observed in HepG2 cells after subcutaneously inoculated in nude mice for 23 days.Conclusion Taken these results together,we suggest that serious variations in COX Ⅱ gene may contribute to HepG2 cells' low tumorigenesis ability.There is a negative correlation between mitochondrial DNA copy number and serum starvation ability,additionally; there is positive correlation between mitochondrial DNA copy number and sensitivity to cisplatin of hepatocellular carcinoma cells. Key words: Hepatocellular carcinoma cells; Mitochondrial DNA; Gene variant