Abstract:

BACKGROUND: Bioartificial liver using porcine hepatocytes or hepatoma cells as sources of transplanted material encounter the danger of zoonotic disease or oncogenicity. The normal adult hepatocytes also have some limitations.
OBJECTIVE: To detect the survival rate, growth curve and cell cycle of normal adult hepatocytes before and after transfection, and to observe the effects of SV40 immortalized gene transfection on growth characteristics of normal adult hepatocyte.
METHODS: Cultured normal adult hepatocytes and immortalized normal adult hepatocytes were performed placenta blue staining and AO-PI staining. MTT staining was used to count the cell survival rate at days 1-8 after culture. MTT assay was used to measure absorbance value of cells, and to draw cell growth curves. The cell growth cycle was detected by flow cytometry.
RESULTS AND CONCLUSION: The three kinds of staining assay showed that the survival rates of the two kinds of cells were 95%-99%, which had no significant difference. The two normal adult hepatocytes had no obviously difference in growth curves, both of which presented as exponential growth at days 3-5 after culture, but the cells grew faster after transfection. Flow cytometry results demonstrated that: after transfection, 65.64% cells were in the S phase, 34.36% cells in the G0-G1 phase, and no cells in G0 phase; however, S phase cells accounted for 21.27%, G0-G1 phase cells accounted for 62.64%, G2 phase cells accounted for 12.09% before transfection. This suggested that the S-phase cells were markedly increased in hepatocytes after transfection, and the proliferation capacity was enhanced. Compared with normal adult hepatocytes, the SV40 immortalized gene trasfected cells exhibit stronger proliferative activity.