A study of serum-free cultivation of human hepatocyte (C3A) cell line

Objective To develop a serum-free culture condition for the cultivation of C3A human hepatocyte cell line.Methods C3A cells were cultivated under a stepwise condition of reducing the serum concentration from 10%,5%,and 2.5% to 0%.The cells cultured in serum-supplemented medium(SSM) served as the positive control and the cells cultured in basic medium served as the blank control.Cellular morphology was observed under an optical microscope,and the growth of C3A cell was tested by using CCK-8.Automatic biochemical analyzer was used to measure the level of ALT and urea in the culture supernatant.In addition,radioimmunoassay and real-time qPCR were adopted to analyze albumin secretion and gene expression.Results Cells began to growth after culturing for 24 h and the growth rates between the experimental group and the positive control groups were similar.The concentrations of urea in the SFM groups were significantly higher than the positive and blank control groups(P0.001) at day-2 after the cultivation.There was also a significant difference between the SFM groups and the control group in the albumin secretion.There was no significant difference in the gene expression of UGT,GST,AFP and ALB between the SFM groups and the positive control group(P=0.640,0.075,0.504,and 1.000,respectively),but the expressions of CK18,CK19 and HNF4α were significantly lower than the positive control group(P 0.001).Conclusion A serum-free culture condition was developed for the cultivation of human C3A cells.