Establishment Of A Mice H22 High-lung-metastatic Hepatocellular Carcinoma Cell Line And Measurement Of The Biological Characteristics And RhoC Gene Expression

ObjectiveTo establish a mice high-metastasis H22 hepatocellular carcinoma cell line, and measure its biologic characters and RhoC gene expression to provide a suitable model for the study of metastasis-related molecular mechanisms.MethodsH22 hepatocellular carcinoma cells (MO for short) were inoculated into the abdominal cavity of mice, collect the abdominal caicinoma cells, inject them to the tail vein of another healthy mice and the metastatic lesions were harvested in the lung, then re-implanted into mice for the second time of in vivo selection. The same procedure was repeated until get the fourth cells(M4 shorted for). Then measure the metastatic ability of M4 cell in vivo, cell proliferation ability in vitro, calculate the cell doubling time and cell division index, observe the chromatosome after Giemsa stain and test the cell cycle by flow cytometry. At last measure the M4 andMO cell RhoC gene mRNA expression with the method of RT-PCR, then analyse it by the Imagequant software.ResultsComparing the MO cell with the high-metastasis cell M4,30 and 40 days after injecting carcinoma cell by tail vein, it is obviously easy to found that the velume of metastatic nodes of M4 cell in vivo is larger than M0. The concentration of M4 cells is larger than MO when they are cultered in vitro at the time point of 24h,36h,48h,60h 及72h. P<0.05, from3×10-9to 7.2×10-26. The cell doubling time of M0 is 71.98h, but M4 is 44.1 Oh, M4 is obviously shorter than MO.and cell division index of M4 is larger than M0(P=0.014). The high-metastasis cell proportion in G0/G1 stage is smaller than M0(P=0.024), in S stage is larger than M0(P=0.022). For the number of chromatosome, they are similar to each other, but there is obvious heteromorphism difference. The RhoC gene mRNA expression of high-metastatic cell is higher than M0; the relative expressive ratio of MO and M4 are 0.486±0.045and 1.011±0.163, (P=0.0029)ConclusionA mice high-lung metastasis hepatocellular carcinoma cell M4 is established. Comparing the MO cell, The metastatic ability of high-lung metastasis hepatocellular carcinoma cell is higher, growing speed is faster,the ratio of division cell is larger, there is obvious heteromorphism difference, proportion in G0/G1 stage is smaller but S stage is larger. The RhoC gene mRNA expression of high-metastatic cell line is higher, RhoC gene excessive expression is related to H22 hepatocellular carcinoma cell Line high-metastatic ability.