Role Of P38MAPK In Cell Proliferation And Apoptosis In Human Hepatocarcinogenesis

Objectives: To investigate the role of p38MAPK in cell proliferation and apoptosisin human hepatocarcinogenesis and study the mechanism of carcinogenesis of HCC and the methods of early pathologic diagnosis of HCC and the strategy to prevent or treat HCC.Methods: There were the tissues of HCC and chronic lesions related with HCC(chronic hepatitis, liver cirrhosis and paratumor cirrhosis) and cultured cell lines (the normal liver cell line HI-7702, the paratumor cirrhosis cell line QSG-7701 and the HCC cell line QGY-7703) . In specimens of tissues, the expressions of HBx protein, p38MAPK, cell cycle G2/M phase related factors (cdc25B, p34cdc2,cyclin B1) , anti-apoptosis factor (survivin) and cell proliferation factor (ki-67) were detected by immunohistochemical and immunocytochemical staining, confocal microscope, electronic microscope, Western Blot. In cultured cells, all targets were observed after cultured cells were treated with DDP (diamminedichloroplatin) and the p38MAPK inhibitor SB203580．Flow cytometer and TUNEL method were used to detect apoptosis and cell cycle.Results: The positive rate of HBx protein in HCC was higher than that in otherelse. In tissues, the positive rates of p38MAPK, cdc25B, cyclin B1, p34cdc2 and survivin increased gradually from normal liver, chronic hepatits, liver cirrhosis, paratumor cirrhosis to HCC. In cultured cells, the positive rates of these targets also had the same trend from normal liver, paratumor cirrhosis to HCC. The proliferation index (PI) was positively correlated with HCC histological grade. Contrary to PI, the apoptosis index (AI) was inversely correlated with the grade. Decreased significantly the apoptotic rates in normal liver cells and HCC cells and increased the rate in paratumor cirrhosis cells were observed through TUNEL method and Flow cytometer after cultured cells were treated with SB2030580．The apoptotic rates in three cell lines pretreated with DDP all increased obviously, and then after SB2030580 treatment, the rates in normal liver cells and HCC cells continued increasing, while in paratumor cirrhosis cells the rate decreased and the cell cycle stopped at S phase.Conclusions:1．It was a novel finding that all the data (including the positive rates ofp38MPK, cell cycle G2/M phase related factors, survivin , ki-67, PI and AI) in paratumor cirrhosis were between those in cirrhosis without HCC and those in HCC, which indicated paratumor cirrhosis cells belonged to an essentially different group from cirrhosis cells without HCC, and they had more feature of a precancerous lesion. It implied paratumor cirrhosis may be the key stage in malignant transformation of hepatic cells. In the paratumor cirrhosis cell line QSG-7701, many items, including the situation of cell cycle, the apoptotic rate, the expressions of p38MAPK and phosphorylated p38MAPK (p-p38) , cell cycle G2/M phase related factors (cdc25B, p34cdc2 and cyclin B1) , survivin and ki-67, and AI, as well as the responses to SB2030580 and DDP, were different from those in the normal liver cell line HL7702 and those in the HCC cell line QGY-7703．The paratumor cirrhosis cells had an individual biology characteristic, and the growth curve of them was similar to that of HCC cells, which demonstrated paratumor cirrhosis cells belonged to precancerous cells transforming.2．In the process of hepatocarcinogenesis, HBx caused a series of abnormal changes of cell cycle, cell apoptosis and proliferation by activating p38MAPK pathway. In chronic hepatitis and liver cirrhosis, p38MAPK found in nucleus induced G2/M phase arrest by inhibiting the activity of cdc25B. In G2/M phase, DNA damage caused by HBV was repaired, but if the repair failed, it would result in apoptosis. In paratumor cirrhosis and HCC, p38MAPK seen in cytoplasm lost the function of inhibiting the activity of cdc25B. Cdc25B activated p34cdc2 by dephosphorylation, and then activated p34cdc2 was transported into nucleus and activated MPF. Activated MPF promoted cell cycle from G2 phase to M phase, causing cells to divide and proliferate continuously, resulting in carcinogenesis of HCC gradually. The results suggested the expression of cdc25B protein was related to the malignant transformation of HCC.3．Survivin played a vital role in carcinogenesis and progress of HCC. The mechanism may be that survivin inhibited the activity of p38MAPK, following the inhibition of apoptosis and the promotion of cell proliferation, resulting in hepatocarcinogenesis.4．In the process of carcinogenesis and progress of HCC, cell proliferation and apoptosis occurred together, but the extent of cell proliferation was more than that of apoptosis. The higher level of cell proliferation promoted carcinogenesis and progress of HCC. Apoptosis of HCC cells was inversely correlated with the HCC histological grade, whereas proliferation of HCC cells was positively correlated with the grade.5．DDP induced apoptosis, which involved the p38MAPK pathway.