Effect of p53 gene on p-glycoprotein expression and chemotherapeutic cytotoxicity of hepatocellular carcinoma cell line Hep3B / 中国病理生理杂志

ABSTRACT

AIM:To test the hypothesis that wild-type p53 regulates the expression of p-glycoprotein. METHODS: While Hep3B cells lack expression of both p53 and retinoblastoma tumor suppressor genes because of deletions, were transfected to a wild-type (wt) p53 cDNA and control vector by a liposome method. After G418 selection, stable wt-p53 transformants, and control vector transformants (pNeo) were obtained. The expression of the p53,p21 and P-gp was analyzed by Northern or Western blot methods. In both transformants, cytotoxic effect of doxorubicin and mitomycin was evaluated by MTT assay, and accumulation of doxorubicin was detected by flow cytometer. RESULTS: In wt-p53 transformants, induction of transcriptionally active p53 was confirmed by the increase of P21 waf1/cip1 protein. Levels of P-gp reduced in the cells expressing wild-type p53 were linked to wt-p53 activity. The wt-p53 transfectants were more sensitive to doxorubicin and mitomycin compared with the pNeo transformants, and the accumulation of doxorubicin in the wt-p53 transfectants was 13 times as much as pNeo transformants. After both transformants were treated with doxorubicin at 0.2 mg/L for 24 hours, apparent apoptosis occurred in the wt-p53 transrormants. CONCLUSION: Restoration of wt-p53 activity in Hep3B leads to more sensitive to chemotherapeutic agents because of decrease of p-glycoprotein expression.