Research On The Mechanisms Based On KLF6 Gene Of Inducing TCA8113 Cells To Differenciation

Objective To understand the role and mechanisms based on KLF6 gene of inducing TCA 8113 cells to differenciation at cellular and molecular level, and to discuss the possibility of KLF6 as a new specific target anti- cancerization of tongue cells.Methods The growth and cell morphology of TCA8113 cells were observed by MTT and under microscope .The expression of KLF6 mRNA were detect by RT-PCR. The expression of KLF6 protein,p53, c-Jun and CyclinD1 were detect by immunohistochemistry.Results After inducing by HMBA,the number of adherent cells decreased obviously along with the concentration of HMBA. The growth inhibition on TCA 8113 cells was in a concentration-time effect relationship by MTT after treating by HMBA. That means the effect of inhibition increased gradually along with the concentration of inducing agent increases, and the time extended. There were some reversal features of developing to normal cells morphologically after induced by HMBA, such as:cell morphology convergence, cell volume increased obviously, and spread out, the proportion of nucleus and cytoplasm decreased, smaller nucleus, the shape of nuclear in most cells became oval, the number of nucleolus reduced obviously,cytoplasm was abundant, the color of total cells uniformed by HE stain and so on. The cell cycle was analysed by flow cytometry. The rate of G1 phase cells was 52.0% before treating by HMBA. The rate of S phase cells was 34.0%, and 14.0% of G2 phase cells. After treating by HMBA,with the time increasing, the number of G1 phase cells increased obviously, while the number of S phase cells reduced significantly,so did the number of G2 phase cells. The cell cycle was obviously arrested in G1 phase(p<0.05).There was apoptosis peak appeared by flow cytometry. The expression of KLF6 mRNA increased significantly after inducing by HMBA by RT-PCR(p<0.05). So did the KLF6 protein and p53 by immunocytochemistry(p<0.05).But the expression of c-Jun and Cyclin D1 were decreased significantly(p<0.05).That means the expression of KLF6 and p53 upregulation after HMBA treatment, while the expression of c-Jun and Cyclin D1 were downregulated.Conclusion Concluding from the results above,the cell proliferation of TCA 8113 was inhibited, and induced to differentiation and apoptosis after treating by HMBA.The mechanism based on KLF6 gene of inducing TCA 8113 cell to differenciation may be upregulated the expression of KLF6 mRNA, increased the expression of KLF6 protein,upregulated p53 gene, downregulated c-Jun and Cyclin D1.Then the inhibition on p21 by c-jun may be released by KLF6.And the complex of Cdk4/6 and Cyclin D1 was inhibited and inactived, so that the cell cycle was arrested in G0/G1 period and to resume TCA 8113 cell to normal and apoptosis at last.