HMBA对人舌癌Tca8113细胞增殖和KLF6及相关蛋白表达的影响

目的: 研究六亚甲基二乙酰胺(hexamethylene bisacetamide, HMBA)对人舌癌Tca8113细胞增殖和Kruppel样因子6(Kruppel-like factor 6, KLF6)及相关蛋白表达的影响。方法: HMBA处理Tca8113细胞后,观察细胞生长和细胞形态;流式细胞术观察细胞周期;RT-PCR检测KLF6 mRNA的表达;免疫细胞化学技术检测KLF6、p53、cyclin D1及c-Jun的蛋白表达。结果: HMBA处理后贴壁细胞的数量随浓度增加明显减少;MTT实验显示其生长抑制作用呈浓度/时间-效应关系;镜下观察显示处理后Tca8113细胞出现向正常细胞形态逆转演变的特征。流式细胞术检测结果显示,对照组Tca8113细胞G₁期比例为(52.00±0.02)%,S期为(34.00±0.08)%,G期为(14.00±0.10)%;HMBA处理后,随作用时间增加,G₁期细胞显著增加,S期细胞显著减少,G₂期细胞也减少,细胞明显被阻滞于G₁期(P<0.05);出现凋亡峰。 RT-PCR结果显示HMBA处理后KLF6 mRNA的表达上调(P<0.05); 免疫组化检测显示HMBA处理后KLF6和p53蛋白表达上调(P<0.05),cyclin D1和c-Jun表达下调(P<0.05)。结论: HMBA对Tca8113细胞增殖具有抑制作用;其作用机制一方面可能通过p53上调p21的表达和活性,另一方面可能通过上调表达的KLF6解除c-Jun对p21的抑制作用,下调cyclin D1,抑制CDK4/6和cyclin D1复合物活性,将Tca8113细胞阻滞于G₀/G₁期,诱导Tca8113细胞向正常细胞分化,恢复正常细胞的表型和功能,并促进细胞凋亡。

Effect of HMBA on proliferation and expression of KLF6 and related proteins in human tongue carcinoma Tca8113 cells

AIM:To study the effect of hexamethylene bisacetamide（HMBA） on the proliferation and expression of KLF6 and related proteins in human tongue carcinoma Tca8113 cells.METHODS;After cultured with HMBA,the growth of the Tca8113 cells was assayed by MTT method,and the morphology of the cells was observed under microscope. The cell cycle was determined by flow cytometry.The mRNA expression of KLF6 was detected by RT-PCR.The protein levels of KLF6,p53,cyclin D1 and c-Jun were measured by the method of immunohistochemistry.RESULTS:The number of adherent cells obviously decreased along with the concentration of HMBA,and the growth inhibition of Tca8113 cells was in a concentration/time-effect relationship after treated with HMBA.Some reversal features of the Tca8113 cells developed to normal cells in morphology after induced by HMBA.The proportion of the cells in G₁ phase was（52.00±0.02）%before treating with HMBA.The proportion of the cells in S phase was（34.00±0.08）%,and（14.00±0.10 ）%of G₂ phase cells.After treated with HMBA,the cell number in G₁ phase significandy increased with the exposure time going on,while the cell number in S phase significantly reduced,so did the cell number in G₂ phase.The cell cycle was significantly arrested in G₁ phase（P<0.05）.The apoptosis peak also appeared.The mRNA expression of KLF6 significantly increased after induced by HMBA（P < 0.05）,so did the protein levels of KLF6 and p53（P < 0.05）,while the expression of cyclin D1 and c-Jun was significantly decreased（P<0.05）.CONCLUSION;HMBA inhibits the proliferation of Tca8113 cells by arresting the cell cycle in G₀/G₁ phase and resuming Tca8113 cells to normal and apoptosis at last.