高密度接种对转化生长因子β1诱导上皮间充质转化的影响**

摘要

BACKGROUND: The cel density is one of the factors involved in the state of cel differentiation, and the effect of cel density on transforming growth factor-β1-induced epithelial-mesenchymal transition of HaCaT cells is stil unclear. OBJECTIVE: To observe the effect of cel density on transforming growth factor-β1-induced epithelial-mesenchymal transition of HaCaT cells. METHODS: HaCaT cells was seeded in 6-wel plates at low density of 103/cm2 and high density of 105/cm2 then treated by 2 μg/L transforming growth factor-β1 for 48 hours, thereafter observed the changes in cel morphology. The transcription levels of epithelial cadherin, tight junction protein-1, vimentin, neuronal-cadherin were detected by real-time PCR, and expression levels of epithelial cadherin and vimentin were detected by Western blot. RESULTS AND CONCLUSION: Cel gap of HaCaT cells grew larger after treated with transforming growth factor-β1 for 48 hours, and cel morphology was long spindle rather than polygonal in low-density group, while in high-density group without obvious morphological changes. The real-time PCR showed that the transcriptions of epithelial cel marker epithelial cadherin and tight junction protein-1 were suppressed when compared with the control group (P < 0.05), and the decreasing deree in the high-density group was higher than that in the low-density group (P <0.05), mesenchymal cel marker neuronal-cadherin and vimentin were upregulated in the high-density group and the low-density group when compared with the control group (P < 0.05), and there were no significant differences between high-density group and low-density group. Western blot results verified the changes of neuronal-cadherin and vimentin expression level. These results suggest that the high seeding density can inhibit transforming growth factor-β1-induced epithelial-mesenchymal transition of HaCaT cells.%　　背景：细胞密度是影响细胞分化状态的因素之一，对于细胞密度在转化生长因子β1诱导 HaCaT 细胞上皮间充质转化中的作用尚缺乏深入研究。目的：观察细胞密度对转化生长因子β1诱导 HaCaT 细胞上皮间充质转化的影响。方法：将HaCaT细胞分别以低密度103/cm2和高密度105/cm2作用48 h，观察细胞形态变化，应用Realtime PCR检测上皮型钙黏蛋白、紧密连接蛋白1和波形蛋白、神经型钙黏蛋白的转录水平，Western blot检测上皮型钙黏蛋白和波形蛋白表达水平。结果与结论：低密度组的 HaCaT 细胞在转化生长因子β1处理48 h 后细胞间隙变大，细胞形态由多角形变为长梭形，高密度组的细胞间隙变大，但形态变化不明显；Realtime PCR 结果显示两组上皮细胞标志物上皮型钙黏蛋白和紧密连接蛋白1的转录均较对照组明显下调(P <0.05)，但高密度组没有低密度组下调明显(P <0.05)，间充质细胞标志物神经型钙黏蛋白和波形蛋白的转录均较对照组明显上调(P <0.05)，而高密度组与低密度组间差异无显著性意义；Western blot 进一步验证了上述上皮型钙黏蛋白和波形蛋白的变化。说明高接种密度抑制转化生长因子β1诱导 HaCaT 细胞上皮间充质转化。