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目的研究沉默TCA8113舌癌细胞整合素连接激酶(integrin-linked kinase,ILK)基因的表达,对其下游底物Akt和GSK3β磷酸化状态及对移植瘤生长和自发性转移的影响。方法通过Lipofectamine 2000介导将特异性siILK表达载体和无同源性的对照载体,稳定转染人舌鳞癌TCA8113细胞,分为TCA8113组、TCA8113 vector组和TCA8113 siILK组,将3组细胞分别注入裸鼠皮下构建移植瘤模型,5周后检测瘤质量,对裸鼠肺及瘤组织形态学行常规病理学检查,使用免疫荧光技术检测体外和体内癌细胞p-Akt和p-GSK3β等的表达,并观察瘤组织内血管生成情况。结果 TCA8113组、TCA8113 vector组肺组织均发生自发性转移瘤,TCA8113 siILK组肺组织未发现自发性转移瘤;TCA8113 siILK组移植瘤细胞形态较其他2组体积减小,核分裂象较少,核浆比例缩小;TCA8113 siILK组在体内和体外实验中的p-Akt和p-GSK3β表达较其他2组均明显下降(P<0.05);TCA8113 siILK组移植瘤质量较其他2组显著降低(P<0.05),抑瘤率分别为84%和92%;TCA8113 siILK组移植瘤血管生成较其他2组也明显减少(P<0.05)。结论沉默TCA8113舌癌细胞ILK表达可抑制其下游信号传导分子Akt和GSK3β的磷酸化,并抑制移植瘤血管生成,从而抑制移植瘤的生长。
Objective To study the expression of integrin-linked kinase (ILK) gene in the silenced TCA8113 tongue cancer cells and its effect on the phosphorylation status of Akt and GSK3β and its effect on the growth and spontaneous metastasis of xenografts. Methods Specific siILK expression vector and control vector without homology were transfected into TCA8113 human tongue squamous cell carcinoma cell line by Lipofectamine 2000 and divided into TCA8113 group, TCA8113 vector group and TCA8113 siILK group. Three groups of cells were injected The tumor model was established subcutaneously in nude mice. The tumor mass was detected 5 weeks later. The lung and tumor morphology of nude mice were examined by routine pathology. The expression of p-Akt and p-GSK3β in cancer cells in vitro and in vivo were detected by immunofluorescence , And observe angiogenesis in tumor tissue. Results No spontaneous metastases were found in the lungs of TCA8113 and TCA8113 vector groups, and no spontaneous metastases were found in TCA8113 siILK group. The tumor size of TCA8113 siILK group was smaller than that of the other two groups, (P <0.05). The expression of p-Akt and p-GSK3β in TCA8113 siILK group was significantly lower than that in other two groups (P <0.05) ), Respectively. The tumor inhibition rates were 84% and 92% respectively. The angiogenesis in TCA8113 siILK group was also significantly decreased compared with the other two groups (P <0.05). Conclusion ILK expression in silencing TCA8113 tongue cancer cells inhibits the phosphorylation of downstream signaling molecules Akt and GSK3β and inhibits xenograft angiogenesis and thus inhibits the growth of xenografted tumors.