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目的:探讨Six1、转化生长因子β(TGF-β)在人喉鳞状细胞癌裸鼠移植瘤淋巴转移中对血管内皮生长因子C(VEGF-C)的作用。方法:利用基因干扰技术,分别沉默人喉鳞状细胞癌细胞内Six1和TGF-β基因的表达,制备Six1和TGF-β靶向siRNA转染至喉鳞状细胞癌细胞,筛选出阳性克隆细胞进行扩增,制备荷瘤裸鼠模型。致瘤小鼠随机分为5组:A组(未转染组),B组(转染空载体组),C组(Six1转染组),D组(TGF-β转染组),E组(Six1+TGF-β转染组)。肿瘤生长到6~12mm时,将裸鼠脱颈处死,观察并记录各组肿瘤的体积及转移情况。Western blot及免疫组织化学检测各组Six1、TGF-β及VEGF-C蛋白的表达;RT-PCR检测各组Six1、TGF-β及VEGF-C mRNA的表达。结果:A、B组比较,平均肿瘤体积及转移只数无明显差异;同A组比较,C、D、E组中平均肿瘤体积无明显减轻,但转移只数减少;与C、D组比较,E组平均肿瘤体积无明显减轻,转移只数无明显减少。Western blot、免疫组织化学及RT-PCR结果显示,A、B组比较,VEGF-C蛋白及其mRNA表达无明显变化;与A组比较,C、D、E组中VEGF-C蛋白及其mRNA表达均出现减弱;与C、D组比较,VEGF-C蛋白及其mRNA的表达在E组未出现进一步降低。结论:Six1和TGF-β均能够调控VEGF-C mRNA及其蛋白的表达,进而调控人喉鳞状细胞癌的淋巴转移,提示Six1、TGF-β的变化可能成为临床抑制人喉鳞状细胞癌淋巴转移的潜在靶点。
Objective: To investigate the effect of Six1 and transforming growth factor β (TGF-β) on vascular endothelial growth factor C (VEGF-C) in human laryngeal squamous cell carcinoma xenografts in nude mice. Methods: Six1 and TGF-β genes were silenced in human laryngeal squamous cell carcinoma cell line by gene interference technique, and Six1 and TGF-β target siRNAs were transfected into laryngeal squamous cell carcinoma cells to screen positive clonal cells Amplification, preparation of tumor-bearing nude mice model. Tumor-bearing mice were randomly divided into 5 groups: group A (untransfected group), group B (empty vector transfected group), group C (Six1 transfected group), group D (TGF- Group (Six1 + TGF-β transfected group). Tumor growth to 6 ~ 12mm, the nude mice were sacrificed, the tumor volume and metastasis were observed and recorded. The expressions of Six1, TGF-β and VEGF-C in each group were detected by Western blot and immunohistochemistry. The expressions of Six1, TGF-β and VEGF-C mRNA in each group were detected by RT-PCR. Results: There was no significant difference in average tumor volume and number of metastasis between group A and group B. Compared with group A, the average tumor volume in groups C, D and E did not reduce obviously, but the number of metastasis decreased; Compared with group C and D The average tumor volume in group E was no significant reduction, no significant reduction in the number of metastases. Western blot, immunohistochemistry and RT-PCR results showed that the expression of VEGF-C protein and its mRNA had no significant changes in group A and group B. Compared with group A, VEGF-C protein and mRNA in group C, D and E The expression of VEGF-C protein and its mRNA did not appear to be further decreased in group E compared with group C and D. Conclusion: Both Six1 and TGF-β can regulate the expression of VEGF-C mRNA and its protein, and then regulate the lymphatic metastasis of human laryngeal squamous cell carcinoma, suggesting that the changes of Six1 and TGF-β may become the clinical inhibitor of human laryngeal squamous cell carcinoma Potential target for lymphatic metastasis.