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目的:探讨热休克蛋白70(HSP70)敲低对结肠癌细胞在裸鼠体内生长的影响。方法:构建两种干扰HSP70的shRNA(HSP70 shRNA-1和HSP70 shRNA-2)质粒表达载体,分别转染到结肠癌HT29细胞,并以空质粒转染(阴性对照)和未转染的HT29细胞(空白对照)为对照,用RT-PCR和Western blot方法检测各组HT29细胞HSP70基因和蛋白的表达;将HSP70shRNA-2转染、空质粒转染及未转染的HT29细胞分别接种至裸鼠皮下建立移3组植瘤模型,观察肿瘤生长情况,3周后剥离瘤块,用HE染色、免疫组化和TUNEL法分别检测移植瘤组织形态学、增殖细胞核抗原(PCNA)的表达及细胞凋亡情况。结果:RT-PCR和Western blot结果显示HSP70 shRNA-1和HSP70 shRNA-2均能明显抑制HT29细胞HSP70基因和蛋白的表达,且HSP70 shRNA-2的抑制作用更为明显,空质粒转染对HSP70的表达无明显影响;与空白对照组比较,HSP70 shRNA-2转染组裸鼠移植瘤生长明显较明显减慢(P<0.01),瘤组织中心出现明显坏死,PCNA表达明显降低,细胞凋亡率明显增加(P<0.01),而空质粒转染组无明显上述改变(P>0.05)。结论:HSP70沉默能抑制结肠癌HT29细胞裸鼠移植瘤的生长,促进细胞凋亡,HSP70可能是治疗结肠癌有效靶点。
Objective: To investigate the effect of heat shock protein 70 (HSP70) knockdown on the growth of colon cancer cells in nude mice. METHODS: Two plasmids expressing HSP70 and HSP70 shRNA-2 were constructed and transfected into HT29 cells respectively. The transfected cells were transfected with empty plasmid (negative control) and untransfected HT29 cells (Blank control) as control, the expression of HSP70 gene and protein in HT29 cells of each group was detected by RT-PCR and Western blot. The HSP70shRNA-2 transfected, empty plasmid transfected and untransfected HT29 cells were inoculated into nude mice Three groups of tumor models were established subcutaneously. The tumor growth was observed. The tumor masses were dissected after 3 weeks. HE staining, immunohistochemistry and TUNEL were used to detect the morphological changes, the expression of proliferating cell nuclear antigen (PCNA) Death situation. Results: The results of RT-PCR and Western blot showed that HSP70 shRNA-1 and HSP70 shRNA-2 can significantly inhibit the expression of HSP70 gene and protein in HT29 cells, and the inhibitory effect of HSP70 shRNA-2 was more obvious. Empty plasmid transfected HSP70 (P <0.01). Compared with the blank control group, the growth of xenografted tumors in HSP70 shRNA-2 transfection group was significantly slower than that in the blank control group (P <0.01). The tumor tissue showed obvious necrosis in the center, the expression of PCNA was significantly decreased, (P <0.01), while the blank plasmid transfected group had no obvious change (P> 0.05). Conclusion: Silencing HSP70 can inhibit the growth of colon cancer HT29 cells in nude mice and promote cell apoptosis. HSP70 may be an effective target for the treatment of colon cancer.