科研之友
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摘要
OBJECTIVE Survivin is a new member of the apoptosis suppressor protein family. It is tumor-specific and only expressed in tumor and embryo tissues, and is closely related to the differentiation, proliferation and infiltration of tumor cells. Utilizing specific short hairpin RNA(shRNA) to silence the expression of Survivin gene of RL95-2 endometrial cancer cells and to investigate the effect of Survivin gene inhibition on growth and apoptosis of endometrial cancer cells derived transplantation tumor. METHODS The nude mice model of xenografted endometrial cancer and the Survivin gene targeted specific shRNA-Lentivirus plasmid were constructed. According to the experiment aim, all tumor-bearing mice were randomly divided into experimental group, control group and blank group. Survivin-shRNA Lentivirus plasmid was intratumorally injected in the experimental group, negative control shRNA Lentivirus plasmid was intratumorally injected in the control group, and the blank group was untreated. Changes of tumor volume in each group were monitored once a week and drew the tumor growth curve. After injection therapy for 4 weeks, nude mice were sacrificed and the expression of Survivin mRNA was measured with qPCR, and the expression of apoptosis-related protein, including Survivin protein and Caspase-3 protein, was measured with Western Blot. After injection therapy for 4 weeks, the cell apoptosis was tested by using Tunel staining and the histologic changes of tumor sites was detected by HE staining. RESULTS After 2 weeks of RL95-2 endometrial cancer cells transplantation in subcutaneous area of nude mice, tumor size is detected at about 1.0×0.8 cm. After injection therapy for 4 weeks, the results of qPCR showed Survivin mRNA expressions in the experimental group, control group and blank group were 0.247±0.035, 0.997±0.028 and 1.007±0.019, respectively, the mean difference was statistically significant among the three groups (F=795.623, P<0.001), and the expression level of Survivin mRNA in the experimental group was lower than that of the other two groups, and the difference was statistically significant (P<0.05); the results of Western Blot showed, the Survivin protein expression in experimental group, control group and blank group were 0.171±0.069, 0.598±0.176 and 0.176±0.125, respectively, the mean difference was statistically significant among the three groups (F=15.691, P=0.004) and the expression level of Survivin protein in the experimental group was lower than that of the other two groups, the difference was statistically significant (P<0.05); the Caspase-3 protein expression in experimental group, control group and blank group were 0.401±0.107, 0.093±0.018 and 0.018±0.026, respectively, the mean difference was statistically significant among the three groups (F=32.33, P=0.001). The expression level of Caspase-3 protein in the experimental group was higher than that of the other two groups, and the difference was statistically significant (P<0.05). Compared with the control group and the blank group, the cell apoptosis rate in the experimental group was obviously higher, P<0.05. Compare the tumor volume of each group, experimental group can significantly narrowed the tumor volume of transplanted endometrial cancer, at the fourth week, the average tumor volume was (0.46±0.07) cm 3 , and the tumor volume decreased by 33%, P<0.05. CONCLUSIONS Silencing Survivin gene expression by the Survivin-shRNA Lentivirus plasmid injection can inhibit the growth of transplantation tumor and induce apoptosis of endometrial cancer cells. Survivin might be a potential target for anti-endometrial cancer therapy.
