腺病毒介导的IL-28B对HeLa细胞宫颈癌抗肿瘤作用的研究

发布时间：2018-09-12 08:16

【摘要】：目的探讨腺病毒载体介导的IL-28B(Ad-h IL-28B)对He La细胞宫颈癌的抗肿瘤作用的机制,通过构建Balb/C裸鼠He La细胞宫颈癌皮下移植瘤模型,探讨其对皮下移植瘤的影响及抗肿瘤作用。方法扩增腺病毒载体(Ad-h IL-28B和Ad-EGFP);测定病毒(Ad-h IL-28B和Ad-EGFP)滴度;体外培养宫颈癌He La细胞,测定MOI值;MTT实验及划痕实验检测Ad-h IL-28B作用He La细胞后,抑制细胞增殖情况;Western blot实验检测STAT3、Bax及Bc L-2蛋白的表达;构建Balb/C裸鼠He La细胞宫颈癌皮下移植瘤模型;观察并测定Ad-h IL-28B处理后Balb/C裸鼠He La细胞宫颈癌皮下移植瘤瘤体的生长情况;HE染色法检测裸鼠He La细胞宫颈癌皮下移植瘤瘤体、肺脏及肝脏组织的病理变化。结果1病毒(Ad-h IL-28B和Ad-EGFP)滴度均为109PFU/100u L,He La细胞MOI测定值为10。2 MTT结果表明Ad-h IL-28B组明显较Ad-EGFP组和PBS对照组吸光度值低,IL-28B对He La细胞增殖有明显抑制作用(F=7.707,P=0.021)。划痕实验结果显示Ad-h IL-28B组于处理后,生长缓慢,划痕与对照组及Ad-EGFP组比较,明显增宽,可见IL-28B能显著抑制He La细胞的增殖。3成功建立Balb/C裸鼠He La细胞宫颈癌皮下移植瘤模型。4与PBS组及Ad-EGFP组比,Ad-h IL-28B组裸鼠移植瘤的体积最小(F=437.597,P0.001),表明Ad-h IL-28B能够明显减小移植瘤的体积,抑制移植瘤的生长。5 HE染色结果表明,Ad-h IL-28B组中肿瘤细胞的异型性数量明显减少,三组裸鼠的肺脏和肝脏组织无明显肿瘤转移。6 Western blot结果表明Ad-h IL-28B显著提升了促细胞凋亡的Bax蛋白(F=117.905,P0.001)的表达,却降低了抗凋亡蛋白Bc L-2蛋白(F=18.398,P=0.001)及STAT3蛋白(F=12.356,P=0.003)的表达。结论1 Ad-h IL-28B能够抑制宫颈癌He La细胞的增殖。2 Ad-h IL-28B能够抑制裸鼠He La细胞宫颈癌皮下移植瘤的生长,抑瘤效果明显。3 Ad-h IL-28B可能通过JAK-STAT途径调节STAT3蛋白、Bc L-2、Bax蛋白水平,进而影响宫颈癌He La细胞的增殖及移植瘤的生长。
[Abstract]:Objective to investigate the antitumor mechanism of adenovirus mediated IL-28B (Ad-h IL-28B) on cervical carcinoma of He La cells. To investigate the effect and antitumor effect of adenovirus vector mediated IL-28B (Ad-h IL-28B) on subcutaneous transplanted tumor of cervical cancer in Balb/C nude mice. Methods the adenovirus vectors (Ad-h IL-28B and Ad-EGFP) were amplified, the titers of Ad-h IL-28B and Ad-EGFP were measured, and the He La cells of cervical cancer were cultured in vitro. The expression of STAT3,Bax and Bc L-2 protein was detected by Western blot assay, and the subcutaneous transplanted tumor model of cervical cancer in He La cells of Balb/C nude mice was established. To observe and measure the growth of He La subcutaneous transplanted tumor of Balb/C nude mice treated with Ad-h IL-28B. The pathological changes of tumor, lung and liver of He La cells in nude mice were detected by HE staining. Results (1) the titer of Ad-h IL-28B and Ad-EGFP were both 109PFU/100u Lhe he La cell MOI. The results showed that the absorbance of Ad-h IL-28B group was significantly lower than that of Ad-EGFP group and PBS control group. IL-28B significantly inhibited the proliferation of He La cells (F7. 707, P0. 021). The results of scratch test showed that the growth of Ad-h IL-28B group was slow after treatment. Compared with the control group and Ad-EGFP group, the scratches were significantly wider. It can be seen that IL-28B can significantly inhibit the proliferation of He La cells. 3 successful establishment of subcutaneous transplanted tumor model of He La cells in Balb/C nude mice compared with PBS group and Ad-EGFP group, the volume of transplanted tumor of nude mice in Ad-h IL-28B group is the smallest (FF437.597 P0.001), which indicates that Ad-h IL-28B can obviously reduce the volume of transplanted tumor. The results of 5. 5 HE staining showed that the number of tumor cells in Ad-h IL-28B group was significantly decreased. No significant tumor metastasis was found in lung and liver tissues of the three groups. The results showed that Ad-h IL-28B significantly increased the expression of apoptosis-promoting Bax protein (Fn117.905, P0.001), but decreased the expression of Bc L-2 protein and STAT3 protein (FN 12.356P0.003). Conclusion 1 Ad-h IL-28B can inhibit the proliferation of cervical cancer He La cells. 2 Ad-h IL-28B can inhibit the growth of cervical cancer subcutaneous transplanted tumor of He La cells in nude mice. The inhibitory effect of 3. 3 Ad-h IL-28B may regulate the level of STAT3 protein Bc- 2, Bax through JAK-STAT pathway. Thus, the proliferation of cervical cancer He La cells and the growth of transplanted tumor were affected.
【学位授予单位】：华北理工大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R737.33

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