【摘要】：目的探討腺病毒載體介導(dǎo)的IL-28B(Ad-h IL-28B)對(duì)He La細(xì)胞宮頸癌的抗腫瘤作用的機(jī)制,通過(guò)構(gòu)建Balb/C裸鼠He La細(xì)胞宮頸癌皮下移植瘤模型,探討其對(duì)皮下移植瘤的影響及抗腫瘤作用。方法擴(kuò)增腺病毒載體(Ad-h IL-28B和Ad-EGFP);測(cè)定病毒(Ad-h IL-28B和Ad-EGFP)滴度;體外培養(yǎng)宮頸癌He La細(xì)胞,測(cè)定MOI值;MTT實(shí)驗(yàn)及劃痕實(shí)驗(yàn)檢測(cè)Ad-h IL-28B作用He La細(xì)胞后,抑制細(xì)胞增殖情況;Western blot實(shí)驗(yàn)檢測(cè)STAT3、Bax及Bc L-2蛋白的表達(dá);構(gòu)建Balb/C裸鼠He La細(xì)胞宮頸癌皮下移植瘤模型;觀察并測(cè)定Ad-h IL-28B處理后Balb/C裸鼠He La細(xì)胞宮頸癌皮下移植瘤瘤體的生長(zhǎng)情況;HE染色法檢測(cè)裸鼠He La細(xì)胞宮頸癌皮下移植瘤瘤體、肺臟及肝臟組織的病理變化。結(jié)果1病毒(Ad-h IL-28B和Ad-EGFP)滴度均為109PFU/100u L,He La細(xì)胞MOI測(cè)定值為10。2 MTT結(jié)果表明Ad-h IL-28B組明顯較Ad-EGFP組和PBS對(duì)照組吸光度值低,IL-28B對(duì)He La細(xì)胞增殖有明顯抑制作用(F=7.707,P=0.021)。劃痕實(shí)驗(yàn)結(jié)果顯示Ad-h IL-28B組于處理后,生長(zhǎng)緩慢,劃痕與對(duì)照組及Ad-EGFP組比較,明顯增寬,可見(jiàn)IL-28B能顯著抑制He La細(xì)胞的增殖。3成功建立Balb/C裸鼠He La細(xì)胞宮頸癌皮下移植瘤模型。4與PBS組及Ad-EGFP組比,Ad-h IL-28B組裸鼠移植瘤的體積最小(F=437.597,P0.001),表明Ad-h IL-28B能夠明顯減小移植瘤的體積,抑制移植瘤的生長(zhǎng)。5 HE染色結(jié)果表明,Ad-h IL-28B組中腫瘤細(xì)胞的異型性數(shù)量明顯減少,三組裸鼠的肺臟和肝臟組織無(wú)明顯腫瘤轉(zhuǎn)移。6 Western blot結(jié)果表明Ad-h IL-28B顯著提升了促細(xì)胞凋亡的Bax蛋白(F=117.905,P0.001)的表達(dá),卻降低了抗凋亡蛋白Bc L-2蛋白(F=18.398,P=0.001)及STAT3蛋白(F=12.356,P=0.003)的表達(dá)。結(jié)論1 Ad-h IL-28B能夠抑制宮頸癌He La細(xì)胞的增殖。2 Ad-h IL-28B能夠抑制裸鼠He La細(xì)胞宮頸癌皮下移植瘤的生長(zhǎng),抑瘤效果明顯。3 Ad-h IL-28B可能通過(guò)JAK-STAT途徑調(diào)節(jié)STAT3蛋白、Bc L-2、Bax蛋白水平,進(jìn)而影響宮頸癌He La細(xì)胞的增殖及移植瘤的生長(zhǎng)。
[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.
【學(xué)位授予單位】：華北理工大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：R737.33

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