本文選題：皮下移植瘤 + 雙基因共表達(dá)慢病毒載體�。� 參考：《醫(yī)學(xué)研究生學(xué)報(bào)》2017年06期

【摘要】：目的在轉(zhuǎn)錄后水平進(jìn)行干預(yù)的RNA干擾技術(shù)在多基因治療中的應(yīng)用越來(lái)越廣泛。文中旨在構(gòu)建人HT-29結(jié)腸癌細(xì)胞裸鼠皮下移植瘤模型,通過(guò)聯(lián)合RNAi和外源性補(bǔ)充的方式研究Survivin shRNA-APC雙基因共表達(dá)慢病毒載體對(duì)結(jié)腸癌裸鼠移植瘤生長(zhǎng)情況的影響。方法選取35只裸鼠隨機(jī)數(shù)字表法分為5組,即Survivin shRNA-APC雙基因組、Survivin shRNA組、APC組、空載組和空白組,每組7只;將已經(jīng)構(gòu)建好的處于對(duì)數(shù)生長(zhǎng)期的雙基因共表達(dá)Survivin shRNA-APC、Survivin shRNA及APC穩(wěn)轉(zhuǎn)株,空載穩(wěn)轉(zhuǎn)株,HT-29結(jié)腸癌細(xì)胞(均為2×106/mL)分別注射至5組裸鼠左前腋下成瘤,構(gòu)建人HT-29結(jié)腸癌細(xì)胞裸鼠皮下移植瘤模型;通過(guò)測(cè)量瘤體體積、重量,檢測(cè)移植瘤生長(zhǎng)抑制率,Real time PCR檢測(cè)移植瘤組織survivin mRNA的表達(dá),免疫組化檢測(cè)Survivin蛋白的表達(dá),TUNEL檢測(cè)凋亡情況。結(jié)果與空白組比較,APC組、Survivin shRNA組、雙基因組移植瘤平均體積、平均重量均減少(P0.05);與空載組比較,APC組、Survivin shRNA組、雙基因組平均體積、平均移植瘤重量亦減少(P0.05),而體積抑制率、瘤重抑制率均增加(P0.05);與雙基因組比較,APC組、Survivin shRNA組移植瘤平均體積、平均重量均增加(P0.05)。與空白組和空載組相比,APC組、Survivin shRNA組、Survivin shRNA-APC雙基因組移植瘤組織Survivin mRNA和蛋白表達(dá)相對(duì)含量明顯降低(P0.05);與Survivin shRNA組、APC組相比,雙基因組移植瘤組織Survivin mRNA和蛋白表達(dá)相對(duì)含量亦明顯降低(P0.05)。與空白組裸鼠移植瘤組織結(jié)腸癌細(xì)胞凋亡指數(shù)[(9.89±0.31)%]比較,APC組、Survivin shRNA組、雙基因組[(31.19±1.79)%、(33.64±2.03)%、(56.72±3.17)%]明顯升高(P0.05),而APC組、Survivin shRNA組、雙基因組亦較空載組[(10.06±0.43)%]明顯升高(P0.05);與Survivin shRNA組、APC組相比,Survivin shRNA-APC雙基因組移植瘤組織癌細(xì)胞凋亡指數(shù)明顯升高(P0.05)。結(jié)論 Survivin shRNA-APC雙基因共表達(dá)慢病毒載體能夠使Survivin基因的表達(dá)水平降低,促進(jìn)結(jié)腸癌細(xì)胞的凋亡,抑制移植瘤的生長(zhǎng),且優(yōu)于單個(gè)基因的影響。
[Abstract]:Objective RNA interference at post-transcriptional level has been widely used in multi-gene therapy. The aim of this study was to establish a subcutaneous xenograft model of human HT-29 colon cancer cells in nude mice, and to study the effect of Survivin shRNA-APC double gene co-expression lentivirus vector on the growth of human colon cancer xenografts in nude mice by combining RNAi and exogenous supplementation. Methods Thirty-five nude mice were randomly divided into five groups, namely, Survivin shRNA-APC double-genome survivin shRNA group, no-load group and blank group with 7 rats in each group, and Survivin shRNA-APC-survivin shRNA and APC stable transformants were coexpressed in logarithmic growth phase. Human HT-29 colon cancer cells were subcutaneously transplanted into the left anterior axillary region of 5 groups of nude mice, and the tumor volume and weight were measured by measuring the volume and weight of human colon cancer cell line (HT-29), which was injected into 5 groups of nude mice respectively by injection of the no-load stable transformation strain (2 脳 10 6 / mL) into the subcutaneous xenograft model of human colon cancer cells. Real time PCR was used to detect the expression of survivin mRNA and the expression of Survivin protein was detected by immunohistochemistry. Tunel was used to detect apoptosis. Results compared with the control group, the average volume and average weight of the tumor were reduced by P0.05, and the mean volume and the mean weight of the tumor were also decreased by P0.05 and volume inhibition rate in the shRNA group compared with the control group, and the mean volume and the mean weight of the tumor were also decreased in the shRNA group compared with the control group, and the volume inhibition rate of the tumor was also decreased compared with the control group. The tumor weight inhibition rate was increased by P0.05G, and the mean volume and weight of the transplanted tumor were increased in the APC group and the survivin shRNA group as compared with those in the double genome group (P 0. 05%, P 0. 05%, P 0. 05%). Compared with the control group and the no-load group, the relative content of Survivin mRNA and protein in the survivin shRNA group was significantly lower than that in the Survivin shRNA group, and that in the Survivin shRNA group was significantly lower than that in the Survivin shRNA group. Compared with the control group, the apoptotic index of colon cancer cells in transplanted tumor tissue of nude mice [9.89 鹵0.31%] was significantly higher in APC group than in survivin shRNA group [31.19 鹵1.79%, 33.64 鹵2.03%, 56.72 鹵3.17%], while in APC group, survivin shRNA group was significantly higher than that in APC group, while in the control group, the apoptosis index of colon cancer cells was 9.89 鹵0.31%, while that in APC group was significantly higher than that in survivin shRNA group. Compared with the control group, the apoptotic index of survivin shRNA-APC was significantly higher than that of the control group [10. 06 鹵0. 43%], and the apoptosis index of survivin shRNA-APC tumor cells was significantly higher than that of the Survivin shRNA group (P 0. 05%, P 0. 05%, P 0. 05%, P 0. 05%, P 0. 05%, P 0. 05%). Conclusion Survivin shRNA-APC double gene co-expression lentivirus vector can reduce the expression level of Survivin gene, promote the apoptosis of colon cancer cells, inhibit the growth of transplanted tumor, and is superior to the effect of single gene.
【作者單位】： 佳木斯大學(xué)附屬第一醫(yī)院消化二科;佳木斯大學(xué)臨床醫(yī)學(xué)院;
【基金】：黑龍江省自然科學(xué)基金(H201368) 2015年佳木斯大學(xué)研究生科技創(chuàng)新項(xiàng)目(LZZ2015_018) 2016年佳木斯大學(xué)研究生科技創(chuàng)新項(xiàng)目(YZ2016_025,YM2016_022)
【分類號(hào)】：R735.35

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本文編號(hào)：1901327