【摘要】：目的本研究前期實驗發(fā)現(xiàn),靶向SIAH2的shRNA載體能顯著抑制HepG2細(xì)胞SIAH2mRNA和蛋白的表達(dá),并抑制細(xì)胞體外增殖。本研究從體內(nèi)水平驗證靶向SIAH2的干擾載體對人肝癌細(xì)胞HepG2細(xì)胞裸鼠移植瘤生長的抑制作用。方法轉(zhuǎn)染pGenesil-SIAH2的HepG2細(xì)胞作為實驗組,命名為HepG2-SIAH2;轉(zhuǎn)染空載體pGenesil-1的HepG2細(xì)胞作為陰性對照組,命名為HepG2-neo;未轉(zhuǎn)染任何質(zhì)粒的HepG2細(xì)胞作為空白對照組,通過G418篩選出穩(wěn)定轉(zhuǎn)染細(xì)胞株。將15只裸鼠隨機(jī)分為3組,接種腫瘤細(xì)胞后觀察裸鼠成瘤情況。4周后測量腫瘤的體積和瘤體質(zhì)量,繪制移植瘤生長曲線,并用實時熒光定量PCR和蛋白質(zhì)印跡法檢測移植瘤中SIAH2的表達(dá)情況。結(jié)果穩(wěn)定轉(zhuǎn)染pGenesil-SIAH2的細(xì)胞株構(gòu)建成功,3組裸鼠接種癌細(xì)胞后均有腫瘤形成。與空白對照組和陰性對照組相比,實驗組腫瘤生長速度明顯減慢,實驗組平均瘤體積(261.57±41.141)mm3,明顯低于陰性對照組(494.35±93.236)mm3(P=0.015)和空白對照組(418.3±28.576 5)mm3(P=0.012),差異有統(tǒng)計學(xué)意義;實驗組平均瘤體質(zhì)量為(0.162±0.02)g,明顯低于陰性對照組(0.358±0.12)g(P=0.032)和空白對照組(0.322±0.24)g(P=0.028)。實驗組瘤體內(nèi)SIAH2mRNA相對表達(dá)量為0.83±0.35,明顯低于陰性對照組2.35±0.96(P=0.003)和空白對照組2.57±0.41(P=0.006),差異有統(tǒng)計學(xué)意義。實驗組瘤體內(nèi)SIAH2蛋白表達(dá)量為0.72±0.02,明顯低于陰性對照組2.61±0.67(P=0.004)和空白對照組2.49±0.91(P=0.007),差異有統(tǒng)計學(xué)意義。結(jié)論靶向SIAH2的shRNA干擾載體能有效抑制人肝癌裸鼠移植瘤的生長,SIAH2有可能成為肝癌基因治療新的分子靶。
[Abstract]:Aim in the previous study, it was found that the shRNA vector targeting SIAH2 could significantly inhibit the expression of SIAH2mRNA and protein in HepG2 cells and inhibit the proliferation of HepG2 cells in vitro. The aim of this study was to investigate the inhibitory effect of the interference vector targeting SIAH2 on the growth of human hepatoma HepG2 cells in nude mice in vivo. Methods HepG2 cells transfected with pGenesil-SIAH2 were used as experimental group, HepG2 cells named as HepG2-SIAH2; transfected empty vector pGenesil-1 as negative control group and HepG2 cells named HepG2-neo; as blank control group. Stable transfection cell lines were screened by G418. Fifteen nude mice were randomly divided into 3 groups. The tumorigenesis of nude mice was observed after inoculation of tumor cells, the tumor volume and tumor mass were measured 4 weeks later, and the growth curve of transplanted tumor was drawn. The expression of SIAH2 in transplanted tumor was detected by real-time fluorescence quantitative PCR and Western blotting. Results the cell lines transfected with pGenesil-SIAH2 stably were successfully constructed and tumor formation was found in all the 3 groups of nude mice inoculated with cancer cells. Compared with the blank control group and the negative control group, the tumor growth rate in the experimental group was significantly slower. The mean tumor volume (261.57 鹵41.141) mm3, in the experimental group was significantly lower than that in the negative control group (494.35 鹵93.236) mm3 (P0. 015) and the blank control group (418.3 鹵28.576 5) mm3 (P0. 012). The mean tumor mass in the experimental group was (0.162 鹵0.02g), which was significantly lower than that in the negative control group (0.358 鹵0.12) g (P0. 032) and the blank control group (0.322 鹵0. 24) g (P0. 028). The relative expression of SIAH2mRNA in the experimental group was 0.83 鹵0.35, which was significantly lower than that in the negative control group (2.35 鹵0.96) and the blank control group (2.57 鹵0.41) (P0. 006). The expression of SIAH2 protein in the experimental group was 0.72 鹵0.02, which was significantly lower than that in the negative control group (2.61 鹵0.67) and the blank control group (2.49 鹵0.91) (P0. 007). Conclusion shRNA interference vector targeting SIAH2 can effectively inhibit the growth of human hepatoma xenografts in nude mice. SIAH2 may be a new molecular target for gene therapy of hepatocellular carcinoma.
【作者單位】： 贛南醫(yī)學(xué)院第一附屬醫(yī)院消化內(nèi)科;贛南醫(yī)學(xué)院第一附屬醫(yī)院醫(yī)務(wù)科;
【基金】：國家自然科學(xué)基金(81160257)
【分類號】：R735.7

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