發(fā)布時間：2018-05-22 07:51

  本文選題：結(jié)直腸癌 + TEAD��； 參考：《中華腫瘤防治雜志》2017年15期

【摘要】：目的結(jié)直腸癌國內(nèi)外高發(fā),是腫瘤死亡的主要病因之一。TEAD4是YAP/TAZ信號通路中的關(guān)鍵因子。本研究旨在探討敲低TEAD4的表達(dá)后對結(jié)直腸癌細(xì)胞增殖的影響和機(jī)制。方法通過在結(jié)直腸癌細(xì)胞系Caco2和HT-29中通過轉(zhuǎn)染TEAD4siRNA和感染慢病毒,以達(dá)到瞬時或穩(wěn)定敲低TEAD4的目的,利用蛋白質(zhì)印跡方法檢測轉(zhuǎn)染效率及相關(guān)蛋白的表達(dá)。用SRB法分別檢測敲低TEAD4后Caco2和HT-29細(xì)胞的存活率,以及使用EdU摻入法測定細(xì)胞DNA合成效率。結(jié)果在腸癌細(xì)胞中使用TEAD4的siRNA,瞬時敲低TEAD4的表達(dá)48h后,蛋白質(zhì)印跡法檢測結(jié)果提示,與對照組相比,Caco2(F=99.22,P0.001)和HT-29(F=167.3,P0.001)細(xì)胞中的TEAD4蛋白水平顯著降低,同時觀察到Caco2(F=45.78,P0.001)和HT-29(F=40.71,P0.001)細(xì)胞中p27蛋白的表達(dá)量有明顯的上升,蛋白表達(dá)差異有統(tǒng)計(jì)學(xué)意義。SRB結(jié)果顯示,使用shRNA慢病毒載體穩(wěn)定敲低腸癌細(xì)胞中TEAD4的表達(dá),與對照組細(xì)胞相比,2d沉默TEAD4基因的Caco2(F=142.9,P0.001)和HT-29(F=141.0,P0.001)細(xì)胞的生長速度變慢;于3d后這種抑制作用更加明顯,Caco2(F=394.5,P0.001)和HT-29(F=305.1,P0.001)細(xì)胞增殖速度明顯變慢。使用EdU摻入法檢測敲低TEAD4后對Caco2和HT-29細(xì)胞DNA合成的影響,結(jié)果顯示,Caco2和HT-29細(xì)胞,Caco2/Consi、Caco2/TEAD4si#2、Caco2/TEAD4si#3、HT-29/Consi、HT-29/TEAD4si#2、HT-29/TEAD4si#3的DNA合成率分別為(43.20±2.18)%、(27.19±2.34)%和(30.14±1.02)%及(28.23±3.11)%、(11.89±1.20)%和(17.91±2.23)%,干擾組Caco2/TEAD4si#2(P=0.001)、Caco2/TEAD4si#3(P0.001)、HT-29/TEAD4si#2(P=0.001)及HT-29/TEAD4si#3(P=0.011)細(xì)胞的DNA合成率顯著低于對照組細(xì)胞,說明在Caco2和HT-29細(xì)胞中敲低TEAD4后,DNA的合成明顯受到了抑制。結(jié)論敲低TEAD4對結(jié)直腸癌細(xì)胞體外增殖和DNA的合成有抑制作用,可能部分通過上調(diào)p27的表達(dá),TEAD4可能是結(jié)直腸癌發(fā)生、發(fā)展中潛在的臨床診斷或治療新靶點(diǎn)。
[Abstract]:Objective Colorectal cancer is a major cause of death at home and abroad. TEAD4 is a key factor in YAP/TAZ signaling pathway. The aim of this study was to investigate the effect and mechanism of knockout TEAD4 expression on the proliferation of colorectal cancer cells. Methods transfection of TEAD4siRNA and lentivirus infection in colorectal cancer cell lines Caco2 and HT-29 were carried out to achieve transient or stable knockdown of TEAD4. Western blotting was used to detect transfection efficiency and expression of related proteins. The survival rate of Caco2 and HT-29 cells after TEAD4 knockout was detected by SRB assay, and the DNA synthesis efficiency was measured by EdU incorporation method. Results TEAD4 siRNAs were used in intestinal cancer cells, and the expression of TEAD4 was instantly knocked down for 48 hours. The results of Western blot analysis showed that the TEAD4 protein levels in the cells were significantly lower than those in the control group (P 0.001) and HT-29FN 167.3P 0.001) cells. It was also observed that the expression of p27 protein increased significantly in Caco2FN 45.78 (P0.001) and HT-29 FN 40.71 P0.001) cells, and the difference in protein expression was statistically significant. The results showed that the expression of p27 protein was stabilized by shRNA lentivirus vector, and the expression of p27 protein in low intestinal cancer cells was stabilized by shRNA lentivirus vector. Compared with the control cells, the growth rate of the cells silencing TEAD4 gene for 2 days was slower than that of the control cells, and the growth rate of the cells was much slower than that of the control cells at 3 days after silencing the TEAD4 gene, and the proliferation rate of the cells was significantly slower than that of the control cells, and the proliferation rate of the cells was significantly slower than that of the control cells. The proliferation rate of the cells was significantly slower than that of the control cells. 浣跨敤EdU鎺哄叆娉曟嫻嬫暡浣嶵EAD4鍚庡Caco2鍜孒T-29緇嗚優(yōu)DNA鍚堟垚鐨勫獎鍝，

本文編號：1921181