中华传染病杂志
中華傳染病雜誌
중화전염병잡지
CHINESE JOURNAL OF INFECTIOUS DISEASES
2009年
7期
398-402
,共5页
李涛%刘国珍%谭德明%吴传湘%郑芳
李濤%劉國珍%譚德明%吳傳湘%鄭芳
리도%류국진%담덕명%오전상%정방
肝炎病毒,乙型%基因转变%肝细胞%质粒
肝炎病毒,乙型%基因轉變%肝細胞%質粒
간염병독,을형%기인전변%간세포%질립
Hepatitis B virus%Gene conversion%Hepatocytes%Plasmids
目的 观察HBV X基因多功能蛋白(HBx)的表达对QSG7701细胞生物学特征的影响及对其的转化作用.方法 采用脂质体法分别将重组质粒pCMV/X及真核表达质粒pRe/CMVZ稳定转染QSG7701细胞,分别获得pCMV X/QSG7701、pReCMV2/QSG7701,设未转染的QSG7701细胞为对照组.Western印迹检测细胞中HBx、c-Myc及Bel-2蛋白的表达,MTT比色法、流式细胞技术、软琼脂克隆形成率实验检测细胞的生物学活性.结果HBx高表达于pCMV X/QSG7701中.pCMV X/QSG7701中c-Mye蛋白的表达水平较其他两组高,Bcl-2蛋白在3组细胞中均有表达,但表达水平差异无统计学意义.在pCMV X/QSG7701、pRcCMV2/QSG7701及未转染的QSG7701 3组细胞中,pCMV X/QSG7701组s期细胞所占百分比较后两组明显增加[分别为(28.80±2.32)%、(15.50+2.64)%、(21.50±3.66)%,LSD 0.05=3.95%,LSD 0.01=5.47%,P<0.01],其G1期细胞所占百分比较后两组明显减少[分别为(62.30±3.85)%、(78.70±4.12)%、(78.105=4.45)%,LSD 0.05=5.63%,LSD 0.01-7.79%,P<0.01],其凋亡率明显高于后两组[分别为(14.90+11 01)%、(8.91±0.48)%、(4.03±0.47)%,LSD 0.05=0.94%,LSD 0.01=1.31%,P<0.01],其细胞株的倍增时间较后两组明显缩短(分别为14 h,29 h,38 h),其软琼脂克隆形成率明显高于后两组[分别为(19.83±1.96)%,(1.764±0.03)%,(1.33±0.18)%,LSD 0.05=1.53%,LSD 0.01=2.11%,P<0.01].结论 HBx能够在体外转化人源性非瘤性肝细胞QSG7701,使细胞具有恶性化生长的倾向.
目的 觀察HBV X基因多功能蛋白(HBx)的錶達對QSG7701細胞生物學特徵的影響及對其的轉化作用.方法 採用脂質體法分彆將重組質粒pCMV/X及真覈錶達質粒pRe/CMVZ穩定轉染QSG7701細胞,分彆穫得pCMV X/QSG7701、pReCMV2/QSG7701,設未轉染的QSG7701細胞為對照組.Western印跡檢測細胞中HBx、c-Myc及Bel-2蛋白的錶達,MTT比色法、流式細胞技術、軟瓊脂剋隆形成率實驗檢測細胞的生物學活性.結果HBx高錶達于pCMV X/QSG7701中.pCMV X/QSG7701中c-Mye蛋白的錶達水平較其他兩組高,Bcl-2蛋白在3組細胞中均有錶達,但錶達水平差異無統計學意義.在pCMV X/QSG7701、pRcCMV2/QSG7701及未轉染的QSG7701 3組細胞中,pCMV X/QSG7701組s期細胞所佔百分比較後兩組明顯增加[分彆為(28.80±2.32)%、(15.50+2.64)%、(21.50±3.66)%,LSD 0.05=3.95%,LSD 0.01=5.47%,P<0.01],其G1期細胞所佔百分比較後兩組明顯減少[分彆為(62.30±3.85)%、(78.70±4.12)%、(78.105=4.45)%,LSD 0.05=5.63%,LSD 0.01-7.79%,P<0.01],其凋亡率明顯高于後兩組[分彆為(14.90+11 01)%、(8.91±0.48)%、(4.03±0.47)%,LSD 0.05=0.94%,LSD 0.01=1.31%,P<0.01],其細胞株的倍增時間較後兩組明顯縮短(分彆為14 h,29 h,38 h),其軟瓊脂剋隆形成率明顯高于後兩組[分彆為(19.83±1.96)%,(1.764±0.03)%,(1.33±0.18)%,LSD 0.05=1.53%,LSD 0.01=2.11%,P<0.01].結論 HBx能夠在體外轉化人源性非瘤性肝細胞QSG7701,使細胞具有噁性化生長的傾嚮.
목적 관찰HBV X기인다공능단백(HBx)적표체대QSG7701세포생물학특정적영향급대기적전화작용.방법 채용지질체법분별장중조질립pCMV/X급진핵표체질립pRe/CMVZ은정전염QSG7701세포,분별획득pCMV X/QSG7701、pReCMV2/QSG7701,설미전염적QSG7701세포위대조조.Western인적검측세포중HBx、c-Myc급Bel-2단백적표체,MTT비색법、류식세포기술、연경지극륭형성솔실험검측세포적생물학활성.결과HBx고표체우pCMV X/QSG7701중.pCMV X/QSG7701중c-Mye단백적표체수평교기타량조고,Bcl-2단백재3조세포중균유표체,단표체수평차이무통계학의의.재pCMV X/QSG7701、pRcCMV2/QSG7701급미전염적QSG7701 3조세포중,pCMV X/QSG7701조s기세포소점백분비교후량조명현증가[분별위(28.80±2.32)%、(15.50+2.64)%、(21.50±3.66)%,LSD 0.05=3.95%,LSD 0.01=5.47%,P<0.01],기G1기세포소점백분비교후량조명현감소[분별위(62.30±3.85)%、(78.70±4.12)%、(78.105=4.45)%,LSD 0.05=5.63%,LSD 0.01-7.79%,P<0.01],기조망솔명현고우후량조[분별위(14.90+11 01)%、(8.91±0.48)%、(4.03±0.47)%,LSD 0.05=0.94%,LSD 0.01=1.31%,P<0.01],기세포주적배증시간교후량조명현축단(분별위14 h,29 h,38 h),기연경지극륭형성솔명현고우후량조[분별위(19.83±1.96)%,(1.764±0.03)%,(1.33±0.18)%,LSD 0.05=1.53%,LSD 0.01=2.11%,P<0.01].결론 HBx능구재체외전화인원성비류성간세포QSG7701,사세포구유악성화생장적경향.
Objective To observe the effects of hepatitis B virus(HBV)X gene muhifunctional protein(HBx)on the biological characteristics of QSG7701 and the transformational effects on QSG7701 cell.Methods QSG7701 ceils were stably transformed by recombinant plasmid pCMV/X and eukaryotic expressed plasmid pRc/CMV2 by liposome-based assay,respectively.Non-transfeeted QSG7701 cells were employed as controls.The expressions of HBx,c-Myc and Bel-2 proteins in QSG7701 cells were detected by Western blot.MTT colorimetric analysis,flow cytometry and soft agar clone-forming assay were performed tO detect the biolo~dcal activity of cells.Results HBx Drotein was highly expressed in pCMV X/QSG7701 cells.The expression level of c-Myc protein in the pCMV X/QSG7701 cells was much higher than those in the other tWO groups of cells.The expression of Bcl2 protein was detected in the three groups of cells,but the expression levels were similar.Percentage of S stage cells in pCMV X/QSG7701 ceils was significantly higher than those in pRcCMV2/QSG7701 and non-transfected QSG7701 cells E(28.80±2.32)%,(15.5±2.64)%and(21.5±3.66)%,LSD 0.05=3.95%,LSD 0.01=5.47%,P<0.01].While percentage of G1 stage cells in pCMV X/QSG7701 cells was significantly lower than those in pRcCMV2/QSG7701 and non-transfected QSG7701 cells[(62.30±3.85)%,(78.70±4.12)%and(78.10q±4.45)%,LSD 0.05=5.63%,LSD 0.01-7.79 %,P<0.01].The apoptosis rate of pCMV X/QSG7701 cells was much higher than those in pRcCMV2/QSG770t and non-transfected QSG7701 cells[(14.90±1.01)%,(8.91±0.48)%and (4.03±0.47)%,LSD 0.05=O.94%,LSD 0.01=1.31%,P<0.01].The population doubling time of pCMV X/QSG7701 cells was shorter than those in pRcCMV2/QSG and non-trarmfected cells(14 h,29 h,38 h,respectively).The cloning ratio in soft agar of pCMV X/QSG7701 cells WaS significantly higher than those of pRcCMV2/QSG and non-transfected QSG7701 cells[(19.83±1.96)%,(1.76±0.03)%and (1.33±0.18)%,LSD 0.05=1.53%,LSD 0.01=2.11%,P<0.01].Conclusion HBx may transform human non-tumor hepatoeyte QSG7701,which makes the cell growth malignizeA.
