中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2010年
1期
47-49
,共3页
乳腺癌%基因%逆转录病毒%脱噬作用
乳腺癌%基因%逆轉錄病毒%脫噬作用
유선암%기인%역전록병독%탈서작용
Breast carcinoma%Gene%Retmvims%Apoptosis
目的 构建表达重组凋亡素(VP3)基因的重组逆转录病毒,探讨其对人乳腺癌MCF-7裸鼠移植瘤模型的凋亡诱导活性及机制.方法 克隆VP3基因,重组法构成逆转录病毒载体pLP-LNCX-VP3(pLLVP3),转染PT67细胞进行病毒包装;雌激素皮贴刺激法建立去卵巢裸鼠乳腺癌皮下移植瘤模型,观察pLLVP3对肿瘤的生长抑制情况,TUNEL法检测肿瘤凋亡,Western blot印迹法检测VP3、Caspase-3和bcl-2蛋白表达.结果 重组载体pLLVP3鉴定正确,滴度为1×10~7 pfu/ml;裸鼠实验pLLVP3组抑瘤率分别为65.52%和68.23%,显著高于对照组(t=4.06,P<0.01),48 h可见VP3蛋白高表达.TUNEL见各组凋亡指数无差异(t_1=1.05,t_2=0.84,均为P>0.05).VP3蛋白高表达组Caspase-3表达亦升高,但bcl-2未见差异.结论 VP3基因能够诱导人乳腺癌细胞MCF-7的凋亡,可能是激活Caspase-3而发生作用.
目的 構建錶達重組凋亡素(VP3)基因的重組逆轉錄病毒,探討其對人乳腺癌MCF-7裸鼠移植瘤模型的凋亡誘導活性及機製.方法 剋隆VP3基因,重組法構成逆轉錄病毒載體pLP-LNCX-VP3(pLLVP3),轉染PT67細胞進行病毒包裝;雌激素皮貼刺激法建立去卵巢裸鼠乳腺癌皮下移植瘤模型,觀察pLLVP3對腫瘤的生長抑製情況,TUNEL法檢測腫瘤凋亡,Western blot印跡法檢測VP3、Caspase-3和bcl-2蛋白錶達.結果 重組載體pLLVP3鑒定正確,滴度為1×10~7 pfu/ml;裸鼠實驗pLLVP3組抑瘤率分彆為65.52%和68.23%,顯著高于對照組(t=4.06,P<0.01),48 h可見VP3蛋白高錶達.TUNEL見各組凋亡指數無差異(t_1=1.05,t_2=0.84,均為P>0.05).VP3蛋白高錶達組Caspase-3錶達亦升高,但bcl-2未見差異.結論 VP3基因能夠誘導人乳腺癌細胞MCF-7的凋亡,可能是激活Caspase-3而髮生作用.
목적 구건표체중조조망소(VP3)기인적중조역전록병독,탐토기대인유선암MCF-7라서이식류모형적조망유도활성급궤제.방법 극륭VP3기인,중조법구성역전록병독재체pLP-LNCX-VP3(pLLVP3),전염PT67세포진행병독포장;자격소피첩자격법건립거란소라서유선암피하이식류모형,관찰pLLVP3대종류적생장억제정황,TUNEL법검측종류조망,Western blot인적법검측VP3、Caspase-3화bcl-2단백표체.결과 중조재체pLLVP3감정정학,적도위1×10~7 pfu/ml;라서실험pLLVP3조억류솔분별위65.52%화68.23%,현저고우대조조(t=4.06,P<0.01),48 h가견VP3단백고표체.TUNEL견각조조망지수무차이(t_1=1.05,t_2=0.84,균위P>0.05).VP3단백고표체조Caspase-3표체역승고,단bcl-2미견차이.결론 VP3기인능구유도인유선암세포MCF-7적조망,가능시격활Caspase-3이발생작용.
Objective By constructing recombinant expression plasmid pLLYP3 expressing recombinant Apoptin (VP3) gene,to discuss the effect of VP3 inducing tumor cells into apoptosis on human breast cancer cell MCF-7 orthotopic transplantation tumors and the mechanism.Methods VP3 gene was cloned into the plasmid pLP-LNCX to form the recombinant plasmid pLLVP3,and MCF-7 orthotopic transplantation tumor model Was established,then pLLVP3 was infected into models.The expression of VP3 gene was detected by Western blot.TdT-mediated duTp nick end labeling (TUNEL) assay was used to verify apoptosis of tnnlor cells.At last Caspase-3 and B cell lymphoma-2 (bcl-2) were detected by Western blot.Results Sequence analysis revealed the recombination of plasmid pLLVP3,and 1×10~7 pfu/ml virus was obtained.The findings of animal experiment showed that tulnor inhibition rate of pLVP3 with low and high doses was 65.52% and 68.23% respectively,evidently higher than that in control group(t=4.06,P<0.01).Besides,Apoptin protein Was over-expressed.Forty-eight h after infection,TUNEL analysis showed obvious apoptosis peaks with the highest percentage rate of apoptotic cells present and cellular Caspase-3 expression could be detected in pLLVP3-infected group.Conclusion VP3 gene could induce apoptosis in human colon cancer cell line MCF-7 in vivo by activating Caspase-3 expression.
