NT4p53(C22)Ant重组腺伴病毒对人Miapaca-Ⅱ胰腺癌细胞的杀伤作用

Killing effects of recombinant AAV expressing fusion peptide NT4p53 (C22) Ant on human pancreatic cancer(Miapaca)-Ⅱcells

目的:研究NT4p53(C22)Ant重组腺伴病毒对人M iapaca-Ⅱ胰腺癌细胞杀伤作用.方法:构建NT4p53(C22)Ant,将其亚克隆于腺伴病毒载体中,经293细胞包装、扩增构成具有感染性的重组腺伴病毒AAV-NT4p53(C22)Ant,转染人胰腺癌M iapaca-Ⅱ细胞,经光镜下观察、MTT比色试验、PI染色实验及乳酸脱氢酶释放检测(LDH),观察AAV-NT4p53(C22)Ant对细胞的杀伤作用.结果:光镜下可见随AAV-NT4p53(C22)Ant感染Miapaca-Ⅱ细胞时间延长,肿瘤细胞数目明显减少;MTT实验检测发现细胞存活率明显降低(P<0.05);流式细胞检测凋亡细胞数目较对照组及空病毒组增加(P<0.05);LDH检测培养基中LDH只有轻度增加(P>0.05).结论:重组腺伴病毒NT4p53(C22)Ant对胰腺癌细胞有杀伤作用,并且是通过引起细胞凋亡而实现的.

AIM： To study the killing effects of recombinant adeno-associated virus （AAV） expressing fusion peptide NT4p53 （C22） Ant on human pancreatic cancer Miapaca-Ⅱ cells. METHODS： NT4p53 （C22）Ant was constructed and subcloned into the vector of recombinant AAV, and that was amplified in 293 packaging cells. The recombinant plasmid AAV-NT4p53 （C22） Ant was transferred into Miapaca-Ⅱ cells. The direct effects on human pancreatic cancer cell line Miapaca- Ⅱ were detected with light microscope, MTT, flow cytometry and lactate dehydrogenase（ LDH ） release assay. RESULTS： It could be observed under microscope that the AAV-NT4p53 （C22）Ant could directly kill the Miapaca- Ⅱ cells. MTT revealed that the survival rate of the cells was decreased obviously. Flow cytometry showed that the number of apoptotic cells was significantly increased in AAV- NT4p53 （ C22 ） Ant group as compared with control group and simple AAV group （ P 〈 0.05 ）. LDH level went up a little （ P 〉 0. 05 ）. CONCLUSION： The AAV-NT4p53 （ C22 ） Ant could cause the apoptosis of human pancreatic cancer Miapaca- Ⅱ cells to kill the cells directly.