Inducing apoptosis of human nasopharyngeal carcinoma cell line CNE-2Z by bcl-xL short hairpin RNA.
OBJECTIVE
Recent studies showed overexpression of bcl-x(L) in human nasopharyngeal carcinoma (NPC) cell line CNE-2Z; it may play a pivotal role in tumorigenesis, metastasis, and drug resistance of NPC. This study was to explore inducing effect of bcl-x(L) short hairpin RNA (shRNA) on apoptosis of CNE-2Z cells.


METHODS
After transfection of recombinant plasmid pmU6-RNAi expressing bcl-x(L) shRNA, apoptotic CNE-2Z cells were detected by fluorescent staining and flow cytometry (FCM). mRNA levels of bcl-x(L), bcl-2, survivin, and caspase-3 was detected by reverse transcription-polymerase chain reaction (RT-PCR); while protein levels of Bcl-x(L), Caspase-3, and P53 were detected by Western blot.


RESULTS
When treated with pmU6-RNAi for 24 h, an obvious apoptotic peak of CNE-2Z cells appeared; cell shrinkage, chromatin condensation, and nuclear fragmentation were observed in most cells under fluorescent microscope. RT-PCR analysis showed that pmU6-RNAi down-regulated mRNA levels of bcl-x(L), bcl-2, and caspase-3, but had little or no effect on mRNA level of survivin; Western blot analysis showed an obvious reduction in protein levels of Bcl-x(L) and Caspase-3, and a great increase in protein level of P53.


CONCLUSIONS
bcl-x(L) shRNA can induce apoptosis of CNE-2Z cells, which may be closely related to down-regulation of bcl-2, caspase-3 and p53. bcl-x(L) shRNA may be helpful for developing gene therapy for NPC.
He
Cheng-Wei
CW
Institute of Biochemistry and Molecular Biology, Guangdong Medical College, Zhanjiang, Guangdong 524023, P. R. China.
Liu
Fang
F
Zhang
Yue-Fei
YF
Liang
Tong
T
Zhou
Ke-Yuan
KY
D017209:Apoptosis; D053148:Caspase 3 / Q000096:biosynthesis / Q000235:genetics; D045744:Cell Line, Tumor; D015316:Genetic Therapy; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins / Q000096:biosynthesis / Q000235:genetics; D009303:Nasopharyngeal Neoplasms / Q000378:metabolism / Q000473:pathology*; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000235:genetics; D010957:Plasmids; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis / Q000235:genetics; D034622:RNA Interference; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D000077022:Survivin; D014162:Transfection; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D051020:bcl-X Protein / Q000096:biosynthesis / Q000235:genetics*
C107489:BIRC5 protein, human; D051034:Inhibitor of Apoptosis Proteins; D008869:Microtubule-Associated Proteins; D009363:Neoplasm Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011994:Recombinant Proteins; D000077022:Survivin; D016159:Tumor Suppressor Protein p53; D051020:bcl-X Protein; D053148:Caspase 3