[Effects of Smac gene overexpression on chemotherapeutic sensitivity of gastric cancer cell line MKN-45].
OBJECTIVE
Abnormal apoptosis is one of the key factors for drug resistance of neoplasms. Smac, a novel gene involved in regulation of apoptosis,plays an important role in tumor cell apoptosis induced by chemotherapeutic drugs. This study was designed to explore the effects of overexpressed Smac gene on chemotherapeutic sensitivity of gastric cancer cell line.


METHODS
By liposome GeneSHUTTLE-40, Smac gene was transfected into gastric cancer cell line MKN-45. Cellular Smac gene expression were determined by reverse transcription-polymerase chain reaction (RT-PCR)and Western blot analysis. Cisplatin (1, 5, 10 microg/ml), mitomycin C (0.1,1,10 microg/ml), and curcumin (10, 20, 40 micromol/L) were selected to treat untransfected and transfected MKN-45 cells. Cellular proliferation activities were assayed by tetrazolium bromide (MTT) colorimetry. The morphological changes of cancer cells were observed under inverted microscope. Cellular apoptosis was determined by Annexin V-FITC and propidium iodide staining flow cytometry.


RESULTS
Compared with untransfected control group,Smac mRNA and protein levels in transfected MKN-45 cells were significantly increased (P< 0.01). The growth inhibition rates of MKN-45 cells treated with various concentration of cisplatin, mitomycin C, and curcumin on MKN-45 cells were increased by 10.10%-23.80% (P< 0.01), 10.01%-15.86% (P< 0.01), 11.28%-22.12% (P< 0.01), respectively, with the cells becoming round, obviously refracted, and fragmented. The cellular apoptosis rates were increased by 6.7%-20.2% (P< 0.01), 5.4%-13.2% (P< 0.01), and 10.6%-20.1% (P< 0.01), respectively.


CONCLUSIONS
Transfection of extrinsic Smac gene results in its over-expression in MKN-45 cells,which could increase the chemotherapeutic sensitivity of MKN-45 cells.
Zheng
Li-Duan
LD
Department of Pathology, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, 430022, PR China. ld_zheng@hotmail.com
Tong
Qiang-Song
QS
Tao
Kai-Xiong
KX
Wang
Liang
L
Zhang
Bo
B
D017209:Apoptosis / Q000187:drug effects; D051017:Apoptosis Regulatory Proteins; D015153:Blotting, Western; D002352:Carrier Proteins / Q000032:analysis / Q000235:genetics*; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D003474:Curcumin / Q000494:pharmacology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins / Q000032:analysis / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D013274:Stomach Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology
D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; C412403:DIABLO protein, human; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins; D012333:RNA, Messenger; D003474:Curcumin