[Experimental study of MAT1 gene silencing mediated by siRNA in pancreatic cancer].
OBJECTIVE
To investigate the inhibitory effect of gene silencing mediated by MAT1-siRNA constructed in vitro transcription for pancreatic cancer in vivo and in vitro.


METHODS
21-nt double strand siRNA targeting MAT1 gene was constructed and labeled with Cy3 fluorescent labeling reagent. Human pancreatic cancer cells of the line BxPC3 were cultured and divided into 4 groups: MAT1-siRNA transfected group, negative siRNA control group, lipid control group, and blank control group. The rate of cell duplication was determined by counting the cells for three consecutive days. Cell cycle profiles were determined by flow cytometry. Semi-quantitative analysis of the level of MAT1-mRNA expression was performed using the RT-PCR technique. The level of MAT1 protein expression was analyzed by Western-blotting. 18 nude mice were injected subcutaneously with BxPC3 cells to establish mouse tumor models, and then divided randomly into 3 equal groups: MAT1-siRNA group undergoing injection of MAT1-siRNA directly into the tumors 2 times a week for 4 weeks, blank control group, and negative MAT1-siRNA group. 4 weeks later the mice were killed to observe the weight and size of tumor and to calculate the tumor inhibition rate.


RESULTS
Two of the 4 designed MAT1-siRNAs significantly suppressed the growth of the BxPC3 cells. 72 h after transfection the cell duplication was inhibited by 34.9% in the MAT1-siRNA transfection group. The cell cycle profile showed 83.9% of the MAT1-siRNA transfected cells were in the G0/G1 phase, a rate significantly higher than that in the blank control group (59.86%, P < 0.01). 48 h later the content of MAT1-mRNA of the MAT1-siRNA transfected cells was significantly reduced by 80.12%, and 72 h after the transfection the content of MAT1 protein was reduced by 50.12%, a rate significantly higher than those of the 2 control groups (both P < 0.01). The weight and volume of the transplant tumors in the MAT1-siRNA injected nude mice were significantly reduced compare with the negative siRNA injected control nude mice and blank control nude mice (both P < 0.05). The inhibition rate was 42.53%.


CONCLUSIONS
MAT1 gene silencing mediated by siRNA significantly suppresses the growth of pancreatic cancer cells in vitro, and significantly achieves an anti-tumor effect on the subcutaneously transplanted pancreatic tumor in vivo.
Liu
Jian-ping
JP
Department of Gastroenterology, Guangdong Frontier Armed Police Hospital, Shenzhen 518029, China.
Yuan
Shi-zhen
SZ
Zhang
Shi-neng
SN
D000818:Animals; D002352:Carrier Proteins / Q000235:genetics*; D018797:Cell Cycle Proteins; D045744:Cell Line, Tumor; D020868:Gene Silencing; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009374:Neoplasms, Experimental / Q000235:genetics / Q000473:pathology / Q000628:therapy; D010190:Pancreatic Neoplasms / Q000235:genetics / Q000473:pathology / Q000628:therapy*; D034741:RNA, Small Interfering / Q000235:genetics*; D014157:Transcription Factors; D014162:Transfection; D023041:Xenograft Model Antitumor Assays; D000095043:Cyclin-Dependent Kinase-Activating Kinase
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D034741:RNA, Small Interfering; D014157:Transcription Factors; C514215:MNAT1 protein, human; D000095043:Cyclin-Dependent Kinase-Activating Kinase