Gastric Cancer
Copyright ©The Author(s) 2003. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Sep 15, 2003; 9(9): 1935-1939
Published online Sep 15, 2003. doi: 10.3748/wjg.v9.i9.1935
Apoptosis-inducing effect of recombinant Caspase-3 expressed by constructed eukaryotic vector on gastric cancer cell line SGC7901
Yuan-Gen Fu, Yao-Jun Qu, Kai-Chun Wu, Hui-Hong Zhai, Zhi-Guo Liu, Dai-Ming Fan
Yuan-Gen Fu, Department of Biochemistry and Molecular Biology, Medical College, Shantou University, Shantou 515031, Guangdong Province, China
Yao-Jun Qu, Department of Digestive Disease, Longgang Center Hospital, Shenzhen 518116, Guangdong Province, China
Yuan-Gen Fu, Kai-Chun Wu, Zhi-Guo Liu, Dai-Ming Fan, Institute of Digestive Diseases, Fourth Military Medical University, Xi’an 710032, Shaanxi Province, China
Hui-Hong Zhai, Department of Digestive Diseases, The First Hospital, Ningxia Medical College, Yinchuan 759901, Ningxia Hui Autonomous Region, China
Author contributions: All authors contributed equally to the work.
Correspondence to: Dai-Ming Fan, Institute of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, 17 Changlexi Road, Xi’an 710032, Shaanxi Province, China.
Telephone: +86-29-3375229 Fax: +86-29-2539041
Received: September 13, 2002
Revised: September 26, 2002
Accepted: October 29, 2002
Published online: September 15, 2003

AIM: To investigate the apoptosis-inducing effect of Caspases-3 expressed by constructed eukaryotic vector on gastric cancer cell line SGC7901.

METHODS: PCR was employed to amplify the sequences of both small and large subunits of Caspases-3. Its products were separately cloned into the Sma I site of pBluescript KS+ to generate both plasmids pBS/SS and pBS/LS. The small subunit fragment was excised from plasmid pBS/SS with BamH I and then inserted into the BamH I site of plasmid pBS/LS preceding that of the large subunit to yield plasmid pBS/Rev-Caspase-3. Rev-Caspase-3 cDNA was excised with Kpn I + Xba I and then subcloned into plasmid pcDNA3.1 (+) to construct Rev-Caspase-3 eukaryotic expression vector pcDNA/Rev-Caspase-3, which was used to transiently transfect SGC7901 cell line. Cell count, MTT assay and electron microscopy were used to confirm the antiproliferation and apoptosis-inducing effect of Rev-Caspase-3 expression on gastric cancer cells.

RESULTS: Plasmid pBS/Rev-Caspase-3 and eukaryotic expression vector pcDNA/Rev-Caspase-3 were successfully constructed. SGC7901 cells were transiently transfected by either pcDNA/Rev-Caspase-3 or pcDNA3.1 (+) for 24, 48, 72, and 96 h respectively. Cell growth was measured by cell count and MTT assay. In cell count assay, the cell numbers were 1.8 × 106, 1.55 × 106, 2.0 × 106, and 3.1 × 106 in the experimental group and 2.5 × 106, 3.1 × 106, 4.0 × 106, and 5.7 × 106 in the control group at 24, 48, 72 and 96 h respectively. The growth of SGC7901 cells was suppressed by Rev-Caspase-3 in a time-dependent manner (P < 0.05). The results of MTT assay were similar to that of cell count (P < 0.05). The characteristics of apoptosis such as chromatin condensation, crescent formation and margination were seen and more obvious with time in the given-experimental period in the experimental group, but not easily observed in the control group.

CONCLUSION: The expression of Rev-Caspase-3 by the constructed eukaryotic vector can significantly induce apoptosis of gastric cancer cell line SGC7901, which may exhibit a potential way in gastric cancer gene therapy.

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