Gastric Carcinoma Cell Line SGC-7901 Research Articles

Construction and identification of recombinant vectors carrying herpes simplex virus thymidine kinase and cytokine genes expressed in gastric carcinoma cell line SGC7901.

To construct and identify the recombinant vectors carrying herpes simplex virus thymidine kinase (HSV-TK) and tumor necrosis factor alpha (TNF-alpha) or interleukin-2 (IL-2) genes expressed in gastric carcinoma cell line SGC7901. The fragments of HSV-TK, internal ribosome entry sites (IRES) and TNF-alpha or IL-2 genes were inserted in a TK-IRES-TNF-alpha or TK-IRES-IL-2 order into pEGFP-N(3) and pLXSN to generate the therapeutic vectors pEGFP-TT, pEGFP-TI, pL(TT)SN and pL(TI)SN respectively, which were structurally confirmed by the digestion analysis of restriction endonuclease. The former two plasmids were used for the transient expression of recombinant proteins in the target cells while pL(TT)SN and pL(TI)SN were transfected into SGC7901 cells by lipofectamine for the stable expression of objective genes through G418 selection. The protein products expressed transiently and stably in SGC7901 cells by the constructed vectors were confirmed by fluorescent microscopy and Western blot respectively. The inserted fragments in all constructed plasmids were structurally confirmed to be consistent with that of the published data. In the transient expression, both pEGFP-TT and pEGFP-TI were shown expressed in nearly 50% of the transfected SGC7901 cells. Similarly, the G418 selected vectors PL(TT)SN and PL(TI)SN were confirmed to be successful in the stable expression of the objective proteins in the target cells. The constructed recombinant vectors in the present study that can express the suicide gene TK in combination with cytokines genes may serve as the potential tools to perform more effective investigations in future for the gene therapy of gastric carcinoma.

Human Gastric Tissues Simultaneously Express the Classical and Alternative Splicing Cholecystokinin-B/Gastrin Receptors

To explore whether cholecystokinin-B/gastrin receptor (CCKBRwt) gene and its alternative splicing variant preserving intron 4 (CCKBRi4sv) are expressed in human gastric carcinoma cell line and tissue, we detect mRNA expression of CCKBRwt and CCKBRi4sv in 30 gastric carcinoma and their corresponding normal tissues, 10 gastritis, and 2 autopsied normal stomach specimens as well as in a gastric carcinoma cell line SGC-7901 cells by RT-PCR and sequencing. The results revealed that human normal, inflammatory, and malignant gastric tissues simultaneously expressed the classical and alternative splicing cholecystokinin-B/gastrin receptor genes. The alternative splicing variant contains the intron 4 of cholecystokinin-B/gastrin receptor gene.

Effects of cyclin D1 antisense oligodeoxyneucleotides on the growth and expression of G1 phase regulators in gastric carcinoma cells.

To investigate the effects of Cyclin D1 antisense oligodeoxyneucleotides (ASODN) on the growth, cell cycle progression and expression of G1 phase regulators in human gastric carcinoma cell lines SGC7901 and HS746T, phosphorothioate-modified Cyclin D1 ASODN were encapsulated by LipofectAMINE2000 and transfected into gastric carcinoma cells. Dose-dependent inhibitory effects were induced by Cyclin D1 ASODN in two gastric carcinoma cell lines. Treatment of gastric carcinoma cells with 0.2 micromol/L Cyclin D1 ASODN for 24 h could significantly inhibit their growth in vitro and in vivo, reduce expression of Cyclin D1 mRNA to 26.3% (SGC7901) and 17.3% (HS746T) respectively. The percentage of cells in G0/G1 phase was increased as revealed by flow cytometry. Immunohistochemical staining showed that the expression of p21 was increased and the expression of Cyclin D1 and pRb was decreased in the two cell lines; the expression of p27 was increased in HS746T, but unchanged in SGC7901. Cyclin D1 ASODN could inhibit the growth and the expression of Cyclin D1 mRNA in gastric carcinoma cells, influence the cell cycle and expression of its regulators.

Inhibitory effects of c9, t11-conjugated linoleic acid on invasion of human gastric carcinoma cell line SGC-7901.

To investigate the effect of c9, t11-conjugated linoleic acid (c9, t11-CLA) on the invasion of human gastric carcinoma cell line and its possible mechanism of preventing metastasis. Using reconstituted basement membrane invasion, chemotaxis, adhesion, PAGE substrate zymography and RT-PCR assays, we analyzed the abilities of invasion, direct migration, adhesion of intracellular matrix, as well as the activity of type IV collagenase and expression of tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2 mRNA in SGC-7901 cells which were treated with gradually increased concentrations (25, 50, 100 and 200 micromol/L) of c9, t11-CLA for 24 h. At the concentrations of 200 micromol/L, 100 micromol/L and 50 micromol/L, c9,t11-CLA suppressed the invasion of SGC-7901 cells into the reconstituted basement membrane by 53.7 %, 40.9 % and 29.3 %, respectively, in comparison with the negative control. Only in the 200 micromol/L c9,t11-CLA group, the chemotaxis of SGC-7901 cells was inhibited by 16.0 % in comparison with the negative control. C9,t11-CLA also could inhibit the adhesion of SGC-7901 cells to laminin, fibronectin and Matrigel, increase the expression of TIMP-1 and TIMP-2 mRNA, and reduce type IV collagenase activities in the serum-free medium supernatant of SGC-7901 cells. c9,t11-CLA can inhibit the invasion of SGC-7901 cells at multiple procedures in tumor metastasis cascade, which may be associated with the induction of TIMP-1 and TIMP-2 mRNA expression.

Effect of antisense human telomerase RNA on malignant phenotypes of gastric carcinoma.

The study was designed to explore the effects of antisense human telomerase RNA (ahTR) on the malignant phenotype of gastric carcinoma cell line SGC-7901, and its potential role in gene therapy for tumors. An ahTR eukaryotic expression vector, including the sequence of template region of telomere repeats, was constructed by recombinant technology of molecules and then transfected into gastric carcinoma cell line SGC-7901 by liposome DOTAP. Subsequently, the expression of hTR RNA and ahTR RNA by reverse transcription-polymerase chain reaction, telomerase activity by telomeric repeat amplification protocol-ELISA (TRAP-ELISA), telomere length by Southern blotting, cell morphology under light microscope, cellular proliferation capacity by 3-(4,5-dimethyl-2 thiazoyl)-2,5-diphenyl-2H-tetrazolium bromide assay, cell-cycle distribution by flow cytometry, efficiency of clone formation in soft agar, and tumorigenecity in nude mice were examined and evaluated in ahTR-transfected cells, control plasmid pCI-neo transfected cells and their parental cells. An ahTR eukaryotic expression vector was constructed and successfully transfected into SGC-7901 cells. The telomerase activity in ahTR-transfected SGC-7901 cells decreased from 100% to approximately 25%, and telomere length in the cells shortened to 3.35 from 4.08 Kb at 60 population doublings. Compared with the parental cells and pCI-neo transfected cells, ahTR-transfected cells displayed some morphological changes, such as decreased atypia, and recovery of contact inhibition and density inhibition under light microscope. Furthermore, ahTR-transfected cells displayed decreased invasive capacity in Borden's chamber invasive model, increased G0/G1 phase rate and apoptotic rate. Surprisingly, ahTR-transfected SGC-7901 cells lost their capacity for clone formation in soft agar and tumorigencity in nude mice. Antisense-hTR transfection can inhibit the growth of SGC-7901 cells and partially reverse the malignant phenotypes. This study provides an exciting approach for cancer therapy by inhibiting telomerase activity using an antisense gene.

The therapeutic effects of recombinant adenovirus RA538 on human gastric carcinoma cells in vitro and in vivo.

AIM:To evaluate the potential of RA-538 gene therapy for gastric carcinoma.METHODS:Human gastric carcinoma cell line SGC7901 treated with Ad-RA538 or Ad-LacZ were analysed by X-gal stain, MTT, DNA ladder, Tunel, flow cytometric analysis, PCR, and Western Blot in vitro. The tumorigenicity and experimental therapy in nude mice model were assessed in vivo.RESULTS:Ad-LacZ could efficiently transfer the LacZ gene into SGC7901 cells. X-gal-positive cells at MOI 25, 50, 100, and 200 were 90%, 100%, 100%, and 100% respectively. Ad-RA538 could strongly inhibit cell growth and induced apoptosis in SGC7901 cells.The proliferation of the Ad-RA538-infected SGC7901 cells was reduced by 76.3%.The mechanism of killing of gastric carcinoma cells by Ad-RA538 was found to be apoptosis by DNA ladder,Tunel and flow cytometric analysis.The tumorigenicity in nude mice using Ad-RA538 showed that all three mice failed to form tumor from 7 to 30 days compared with Ad-LacZ and parent SGC7901 cells. Experimental therapy on the nude mice model bearing subcutaneous tumor of SGC7901 cells showed that intratumor instillation of Ad-RA538 inhibited the growth of the tumors. Ad-RA538-treated tumors were inhibited by 60.66%, compared with that of the tumor injected with Ad-LacZ and mock.CONCLUSION: The expression of Ad RA538 can inhibit growth and induce apoptosis of gastric cancer cell in vitro and in vivo. Ad RA538 can be used potentially in gene therapy for gastric carcinoma.