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. 2008 Apr 14;14(14):2162-7.
doi: 10.3748/wjg.14.2162.

Effect of NHE1 antisense gene transfection on the biological behavior of SGC-7901 human gastric carcinoma cells

Hai-Feng Liu  1 , Xiao-Chun Teng, Jing-Chen Zheng, Gang Chen, Xing-Wei Wang
Affiliations

Effect of NHE1 antisense gene transfection on the biological behavior of SGC-7901 human gastric carcinoma cells

Hai-Feng Liu et al. World J Gastroenterol. .

Abstract

Aim: To study the effect of type 1 Na(+)/H(+) exchanger (NHE1) antisense human gene transfection on the biological behavior of gastric carcinoma cell line SGC-7901.

Methods: Antisense NHE1 eukaryotic expression on vector pcDNA3.1 was constructed by recombinant DNA technique and transfected into gastric carcinoma cell line SGC-7901 with DOTAP liposome transfection method. Morphological changes of cells were observed with optic and electron microscopes. Changes in cell proliferative capacity, apoptosis, intracellular pH (pH(i)), cell cycle, clone formation in two-layer soft agar, and tumorigenicity in nude mice were examined.

Results: Antisense eukaryotic expressing vectors were successfully constructed and transfected into SGC-7901. The transfectant obtained named 7901-antisense (7901-AS) stablely produced antisense NHE1. There was a significant difference between the pH(i) of 7901-AS cells (6.77 +/- 0.05) and that of 7901-zeo cells and SGC-7901 cells (7.24 +/- 0.03 and 7.26 +/- 0.03, P < 0.01). Compared with SGC-7901 and 7901-zeo cells, 7901-AS cells mostly showed cell proliferation inhibition, G1/G0 phase arrest, increased cell apoptotic rate, recovery of contact inhibition, and density contact. The tumorigenicity in nude mice and cloning efficiency in the two-layer soft agar were clearly inhibited.

Conclusion: NHE1 antisense gene significantly restrains the malignant behavior of human gastric carcinoma cells, suppresses cell growth and induces cell apoptosis, and partially reverses the malignant phenotypes of SGC-7901. These results suggest a potential role for human tumor gene therapy.

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Figures

Figure 1
Figure 1
Identification of antisense human NHE1 cDNA vector digested by restriction endonuclease. Lane 1: Marker (λDNA/EcoRI+ HindIII); Lane 2: pcDNA3.1 (-)/Zeo plasmid; Lane 3: pcDNA3.1-NHE1 plasmid; Lane 4: pcDNA3.1-NHE1 plasmid digested by EcoRI and HindIII (5.0 and 3.6 kb).
Figure 2
Figure 2
Analysis of exogenous gene integration in nucleic DNA of SGC-7901. Lane 1: Polymerase chain reaction marker; Lane 2: 7901-zeo; Lane 3: 7901-AS; Lane 4: SGC-7901.
Figure 3
Figure 3
Standard curve of intracellular pH.
Figure 4
Figure 4
Morphorlogical changes in the transfected cells and parental cells. The first row was observed under inverted microscope. The second row was stained with hematoxylin and eosin, and examined by light microscopy. (A) and (D) SGC-7901; (B) and (E) 7901-zeo; (C) and (F) 7901-AS.
Figure 5
Figure 5
Comparison of growth curves of transfected cells and parental cells.
See this image and copyright information in PMC

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