Abstract
We investigated whether endothelial nitrite oxide synthase (NOS) gene transfer inhibited cellular proliferation. Endothelial NOS and endothelin type A receptor genes were transferred into 293 cells, a human embryonic kidney cell line, by calcium-phosphate coprecipitation. The cytosolic free Ca2+ levels ([Ca2+]i) of transfected cells were estimated with fura-2 fluorescence. Thymidine incorporation was increased by endothelin-1 in type A receptor-transfected cells. The endothelial NOS gene transfer did not affect endothelin-1-induced increase in [Ca2+]i of type A receptor-transfected cells, but markedly inhibited mitogen-activated protein kinase andc-fos promoter activities. The endothelial NOS gene transfer also inhibited thymidine incorporation into type A receptor-transfected cells in response to endothelin-1, which was abolished in the presence of the NOS inhibitorNG-monomethyl-l-arginine acetate. The endothelin-1-induced increase in cell number was significantly suppressed by endothelial NOS gene transfer as well as by the mitogen-activated protein kinase inhibitor PD98059. These results indicate that endothelial NOS gene transfer inhibits cellular proliferation via inhibition of the mitogen-activated protein kinase cascade.
Footnotes
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Send reprint requests to: Uichi Ikeda, M.D., Ph.D., Department of Cardiology, Jichi Medical School, Minamikawachi-machi, Tochigi 329-0498, Japan. E-mail:uikeda{at}jichi.ac.jp
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↵1 This work was supported by grants from the Ministry of Health and Welfare (no. 10C-1) and the Ministry of Education, Science, Sports and Culture (no. 10670675).
- Abbreviations:
- NO
- nitric oxide
- NOS
- nitric-oxide synthase
- DMEM/F12
- Dulbecco's modified Eagle's medium/nutrient mixture F12
- fura-2/AM
- fura-2 acetoxymethyl ester
- ETAR
- endothelin type A receptor
- Received April 7, 1999.
- Accepted October 1, 1999.
- The American Society for Pharmacology and Experimental Therapeutics
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