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Am J Physiol Heart Circ Physiol 277: H1317-H1325, 1999;
0363-6135/99 $5.00
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Vol. 277, Issue 4, H1317-H1325, October 1999

NO from smooth muscle cells decreases NOS expression in endothelial cells: role of TNF-alpha

Trinidad de Frutos, Lourdes Sánchez de Miguel, Margarita García-Durán, Fernando González-Fernández, Juan A. Rodríguez-Feo, Mercedes Montón, José Guerra, Jerónimo Farré, Santos Casado, and Antonio López-Farré

Hypertension and Cardiovascular Research Laboratory, Fundación Jiménez Díaz, 28040 Madrid, Spain

Despite the evidence that cytokines stimulate nitric oxide (NO) production by inducible nitric oxide synthase (iNOS), several reports recently demonstrated that the hypotensive response related to endothelial nitric oxide synthase (eNOS) activity could be inhibited by the same cytokines. The aim of the present work was to analyze whether NO generated by vascular smooth muscle cells (VSMC) could modify eNOS protein expression in endothelial cells. Bovine aortic endothelial cells (BAEC) and bovine VSMC (BVSMC) in coculture were used for the study. Interleukin-1beta (IL-1beta , 10 ng/ml)-treated BVSMC, which expressed iNOS protein, decreased eNOS protein expression in BAEC. The presence of NO antagonists Nomega -nitro-L-arginine methyl ester (10-3 mol/l) or NG-monomethyl-L-arginine (10-3 mol/l) prevented the decrease in eNOS protein expression induced by IL-1beta -treated BVSMC. Surprisingly, two different NO donors, 3-morpholinosydnonimine (10-4 mol/l) and S-nitroso-N-acetyl-D,L-penicillamine (10-4 mol/l), failed to modify eNOS expression in BAEC, suggesting the existence of a diffusible mediator released from IL-1beta -treated BVSMC that acts on endothelial cells by reducing eNOS expression. The presence of NO antagonists reduced tumor necrosis factor-alpha (TNF-alpha ) production by IL-1beta -stimulated BVSMC. This effect was also produced in the presence of a protein kinase G inhibitor, guanosine-5'-O-(2-thiodiphosphate) trilithium salt. A polyclonal antibody against TNF-alpha prevented eNOS expression in the BAEC-BVSMC coculture. In conclusion, NO by itself failed to modify eNOS protein expression in endothelial cells but increased TNF-alpha generation by IL-1beta -stimulated BVSMC and, in this way, reduced eNOS expression in the endothelium.

coculture; endothelial dysfunction; interleukin-1beta ; nitric oxide donors; tumor necrosis factor-alpha


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