EXPRESSION OF ENDOTHELIAL NITRIC-OXIDE SYNTHASE IN HUMAN AND RABBIT GASTROINTESTINAL SMOOTH-MUSCLE CELLS

The aim of this study was to identify the nitric oxide synthase (NOS) isoform expressed in freshly dispersed rabbit gastric smooth muscle ce lls and in cultured rabbit gastric, human intestinal, and guinea pig t aenia coli smooth muscle cells. RT-PCR products of the predicted size (354 bp) were obtained with endothelial NOS (eNOS)-specific primers, b ut not neuronal NOS (nNOS)- or inducible NOS (iNOS)-specific primers, in all smooth muscle preparations except guinea pig taenia coli. Contr ol RT-PCR studies showed absence of the endothelial markers, platelet endothelial cell adhesion molecule-1 (PECAM-1) and vascular endothelia l growth factor receptor (VEGFR), and the interstitial cell marker, c- hit, from cultures of smooth muscle cells. Cloning and sequence analys is showed that the predicted amino acid sequence (117 residues) in rab bit and human smooth muscle cells differed by only one residue from th at of human eNOS. Northern blot analysis, using the PCR-generated and cloned eNOS cDNA from rabbits and humans as probes, demonstrated the e xpression of eNOS mRNA (4.4 kb) in both species. eNOS, but not nNOS or iNOS, transcripts were localized by in situ RT-PCR in single, freshly dispersed rabbit gastric smooth muscle cells; expression was evident in the majority of cells in each preparation. We conclude that eNOS is selectively expressed in rabbit gastric and human intestinal smooth m uscle cells. The results confirm functional evidence for the existence of a constitutive NOS in smooth muscle cells of the gut in different species, except for guinea pig taenia coli.