In vitro interactions between ramiprilat and angiotensin I-converting enzyme in endothelial cells.

Ramiprilat is an angiotensin I-converting enzyme (ACE) inhibitor whose particular lipophilicity may modify its inhibitory activity on the cellular form of this enzyme in comparison to ACE inhibitors that are more hydrophilic. The inhibitory activity of ramiprilat on cellular ACE and its binding to plasma membrane ACE have been studied in cultures of pig vascular endothelial cells. ACE activity in pig pulmonary artery endothelial cells is completely inhibited by 100 nM ramiprilat; the IC50 is 2 nM, whatever the form of ACE: soluble ACE released into the culture medium, cellular ACE studied in a cell monolayer homogenate, or tissue ACE purified from pig lung tissue. When added directly to whole living cells in their culture medium, 100 nM ramiprilat inhibits less than 80% of ACE activity in the supernatant while this dose inhibits the cellular form of the enzyme completely, suggesting that ramiprilat reaches and more specifically inhibits membrane-bound ACE rather than ACE secreted by the monolayer of endothelial cells. [3H]ramiprilat binds specifically to the membrane of cultured endothelial cells, in a time and dose-dependent manner (Kd = 6 nM; Bmax = 1,600 fmol/mg of protein); specificity is confirmed by the fact that an anti-ACE antibody prevents binding of [3H]ramiprilat and that both cold ramiprilat as well as other synthetic inhibitor compounds (captopril and enalaprilat) displace [3H]ramiprilat in a dose-dependent manner. Angiotensin I and bradykinin, natural substrates of ACE, do not induce displacement of [3H]ramiprilat bound to the luminal plasma membrane of endothelial cells.(ABSTRACT TRUNCATED AT 250 WORDS)