Conversion of T-kinin to bradykinin by the rat kidney

Isolated rat kidneys were perfused with T-kinin (TK, Ile-Ser-BK) and bradykinin (BK). HPLC analysis of perfusate samples taken at 2-10 min during the TK perfusion (0.5 nmol/mL initial concentration) showed two peptide peaks, the first one eluting at 14.42 min, the same retention time for standard BK, and the second at 16.20 min, corresponding to that of TK. When BK (0.5 nmol/mL) was perfused, only its corresponding peak was obtained although total BK recovery was reduced quickly, as expected. Using both HPLC analysis and a kinin bioassay on the isolated guinea pig ileum, it was found that 12% of the added TK was converted to BK during the first perfusion cycle (2 min). While the BK recovered (12-14% from the initial TK concentration) was maintained at a similar proportion between the 2nd and the 10th min of perfusion, the rate of TK disappearance, as well as its full recovery from the perfusate, indicated further fragmentation of peptides during kinin perfusion. In the presence of 5 microM DL-mercaptomethyl-3-guanidino-ethylthiopropanoic acid (Mergetpa), an inhibitor of plasma carboxypeptidase N (EC 3.4.17.3), the rate of conversion of TK to BK was not affected. On the other hand, the kinase II inhibitor bradykinin potentiating peptide 9a (BPP9a) increased both the proportion of TK converted to BK and the disappearance rate of TK from the perfusate. In the presence of BPP9a, the rate of BK production increased from 1.5 +/- 0.2 to 7.6 +/- 0.9 nmol/min. Furthermore, the recovery of BK was reduced during the first 2 min of perfusion to 7.6% and the conversion rate to 0.9 nmol/min when TK was perfused into the kidney in the presence of 10 microM bestatin, a known inhibitor of aminopeptidases. These data indicate that in the kidney TK is converted to BK, probably by aminopeptidase M, thus suggesting that BK is, in fact, an additional and functional kinin, inducing physiological and/or pathophysiological effects in the rat kidney in which TK is the main kinin released.