Submitted on July 6, 2007
Accepted on September 20, 2007
Carboxypeptidase B and other kininases of the rat coronary and mesenteric arterial bed perfusates
Eduardo B Oliveira1, Laura L Souza1, Disney O Sivieri Jr2, Luiz B Bispo-da-Silva3, Hugo J V Pereira4, Claudio M Costa Neto5, Marcelo V Sousa6, and Maria Cristina Oliveira Salgado7*
1 Biochemisty and Imunology, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil; , Brazil
2 Pharmacology, University of Sao Paulo, Ribeirao Preto, Brazil; , Brazil
3 Pharmacology, University of Sao Paulo, Brazil
4 Biochemisty and Imunology, University of Sao Paulo, Brazil; , Brazil
5 Biochemisty and Imunology, University of Sao Paulo, Brazil
6 Celular Biology, University of Brasilia, Brasilia, Distrito Federal, Brazil
7 Pharmacology, University of Sao Paulo, Ribeirao Preto, Sao Paulo, Brazil; , Brazil
* To whom correspondence should be addressed. E-mail: mcosalgado{at}gmail.com.
We describe the enzymes that constitute the major bradykinin (BK)-processing pathways in the perfusates of mesenteric arterial bed (MAB) and coronary vessels isolated from Wistar normotensive (WNR) and spontaneously hypertensive (SHR) rats. The contribution of particular proteases to BK degradation was revealed by the combined analysis of fragments generated during incubation of BK with representative perfusate samples and the effect of selective inhibitors on the respective reactions. Marked differences were seen among the perfusates studied; MAB secretes, per minute of perfusion, kininase activity capable of hydrolyzing about 300 pmol of BK/min, which is about 250-fold larger amount on a per unit time basis than that of its coronary counterpart. BK degradation in the coronary perfusate seems to be mediated by ACE, NEP-like enzyme and a MGTA-sensitive basic carboxypeptidase; coronary perfusate of WNR contains an additional BK-degrading enzyme whose specificity resembles that of neurolysin or thimet oligopeptidase. Diversely, a des-Arg9-BK-forming enzyme, responsible for nearly all of the kininase activity of MAB perfusates of WNR and SHR, could be purified by a procedure that involved affinity chromatography over potato carboxypeptidase inhibitor-Sepharose column and shown to be structurally identical with rat pancreatic CPB. Comparable levels of CPB mRNA expression were observed in pancreas, liver, mesentery and kidney, but very low levels were detected in lung, heart, aorta and carotid artery. In conclusion, distinct BK-processing pathways operate in the perfusates of rat MAB and coronary bed, with a substantial participation of a des-Arg9-BK-forming enzyme identical with pancreatic CPB.
