Interactions between carboxypeptidase M and kinin B1 receptor in endothelial cells.

Introduction: Carboxypeptidase M (CPM) is a glycosylphosphatidylinositol anchored enzyme that plays an important role in the kallikrein-kinin system (KKS). CPM catalytic domain hydrolyzes Arg from C-terminal peptides (i.e., bradykinin and kallidin), generating des-Arg-kinins, the agonists of B receptor (BR). It is known that CPM and kinin BR are co-localized in the plasma membrane microdomains, where they interact with each other, facilitating receptor signaling.

Aims: We hypothesized here that this CPM-BR interaction could also affect the activity of the enzyme.

Methods: Thus, in this work, we evaluated the impact of BR presence or absence on CPM activity and expression, using primary culture of microvascular endothelial cells from wild-type, kinin BR knockout mice (B ), and transgenic rats overexpressing B receptor exclusively in the endothelium. In addition, HEK293T cells, as wells as B primary culture of endothelial cells, both transfected with BR, were also used.

Results: CPM expression and activity were downregulated in cells of knockout mice compared to control and this reduction was rescued after BR transfection. Cells overexpressing BR presented higher levels of CPM mRNA, protein, and activity. This profile was reverted by pre-incubation with the BR antagonist, R715, in highly expressing receptor cells.

Conclusions: Our data show that kinin BR positively modulates both CPM expression and activity, suggesting that CPM-BR interaction in membrane microdomains might affect enzyme activity, beyond interfering in receptors signaling. This work highlights the interactions among different components of KKS and contributes to a better understanding of its patho-physiological role.