3.2 Endogenous BK-(1-9) fragments induce NO production in vitro
To test whether BK-(1-9) fragments are biologically active, we tested these BK fragments for their ability to induce NO production in different cell types. We used cells from different species (i.e., human, rat and mouse) and known to express the kinin B1 and B2 receptors. As expected, incubation with BK-(1-9) (100 nM) increased NO production in immortalized human glioblastoma cells (U-87 MG), neonatal male rat cardiomyocytes and male adult mouse ventricular myocytes. BK-(1-9) fragments, BK-(1-7) and BK-(1-5), at the same concentration, also induced NO production in these cell types (Figure 2).
We then generated a concentration-response curve for NO production mediated by BK-(1-9), BK-(1-7) and BK-(1-5) in neonatal male rat cardiomyocytes. At all concentrations tested (1, 10, and 100 nM), BK-(1-9) was able to elicit NO production (Figure 3A). The heptapeptide fragment BK-(1-7) induced NO production in neonatal male rat cardiomyocytes but only at the two higher concentrations (10 and 100 nM) in a dose-dependent manner (Figure 3B). The pentapeptide metabolite BK-(1-5) (Figure 3C) was able to induce NO production at all concentration tested, including 1nM, but no significant difference was observed between 10 and 100 nM. These results suggest that BK-(1-7) and BK-(1-5) are not only metabolites of BK-(1-9) in vivo but, in vitro, they were able to induce NO production in human and rodent cells.