Effect of bradykinin on cultured bovine corneal endothelial cells

PURPOSE: To clarify the effect of bradykinin on cytosolic free calcium mobilization and cell proliferation in cultured bovine corneal endothelial cells (BCEC). METHODS: The cytosolic free calcium concentration (Ca2+]i) was measured with the InCa(TM) Imaging System after the treatment of bradykinin (10(-11) to 10(-7) M) alone or with the pretreatments of EGTA, bradykinin receptor (Bk1 and Bk2) antagonists and an inhibition of phospholipase C (U-73122). Also, the effect of bradykinin on cell proliferation in BCEC was evaluated using cell counts. RESULTS: In BCEC, [Ca2+]i in the resting state was 87 +/- 9 nM. Bradykinin induced an increment of [Ca2+]i in a concentration-dependent manner and its 50% effective concentration was approximately 5 x 10(-11) M. A [Ca2+]i increment at 10(-8) M bradykinin was inhibited with the pretreatment of EGTA, an extracellular calcium chelator. U-73122 (5 x 10(-6) M) attenuated the bradykinin-induced [Ca2+]i increment. The pretreatment of HOE-140 (Bk2 antagonist) almost attenuated the bradykinin (10(-8) M)-induced [Ca2+]i increase, but des-Arg9-[Leu(8)]-bradykinin (Bk1 antagonist) did not suppress it. To investigate the physiological effect of bradykinin, the effect of bradykinin on cell proliferation was studied. 10(-8) M of bradykinin produced a significant increase in cell numbers. This mitogenic effect of bradykinin was inhibited by the Bk2 antagonist. CONCLUSIONS: Bradykinin-induced stimulation of the signal transduction pathway in BCEC is coupled with the Bk2 type receptor. Furthermore, bradykinin produces the mitogenic effect in BCEC.