Up‐regulation of [3H]‐des‐Arg10‐kallidin binding to the bradykinin B1 receptor by interleukin‐1β in isolated smooth muscle cells: correlation with B1 agonist‐induced PGI2 production

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Abstract
1 Binding of the specific bradykinin B1 receptor agonist, [3H]-des-Arg10-kallidin (-KD) was investigated in smooth muscle cells (SMC) isolated from rabbit mesenteric arteries (RMA). 2 [3H]-des-Arg10-KD specifically bound to interleukin-1 (IL-1)-treated RMA-SMC in a saturable fashion with an equilibrium dissociation constant (KD) of 0.3–0.5 nm. The number of binding sites per cell was 20,000–35,000. Kinins inhibited [3H]-des-Arg10-KD binding to RMA-SMC with an order of potency very similar to that observed in typical B1 specific bioassays: des-Arg9-bradykinin (BK) ≅ KD > > BK. Furthermore, the B1 receptor antagonist [Leu8]des-Arg9-BK inhibited [3H]-des-Arg10-KD binding with an IC50 of 43 nm as expected for its effect at B1 receptors. The B2 receptor antagonists, NPC 567 and Hoe 140 only affected [3H]-des-Arg10-KD binding at very high concentrations (IC50 = 0.8 μm and IC50 > 10 μm, respectively). 3 Des-Arg9-BK (BL agonist) and [Hyp3]Tyr(Me)8-BK (B2 agonist) did not induce prostacyclin (PGI2) production by RMA-SMC. Lipopolysaccharide (LPS) treatment of the cells did not affect the B1 agonist response whereas IL-1β treatment produced a 7 fold increase in des-Arg9-BK-stimulated PGI2 production. IL-1β also stimulated the response to B2 agonists. 4 Des-Arg9-BK-induced PGI2 secretion in IL-1-primed RMA-SMC was mediated by B1 receptors since it was inhibited by [Leu8]des-Arg9-BK (IC50 = 56–73 nm) but not by Hoe 140. High concentrations of NPC 567 (IC50 = 2.4 μm) were required to inhibit PGI2 production induced by B1 agonists. 5 IL-1-treated RMA-SMC displayed a 5 fold increase in the number of B1 receptors without modification of the affinity constant, thus establishing a possible relationship between the receptor density and the IL-1-primed B1 response. 6 LPS treatment of the cells induced a 4 fold increase in B1 receptor number without modifying PGI2 secretion. This observation suggests that IL-1 but not LPS, in addition to increase in the number of receptors, signals the cell to permit the coupling of B1 receptors to the PLA2/cyclo-oxygenase pathway.

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