Haddad Youssef, Couture Réjean
Department of Pharmacology and Physiology, Faculty of Medicine, Université de Montréal, Montréal, QC, Canada.
Front Physiol. 2017 Oct 31;8:861. doi: 10.3389/fphys.2017.00861. eCollection 2017.
Kinin B1 receptor (B1R) enhanced superoxide anion ([Formula: see text]) production in the vasculature of diabetic rats. This study investigates the induction and distribution of B1R in diabetic blood vessels and addresses the hypothesis that B1R is co-localized with NADPH oxidase (NOX1 and NOX2) and produces its activation via protein kinase C (PKC). Diabetes was induced in rats with streptozotocin (STZ 65 mg.kg, i.p.). Two weeks later, the production of [Formula: see text] was measured in aorta rings in response to the B1R agonist (Sar[D-Phe]-des-Arg-BK, 20 μM) by the method of lucigenin-enhanced chemiluminescence. Various inhibitors were added (10 μM) to block PKC (Ro-31-8220), PKCβ1/2 (LY333531), or NADPH oxidase (Diphenyleneiodonium). The cellular localization of B1R was studied in the aorta, popliteal artery, and renal glomerulus/arteries by immunofluorescence and confocal microscopy with markers of endothelial cells (anti-RECA-1), macrophages (anti-CD11), vascular smooth muscle cells (anti-SMA), and NADPH oxidase (anti-NOX1 and NOX2). Although B1R was largely distributed in resistant vessels, it was sparsely expressed in the aorta's endothelium. The greater basal production of [Formula: see text] in STZ-diabetic aorta was significantly enhanced by the B1R agonist (15-45 min). The peak response to the agonist (30 min) was inhibited by all inhibitors. Immunofluorescent staining for B1R, NOX1, and NOX2 was significantly increased in endothelial cells, vascular smooth muscle cells, and macrophages of STZ-diabetic aorta on which they were found co-localized. Data showed that B1R enhanced [Formula: see text] by activating vascular NADPH oxidase through PKCβ1/2. This was substantiated by the cellular co-localization of B1R with NOX1 and NOX2 and opens the possibility that B1R-enhanced oxidative stress is derived from vascular and infiltrating immune cells in diabetes.
激肽B1受体（B1R）可增强糖尿病大鼠血管中超氧阴离子（[公式：见原文]）的产生。本研究调查了B1R在糖尿病血管中的诱导情况和分布，并探讨了B1R与NADPH氧化酶（NOX1和NOX2）共定位并通过蛋白激酶C（PKC）激活的假说。用链脲佐菌素（STZ 65 mg·kg，腹腔注射）诱导大鼠患糖尿病。两周后，采用光泽精增强化学发光法测定主动脉环对B1R激动剂（Sar[D-Phe]-des-Arg-BK，20 μM）刺激的[公式：见原文]产生量。加入各种抑制剂（10 μM）以阻断PKC（Ro-31-8220）、PKCβ1/2（LY333531）或NADPH氧化酶（二亚苯基碘鎓）。通过免疫荧光和共聚焦显微镜，使用内皮细胞标志物（抗RECA-1）、巨噬细胞标志物（抗CD11）、血管平滑肌细胞标志物（抗平滑肌肌动蛋白）和NADPH氧化酶标志物（抗NOX1和NOX2），研究B1R在主动脉、腘动脉和肾小球/动脉中的细胞定位。虽然B1R主要分布在阻力血管中，但在主动脉内皮中表达稀少。B1R激动剂可显著增强STZ糖尿病大鼠主动脉中更高的基础[公式：见原文]产生量（15 - 45分钟）。所有抑制剂均抑制了对激动剂的峰值反应（30分钟）。在STZ糖尿病大鼠主动脉的内皮细胞、血管平滑肌细胞和巨噬细胞中，B1R、NOX1和NOX2的免疫荧光染色显著增加，且发现它们共定位。数据表明，B1R通过PKCβ1/2激活血管NADPH氧化酶来增强[公式：见原文]。B1R与NOX1和NOX2的细胞共定位证实了这一点，并揭示了B1R增强的氧化应激可能源自糖尿病中的血管和浸润免疫细胞的可能性。
