Aims Human and mouse cardiac beta3-adrenergic receptors (beta3AR) exert antipathetic effects to those of beta1-2AR stimulation. We examined their role in modulating myocardial remodelling, particularly fibrosis in response to haemodynamic stress. Methods and results Mice with cardiac myocyte-specific expression of beta3AR (ADRB3-tg) or tamoxifen-inducible homozygous deletion (c-Adrb3-ko, with loxP-targeted Adrb3) were submitted to transaortic constriction. A superfusion assay was used for proteomic analysis of paracrine mediators between beta3AR-expressing cardiac myocytes and cardiac fibroblasts cultured separately. We show that cardiac beta3AR attenuate myocardial fibrosis in response to haemodynamic stress. Interstitial fibrosis and collagen content were reduced in ADRB3-tg, but augmented in c-Adrb3-ko. ADRB3 and collagen (COL1A1) expression were also inversely related in ventricular biopsies of patients with valve disease. Incubation of cardiac fibroblasts with media conditioned by hypertrophic myocytes induced fibroblast proliferation, myo-differentiation, and collagen production. These effects were abrogated upon ADRB3 expression in myocytes. Comparative shotgun proteomic analysis of the myocyte secretomes revealed a number of factors differentially regulated by beta3AR, among which connective tissue growth factor [CTGF (CCN2)] was prominently reduced. CTGF was similarly reduced in stressed hearts from ADRB3-tg, but increased in hearts from c-Adrb3-ko mice. CTGF expression was mediated by reactive oxygen species production which was reduced by ADRB3 expression in vitro and in vivo. This antioxidant and anti-fibrotic effect involved beta3AR coupling to the neuronal isoform of nitric oxide synthase (nNOS) in cardiac myocytes, as both were abrogated upon nNOS inhibition or Nos1 homozygous deletion. Conclusion Cardiac beta3AR protect from fibrosis in response to haemodynamic stress by modulating nitric oxide and oxidant stress-dependent paracrine signaling to fibroblasts. Specific agonism at beta3AR may offer a new therapeutic modality to prevent cardiac fibrosis.

中文翻译：

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心肌细胞β3-肾上腺素能受体通过调节氧化应激依赖性旁分泌信号传导来预防心肌纤维化

目的 人和小鼠心脏β3-肾上腺素能受体（β3AR）对β1-2AR 刺激产生抗病作用。我们检查了它们在调节心肌重塑中的作用，特别是响应血流动力学压力的纤维化。方法和结果 心肌细胞特异性表达β3AR (ADRB3-tg) 或他莫昔芬诱导的纯合缺失(c-Adrb3-ko，具有loxP 靶向Adrb3) 的小鼠接受经主动脉缩窄。灌注试验用于表达β3AR的心肌细胞和分别培养的心脏成纤维细胞之间的旁分泌介质的蛋白质组学分析。我们表明心脏β3AR 减弱心肌纤维化以响应血流动力学压力。ADRB3-tg 中的间质纤维化和胶原蛋白含量减少，但在 c-Adrb3-ko 中增加。ADRB3 和胶原蛋白 (COL1A1) 的表达在瓣膜病患者的心室活检中也呈负相关。心脏成纤维细胞与肥大肌细胞调节的培养基一起孵育可诱导成纤维细胞增殖、肌分化和胶原蛋白的产生。这些影响在肌细胞中的 ADRB3 表达后被消除。肌细胞分泌组的比较鸟枪法蛋白质组学分析揭示了许多受 β3AR 差异调节的因子，其中结缔组织生长因子 [CTGF (CCN2)] 显着降低。CTGF 在来自 ADRB3-tg 的应激心脏中同样减少，但在来自 c-Adrb3-ko 小鼠的心脏中增加。CTGF 表达由活性氧物质产生介导，活性氧物质在体外和体内被 ADRB3 表达降低。这种抗氧化和抗纤维化作用涉及 β3AR 与心肌细胞中一氧化氮合酶 (nNOS) 的神经元异构体的偶联，因为两者在 nNOS 抑制或 Nos1 纯合缺失后都被废除。结论心脏β3AR通过调节一氧化氮和氧化应激依赖性旁分泌信号传导至成纤维细胞来防止纤维化以响应血流动力学应激。β3AR 的特异性激动可能提供一种新的治疗方式来预防心脏纤维化。