Aims Transforming growth factor (TGF)-β is up-regulated in the diabetic myocardium and may mediate fibroblast activation. We aimed at examining the role of TGF-β-induced fibroblast activation in the pathogenesis of diabetic cardiomyopathy. Methods and results We generated lean and obese db/db mice with fibroblast-specific loss of TbR2, the Type 2 receptor-mediating signaling through all three TGF-β isoforms, and mice with fibroblast-specific Smad3 disruption. Systolic and diastolic function, myocardial fibrosis, and hypertrophy were assessed. Transcriptomic studies and in vitro experiments were used to dissect mechanisms of fibroblast activation. Fibroblast-specific TbR2 loss attenuated systolic and diastolic dysfunction in db/db mice. The protective effects of fibroblast TbR2 loss in db/db mice were associated with attenuated fibrosis and reduced cardiomyocyte hypertrophy, suggesting that in addition to their role in fibrous tissue deposition, TGF-β-stimulated fibroblasts may also exert paracrine actions on cardiomyocytes. Fibroblast-specific Smad3 loss phenocopied the protective effects of fibroblast TbR2 loss in db/db mice. Db/db fibroblasts had increased expression of genes associated with oxidative response (such as Fmo2, encoding flavin-containing monooxygenase 2), matricellular genes (such as Thbs4 and Fbln2), and Lox (encoding lysyl oxidase). Ingenuity pathway analysis (IPA) predicted that neurohumoral mediators, cytokines, and growth factors (such as AGT, TGFB1, and TNF) may serve as important upstream regulators of the transcriptomic profile of diabetic mouse fibroblasts. IPA of scRNA-seq data identified TGFB1, p53, MYC, PDGF-BB, EGFR, and WNT3A/CTNNB1 as important upstream regulators underlying fibroblast activation in db/db hearts. Comparison of the transcriptome of fibroblasts from db/db mice with fibroblast-specific Smad3 loss and db/db Smad3 fl/fl controls identified Thbs4 [encoding thrombospondin-4 (TSP-4), a marker of activated fibroblasts] as a candidate diabetes-induced fibrogenic mediator. However, in vitro experiments showed no significant activating effects of matricellular or intracellular TSP-4 on cardiac fibroblasts. Conclusion Fibroblast-specific TGF-β/Smad3 signaling mediates ventricular fibrosis, hypertrophy, and dysfunction in Type 2 diabetes.

中文翻译：

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成纤维细胞特异性 TGF-β 信号传导介导肥胖糖尿病小鼠的心功能不全、纤维化和肥大


目的 转化生长因子 （TGF）-β 在糖尿病心肌中上调，可能介导成纤维细胞活化。我们旨在检查 TGF β诱导的成纤维细胞活化在糖尿病心肌病发病机制中的作用。方法和结果我们生成了成纤维细胞特异性丢失 TbR2 的瘦和肥胖 db/db 小鼠，TbR2 是通过所有三种 TGF-β 亚型的 2 型受体介导信号，以及成纤维细胞特异性 Smad3 破坏的小鼠。评估收缩和舒张功能、心肌纤维化和肥大。转录组学研究和体外实验用于剖析成纤维细胞活化的机制。成纤维细胞特异性 TbR2 丢失减轻了 db/db 小鼠的收缩和舒张功能障碍。db/db 小鼠成纤维细胞 TbR2 丢失的保护作用与纤维化减弱和心肌细胞肥大减少有关，表明除了在纤维组织沉积中的作用外，TGF β刺激的成纤维细胞还可能对心肌细胞发挥旁分泌作用。成纤维细胞特异性 Smad3 丢失表型复制了成纤维细胞 TbR2 丢失在 db/db 小鼠中的保护作用。Db/db 成纤维细胞与氧化反应相关的基因 （如 Fmo2，编码含黄素的单加氧酶 2）、基质细胞基因 （如 Thbs4 和 Fbln2） 和 Lox （编码赖氨酰氧化酶） 的表达增加。Ingenuity Pathway Analysis （IPA） 预测神经体液介质、细胞因子和生长因子 （如 AGT、TGFB1 和 TNF） 可能是糖尿病小鼠成纤维细胞转录组谱的重要上游调节因子。scRNA-seq 数据的 IPA 确定 TGFB1 、 p53 、 MYC 、 PDGF-BB 、 EGFR 和 WNT3A/CTNNB1 是 db/db 心脏成纤维细胞活化的重要上游调节因子。 来自成纤维细胞特异性 Smad3 丢失的 db/db 小鼠成纤维细胞转录组与 db/db Smad3 fl/fl 对照的比较发现 Thbs4 [编码血小板反应蛋白-4 （TSP-4），活化成纤维细胞的标志物] 是候选糖尿病诱导的纤维化介质。然而，体外实验显示基质细胞或细胞内 TSP-4 对心脏成纤维细胞没有显着的激活作用。结论 成纤维细胞特异性 TGF-β/Smad3 信号传导介导 2 型糖尿病患者的心室纤维化、肥大和功能障碍。