Abstract：
OBJECTIVE: Excessive fibrogenesis is associated with adverse cardiac remodelling and heart failure. The myofibroblast, primarily derived from resident fibroblast, is the effector cell type in cardiac fibrosis. Megakaryocytic leukaemia 1 (MKL1) is considered the master regulator of fibroblast-myofibroblast transition (FMyT). The underlying transcriptional mechanism is not completely understood. Our goal was to identify novel transcriptional targets of MKL1 that might regulate FMyT and contribute to cardiac fibrosis.
METHODS: RNA sequencing (RNA-seq) performed in primary cardiac fibroblasts identified insulin-like growth factor binding protein 5 (IGFBP5) as one of the genes most significantly up-regulated by constitutively active (CA) MKL1 over-expression. IGFBP5 expression was detected in heart failure tissues using RT-qPCR and western blots.
RESULTS: Once activated, IGFBP5 translocated to the nucleus to elicit a pro-FMyT transcriptional programme. Consistently, IGFBP5 knockdown blocked FMyT in vitro and dampened cardiac fibrosis in mice. Of interest, IGFBP5 interacted with nuclear factor of activated T-cell 4 (NFAT4) to stimulate the transcription of microfibril-associated protein 5 (MFAP5). MFAP5 contributed to FMyT and cardiac fibrosis by enabling sterol response element binding protein 2 (SREBP2)-dependent cholesterol synthesis.
CONCLUSIONS: Our data unveil a previously unrecognized transcriptional cascade, initiated by IGFBP5, that promotes FMyT and cardiac fibrosis. Screening for small-molecule compounds that target this axis could yield potential therapeutics against adverse cardiac remodelling.
METHODS: RNA sequencing (RNA-seq) performed in primary cardiac fibroblasts identified insulin-like growth factor binding protein 5 (IGFBP5) as one of the genes most significantly up-regulated by constitutively active (CA) MKL1 over-expression. IGFBP5 expression was detected in heart failure tissues using RT-qPCR and western blots.
RESULTS: Once activated, IGFBP5 translocated to the nucleus to elicit a pro-FMyT transcriptional programme. Consistently, IGFBP5 knockdown blocked FMyT in vitro and dampened cardiac fibrosis in mice. Of interest, IGFBP5 interacted with nuclear factor of activated T-cell 4 (NFAT4) to stimulate the transcription of microfibril-associated protein 5 (MFAP5). MFAP5 contributed to FMyT and cardiac fibrosis by enabling sterol response element binding protein 2 (SREBP2)-dependent cholesterol synthesis.
CONCLUSIONS: Our data unveil a previously unrecognized transcriptional cascade, initiated by IGFBP5, that promotes FMyT and cardiac fibrosis. Screening for small-molecule compounds that target this axis could yield potential therapeutics against adverse cardiac remodelling.
摘要：
目的：过度的纤维化与不良心脏重塑和心力衰竭有关。成肌纤维细胞,主要来源于常驻成纤维细胞，是心脏纤维化中的效应细胞类型。巨核细胞白血病1（MKL1）被认为是成纤维细胞-肌成纤维细胞转化（FMyT）的主要调节因子。潜在的转录机制尚未完全了解。我们的目标是鉴定可能调节FMyT并有助于心脏纤维化的MKL1的新转录靶标。
方法：在原代心脏成纤维细胞中进行的RNA测序（RNA-seq）鉴定胰岛素样生长因子结合蛋白5（IGFBP5）是组成型活性（CA）MKL1过表达最显著上调的基因之一。使用RT-qPCR和蛋白质印迹在心力衰竭组织中检测IGFBP5表达。
结果：一旦激活，IGFBP5易位到细胞核以引发pro-FMyT转录程序。始终如一，IGFBP5敲低在体外阻断FMyT并抑制小鼠心脏纤维化。感兴趣的，IGFBP5与活化T细胞核因子4（NFAT4）相互作用，刺激微纤维相关蛋白5（MFAP5）的转录。MFAP5通过促进固醇反应元件结合蛋白2（SREBP2）依赖性胆固醇合成而促进FMyT和心脏纤维化。
结论：我们的数据揭示了以前未被识别的转录级联，由IGFBP5发起，促进FMyT和心脏纤维化。筛选靶向该轴的小分子化合物可以产生针对不利心脏重塑的潜在治疗剂。
方法：在原代心脏成纤维细胞中进行的RNA测序（RNA-seq）鉴定胰岛素样生长因子结合蛋白5（IGFBP5）是组成型活性（CA）MKL1过表达最显著上调的基因之一。使用RT-qPCR和蛋白质印迹在心力衰竭组织中检测IGFBP5表达。
结果：一旦激活，IGFBP5易位到细胞核以引发pro-FMyT转录程序。始终如一，IGFBP5敲低在体外阻断FMyT并抑制小鼠心脏纤维化。感兴趣的，IGFBP5与活化T细胞核因子4（NFAT4）相互作用，刺激微纤维相关蛋白5（MFAP5）的转录。MFAP5通过促进固醇反应元件结合蛋白2（SREBP2）依赖性胆固醇合成而促进FMyT和心脏纤维化。
结论：我们的数据揭示了以前未被识别的转录级联，由IGFBP5发起，促进FMyT和心脏纤维化。筛选靶向该轴的小分子化合物可以产生针对不利心脏重塑的潜在治疗剂。
