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Abstract：
OBJECTIVE: Fibrosis contributes to 45% of deaths in industrialized nations and is characterized by an abnormal accumulation of extracellular matrix (ECM). There are no specific anti-fibrotic treatments for liver fibrosis, and previous unsuccessful attempts at drug development have focused on preventing ECM deposition. Because liver fibrosis is largely acknowledged to be reversible, regulating fibrosis resolution could offer novel therapeutical options. However, little is known about the mechanisms controlling ECM remodeling during resolution. Changes in proteolytic activity are essential for ECM homeostasis and macrophages are an important source of proteases. Herein, in this study we evaluate the role of macrophage-derived cathepsin D (CtsD) during liver fibrosis.
METHODS: CtsD expression and associated pathways were characterized in single-cell RNA sequencing and transcriptomic datasets in human cirrhosis. Liver fibrosis progression, reversion and functional characterization were assessed in novel myeloid-CtsD and hepatocyte-CtsD knock-out mice.
RESULTS: Analysis of single-cell RNA sequencing datasets demonstrated CtsD was expressed in macrophages and hepatocytes in human cirrhosis. Liver fibrosis progression, reversion and functional characterization were assessed in novel myeloid-CtsD (CtsDΔMyel) and hepatocyte-CtsD knock-out mice. CtsD deletion in macrophages, but not in hepatocytes, resulted in enhanced liver fibrosis. Both inflammatory and matrisome proteomic signatures were enriched in fibrotic CtsDΔMyel livers. Besides, CtsDΔMyel liver macrophages displayed functional, phenotypical and secretomic changes, which resulted in a degradomic phenotypical shift, responsible for the defective proteolytic processing of collagen I in vitro and impaired collagen remodeling during fibrosis resolution in vivo. Finally, CtsD-expressing mononuclear phagocytes of cirrhotic human livers were enriched in lysosomal and ECM degradative signaling pathways.
CONCLUSIONS: Our work describes for the first-time CtsD-driven lysosomal activity as a central hub for restorative macrophage function during fibrosis resolution and opens new avenues to explore their degradome landscape to inform drug development.
METHODS: CtsD expression and associated pathways were characterized in single-cell RNA sequencing and transcriptomic datasets in human cirrhosis. Liver fibrosis progression, reversion and functional characterization were assessed in novel myeloid-CtsD and hepatocyte-CtsD knock-out mice.
RESULTS: Analysis of single-cell RNA sequencing datasets demonstrated CtsD was expressed in macrophages and hepatocytes in human cirrhosis. Liver fibrosis progression, reversion and functional characterization were assessed in novel myeloid-CtsD (CtsDΔMyel) and hepatocyte-CtsD knock-out mice. CtsD deletion in macrophages, but not in hepatocytes, resulted in enhanced liver fibrosis. Both inflammatory and matrisome proteomic signatures were enriched in fibrotic CtsDΔMyel livers. Besides, CtsDΔMyel liver macrophages displayed functional, phenotypical and secretomic changes, which resulted in a degradomic phenotypical shift, responsible for the defective proteolytic processing of collagen I in vitro and impaired collagen remodeling during fibrosis resolution in vivo. Finally, CtsD-expressing mononuclear phagocytes of cirrhotic human livers were enriched in lysosomal and ECM degradative signaling pathways.
CONCLUSIONS: Our work describes for the first-time CtsD-driven lysosomal activity as a central hub for restorative macrophage function during fibrosis resolution and opens new avenues to explore their degradome landscape to inform drug development.
摘要：
目的：在工业化国家，纤维化导致45％的死亡，其特征是细胞外基质（ECM）的异常积累。没有针对肝纤维化的特异性抗纤维化治疗,以前在药物开发方面的失败尝试都集中在预防ECM沉积上。因为肝纤维化在很大程度上被认为是可逆的，调节纤维化分辨率可以提供新的治疗选择。然而,关于在分辨过程中控制ECM重塑的机制知之甚少。蛋白水解活性的变化对于ECM稳态是必需的,并且巨噬细胞是蛋白酶的重要来源。在这里，在这项研究中，我们评估了巨噬细胞衍生的组织蛋白酶D（CtsD）在肝纤维化中的作用。
方法：在人类肝硬化的单细胞RNA测序和转录组数据集中对CtsD表达和相关通路进行了表征。肝纤维化进展，在新型髓样CtsD和肝细胞CtsD敲除小鼠中评估了逆转和功能表征。
结果：单细胞RNA测序数据集的分析表明，CtsD在人肝硬化的巨噬细胞和肝细胞中表达。肝纤维化进展，在新型髓样CtsD（CtsDΔMyel）和肝细胞CtsD敲除小鼠中评估了逆转和功能表征。巨噬细胞中的CtsD缺失，但不是在肝细胞中，导致肝纤维化增强。在纤维化CtsDΔMyel肝中富含炎性和基质蛋白质组特征。此外,CtsDΔMyel肝巨噬细胞显示出功能性，表型和分泌组学变化，这导致了退化的表型转变，负责胶原蛋白I的体外蛋白水解加工缺陷和体内纤维化解决过程中胶原蛋白重塑受损。最后,肝硬化人肝脏中表达CtsD的单核吞噬细胞在溶酶体和ECM降解信号通路中富集。
结论：我们的工作首次描述了CtsD驱动的溶酶体活性作为纤维化解决过程中恢复巨噬细胞功能的中心枢纽，并开辟了新的途径来探索其降解组景观，为药物开发提供信息。
方法：在人类肝硬化的单细胞RNA测序和转录组数据集中对CtsD表达和相关通路进行了表征。肝纤维化进展，在新型髓样CtsD和肝细胞CtsD敲除小鼠中评估了逆转和功能表征。
结果：单细胞RNA测序数据集的分析表明，CtsD在人肝硬化的巨噬细胞和肝细胞中表达。肝纤维化进展，在新型髓样CtsD（CtsDΔMyel）和肝细胞CtsD敲除小鼠中评估了逆转和功能表征。巨噬细胞中的CtsD缺失，但不是在肝细胞中，导致肝纤维化增强。在纤维化CtsDΔMyel肝中富含炎性和基质蛋白质组特征。此外,CtsDΔMyel肝巨噬细胞显示出功能性，表型和分泌组学变化，这导致了退化的表型转变，负责胶原蛋白I的体外蛋白水解加工缺陷和体内纤维化解决过程中胶原蛋白重塑受损。最后,肝硬化人肝脏中表达CtsD的单核吞噬细胞在溶酶体和ECM降解信号通路中富集。
结论：我们的工作首次描述了CtsD驱动的溶酶体活性作为纤维化解决过程中恢复巨噬细胞功能的中心枢纽，并开辟了新的途径来探索其降解组景观，为药物开发提供信息。
