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.
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。