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Single-cell RNA sequencing reveals the heterogeneity and intercellular communication of hepatic stellate cells and macrophages during liver fibrosis.

ABSTRACT: Uncontrolled and excessive progression of liver fibrosis is thought to be the prevalent pathophysiological cause of liver cirrhosis and hepatocellular cancer, and there are currently no effective antifibrotic therapeutic options available. Intercellular communication and cellular heterogeneity in the liver are involved in the progression of liver fibrosis, but the exact nature of the cellular phenotypic changes and patterns of interregulatory remain unclear. Here, we performed single-cell RNA sequencing on nonparenchymal cells (NPCs) isolated from normal and fibrotic mouse livers. We identified eight main types of cells, including endothelial cells, hepatocytes, dendritic cells, B cells, natural killer/T (NK/T) cells, hepatic stellate cells (HSCs), cholangiocytes and macrophages, and revealed that macrophages and HSCs exhibit the most variance in transcriptional profile. Further analyses of HSCs and macrophage subpopulations and ligand-receptor interaction revealed a high heterogeneity characterization and tightly interregulated network of these two groups of cells in liver fibrosis. Finally, we uncovered a profibrotic Thbs1+ macrophage subcluster, which expands in mouse and human fibrotic livers, activating HSCs via PI3K/AKT/mTOR signaling pathway. Our findings decode unanticipated insights into the heterogeneity of HSCs and macrophages and their intercellular crosstalk at a single-cell level, and may provide potential therapeutic strategies in liver fibrosis.

SUBMITTER: Cheng S

PROVIDER: S-EPMC10505372 | biostudies-literature | 2023 Oct

REPOSITORIES: biostudies-literature

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Single-cell RNA sequencing reveals the heterogeneity and intercellular communication of hepatic stellate cells and macrophages during liver fibrosis.

Cheng Sheng S Zou Yunhan Y Zhang Man M Bai Shihao S Tao Kun K Wu Jiaoxiang J Shi Yi Y Wu Yuelan Y Lu Yinzhong Y He Kunyan K Sun Peng P Su Xianbin X Hou Shangwei S Han Bo B

MedComm 20230917 5

Uncontrolled and excessive progression of liver fibrosis is thought to be the prevalent pathophysiological cause of liver cirrhosis and hepatocellular cancer, and there are currently no effective antifibrotic therapeutic options available. Intercellular communication and cellular heterogeneity in the liver are involved in the progression of liver fibrosis, but the exact nature of the cellular phenotypic changes and patterns of interregulatory remain unclear. Here, we performed single-cell RNA se ...[more]

PMID: 37724132

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Project description:Background & aimsThe multiple vital functions of the human liver are performed by highly specialised parenchymal and non-parenchymal cells organised in complex collaborative sinusoidal units. Although crucial for homeostasis, the cellular make-up of the human liver remains to be fully elucidated. Here, single-cell RNA-sequencing was used to unravel the heterogeneity of human liver cells, in particular of hepatocytes (HEPs) and hepatic stellate cells (HSCs).MethodThe transcriptome of ~25,000 freshly isolated human liver cells was profiled using droplet-based RNA-sequencing. Recently published data sets and RNA in situ hybridisation were integrated to validate and locate newly identified cell populations.ResultsIn total, 22 cell populations were annotated that reflected the heterogeneity of human parenchymal and non-parenchymal liver cells. More than 20,000 HEPs were ordered along the portocentral axis to confirm known, and reveal previously undescribed, zonated liver functions. The existence of 2 subpopulations of human HSCs with unique gene expression signatures and distinct intralobular localisation was revealed (i.e. portal and central vein-concentrated GPC3 + HSCs and perisinusoidally located DBH + HSCs). In particular, these data suggest that, although both subpopulations collaborate in the production and organisation of extracellular matrix, GPC3 + HSCs specifically express genes involved in the metabolism of glycosaminoglycans, whereas DBH + HSCs display a gene signature that is reminiscent of antigen-presenting cells.ConclusionsThis study highlights metabolic zonation as a key determinant of HEP transcriptomic heterogeneity and, for the first time, outlines the existence of heterogeneous HSC subpopulations in the human liver. These findings call for further research on the functional implications of liver cell heterogeneity in health and disease.Lay summaryThis study resolves the cellular landscape of the human liver in an unbiased manner and at high resolution to provide new insights into human liver cell biology. The results highlight the physiological heterogeneity of human hepatic stellate cells.

Project description:The fibrogenic wound-healing response in liver increases stiffness. Stiffness mechanotransduction, in turn, amplifies fibrogenesis. Here, we aimed to understand the distribution of stiffness in fibrotic liver, how it impacts hepatic stellate cell (HSC) heterogeneity, and identify mechanisms by which stiffness amplifies fibrogenic responses. Magnetic resonance elastography and atomic force microscopy demonstrated a heterogeneous distribution of liver stiffness at macroscopic and microscopic levels, respectively, in a carbon tetrachloride (CCl4) mouse model of liver fibrosis as compared with controls. High stiffness was mainly attributed to extracellular matrix dense areas. To identify a stiffness-sensitive HSC subpopulation, we performed single-cell RNA sequencing (scRNA-seq) on primary HSCs derived from healthy versus CCl4-treated mice. A subcluster of HSCs was matrix-associated with the most upregulated pathway in this subpopulation being focal adhesion signaling, including a specific protein termed four and a half LIM domains protein 2 (FHL2). In vitro, FHL2 expression was increased in primary human HSCs cultured on stiff matrix as compared with HSCs on soft matrix. Moreover, FHL2 knockdown inhibited fibronectin and collagen 1 expression, whereas its overexpression promoted matrix production. In summary, we demonstrate stiffness heterogeneity at the whole organ, lobular, and cellular level, which drives an amplification loop of fibrogenesis through specific focal adhesion molecular pathways.NEW & NOTEWORTHY The fibrogenic wound-healing response in liver increases stiffness. Here, macro and microheterogeneity of liver stiffness correlate with HSC heterogeneity in a hepatic fibrosis mouse model. Fibrogenic HSCs localized in stiff collagen-high areas upregulate the expression of focal adhesion molecule FHL2, which, in turn, promotes extracellular matrix protein expression. These results demonstrate that stiffness heterogeneity at the whole organ, lobular, and cellular level drives an amplification loop of fibrogenesis through specific focal adhesion molecular pathways.

Dataset Information

Single-cell RNA sequencing reveals the heterogeneity and intercellular communication of hepatic stellate cells and macrophages during liver fibrosis.

Publications

Single-cell RNA sequencing reveals the heterogeneity and intercellular communication of hepatic stellate cells and macrophages during liver fibrosis.

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