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Nonalcoholic liver disease is a component of metabolic syndrome associated with obesity, insulin resistance, and hyperlipidemia. Excessive alcohol consumption may accelerate the progression of steatosis, steatohepatitis, and fibrosis. While simple steatosis is considered a benign condition, nonalcoholic steatohepatitis with inflammation and fibrosis may progress to cirrhosis, liver failure, and hepatocellular cancer. Studies in rodent experimental models and primary cell cultures have demonstrated several common cellular and molecular mechanisms in the pathogenesis and regression of liver fibrosis. Chronic injury and death of hepatocytes cause the recruitment of myeloid cells, secretion of inflammatory and fibrogenic cytokines, and activation of myofibroblasts, resulting in liver fibrosis. In this review, we discuss the role of metabolically injured hepatocytes in the pathogenesis of nonalcoholic steatohepatitis and alcohol-associated liver disease. Specifically, the role of chemokine production and de novo lipogenesis in the development of steatotic hepatocytes and the pathways of steatosis regulation are discussed.

Advanced hepatic fibrosis, driven by the activation of hepatic stellate cells (HSCs), affects millions worldwide and is the strongest predictor of mortality in nonalcoholic steatohepatitis (NASH); however, there are no approved antifibrotic therapies. To identify antifibrotic drug targets, we integrated progressive transcriptomic and morphological responses that accompany HSC activation in advanced disease using single-nucleus RNA sequencing and tissue clearing in a robust murine NASH model. In advanced fibrosis, we found that an autocrine HSC signaling circuit emerged that was composed of 68 receptor-ligand interactions conserved between murine and human NASH. These predicted interactions were supported by the parallel appearance of markedly increased direct stellate cell-cell contacts in murine NASH. As proof of principle, pharmacological inhibition of one such autocrine interaction, neurotrophic receptor tyrosine kinase 3–neurotrophin 3, inhibited human HSC activation in culture and reversed advanced murine NASH fibrosis. In summary, we uncovered a repertoire of antifibrotic drug targets underlying advanced fibrosis in vivo. The findings suggest a therapeutic paradigm in which stage-specific therapies could yield enhanced antifibrotic efficacy in patients with advanced hepatic fibrosis. This is all of the human data samples and analysis for the paperAn autocrine signaling circuit in hepatic stellate cells underlies advanced fibrosis in nonalcoholic steatohepatitis. GEO data here: GSE212837 The folderliver_data_cellranger_all.zipcontains the raw and filtered cellranger output for every sample. This allows reprocessing of all cells including the use of methods that use empty droplets to calculate ambient RNA. The folderAutocrine_liver_paper_jupyter_notebooks.zipcontains all of the notebooks used for analysis: Autocrine_signaling_liver_data_merge_all_select_celltype_subsets.ipynb is the initial noteboook for processing all datasets and merging. It also contains in the first cell instructions for creating a conda environment that will allow all of the tools to run. There are 5 notebooks for qc and cleaning of each celltype: Autocrine_signaling_liver_data_Stellate_cell_recluster.ipynb,Autocrine_signaling_liver_data_Hepatocyte_cell_recluster.ipynb, Autocrine_signaling_liver_data_Endo_cell_recluster.ipynb,Autocrine_signaling_liver_data_Cholangiocyte_cell_recluster.ipynb, and Autocrine_signaling_liver_data_NKTcell_cell_recluster.ipynb There is a final notebook for re-merging the cleaned celltype objects and final clustering, DE and analysis:Autocrine_signaling_liver_data_recluster_all_cleaned_celltypes_analysis.ipynb The fileraw_all_cellranger.h5ad.gzis all cells passing cellranger filter annotated with celltype and ambient RNA and doublet scoring, but otherwise no cells or genes have been removed. The foldercelltype_subsets_v1.zipcontains the roughly filtered celltypes for each of the celltypes. The foldercleaned_celltype_subsets_for_merge.zipcontains the final qc'd and filtered celltypes that are merged for final analysis. It also contains the final merged and batch normalized object. The python scriptcellphone_db_liver_all_cells_clean.pycontains a script for running cellphoneDB on final data.

Background and aims In clinical and experimental NASH, the origin of the scar-forming myofibroblast is the HSC. We used foz/foz mice on a Western diet to characterize in detail the phenotypic changes of HSCs in a NASH model. Approach and results We examined the single-cell expression profiles (scRNA sequencing) of HSCs purified from the normal livers of foz/foz mice on a chow diet, in NASH with fibrosis of foz/foz mice on a Western diet, and in livers during regression of NASH after switching back to a chow diet. Selected genes were analyzed using immunohistochemistry, quantitative real-time PCR, and short hairpin RNA knockdown in primary mouse HSCs. Our analysis of the normal liver identified two distinct clusters of quiescent HSCs that correspond to their acinar position of either pericentral vein or periportal vein. The NASH livers had four distinct HSC clusters, including one representing the classic fibrogenic myofibroblast. The three other HSC clusters consisted of a proliferating cluster, an intermediate activated cluster, and an immune and inflammatory cluster. The livers with NASH regression had one cluster of inactivated HSCs, which was similar to, but distinct from, the quiescent HSCs. Conclusions Analysis of single-cell RNA sequencing in combination with an interrogation of previous studies revealed an unanticipated heterogeneity of HSC phenotypes under normal and injured states.

Background & Aims How benign liver steatosis progresses to nonalcoholic steatohepatitis (NASH), fibrosis, and hepatocellular carcinoma (HCC) remains elusive. NASH progression entails diverse pathogenic mechanisms and relies on complex cross-talk between multiple tissues such as the gut, adipose tissues, liver, and the brain. Using a hyperphagic mouse fed with a Western diet (WD), we aimed to elucidate the cross-talk and kinetics of hepatic and extrahepatic alterations during NASH–HCC progression, as well as regression. Methods Hyperphagic mice lacking a functional Alms1 gene (Foz/Foz) and wild-type littermates were fed WD or standard chow for 12 weeks for NASH/fibrosis and for 24 weeks for HCC development. NASH regression was modeled by switching back to normal chow after NASH development. Results Foz+WD mice were steatotic within 1 to 2 weeks, developed NASH by 4 weeks, and grade 3 fibrosis by 12 weeks, accompanied by chronic kidney injury. Foz+WD mice that continued on WD progressed to cirrhosis and HCC within 24 weeks and had reduced survival as a result of cardiac dysfunction. However, NASH mice that were switched to normal chow showed NASH regression, improved survival, and did not develop HCC. Transcriptomic and histologic analyses of Foz/Foz NASH liver showed strong concordance with human NASH. NASH was preceded by an early disruption of gut barrier, microbial dysbiosis, lipopolysaccharide leakage, and intestinal inflammation. This led to acute-phase liver inflammation in Foz+WD mice, characterized by neutrophil infiltration and increased levels of several chemokines/cytokines. The liver cytokine/chemokine profile evolved as NASH progressed, with subsequent predominance by monocyte recruitment. Conclusions The Foz+WD model closely mimics the pathobiology and gene signature of human NASH with fibrosis and subsequent HCC. Foz+WD mice provide a robust and relevant preclinical model of NASH, NASH-associated HCC, chronic kidney injury, and heart failure., Supplemental Graphical Summary

Chronic liver injury with any etiology can progress to fibrosis and the end-stage diseases cirrhosis and hepatocellular carcinoma. The progression of liver disease is controlled by a variety of factors, including liver injury, inflammatory cells, inflammatory mediators, cytokines, and the gut microbiome. In the current review, we discuss recent data on a large number of cytokines that play important roles in regulating liver injury, inflammation, fibrosis, and regeneration, with a focus on interferons and T helper (Th) 1, Th2, Th9, Th17, interleukin (IL)-1 family, IL-6 family, and IL-20 family cytokines. Hepatocytes can also produce certain cytokines (such as IL-7, IL-11, and IL-33), and the functions of these cytokines in the liver are briefly summarized. Several cytokines have great therapeutic potential, and some are currently being tested as therapeutic targets in clinical trials for the treatment of liver diseases, which are also described.

Benign hepatosteatosis, affected by lipid uptake, de novo lipogenesis and fatty acid (FA) oxidation, progresses to non-alcoholic steatohepatitis (NASH) on stress and inflammation. A key macronutrient proposed to increase hepatosteatosis and NASH risk is fructose. Excessive intake of fructose causes intestinal-barrier deterioration and endotoxaemia. However, how fructose triggers these alterations and their roles in hepatosteatosis and NASH pathogenesis remain unknown. Here we show, using mice, that microbiota-derived Toll-like receptor (TLR) agonists promote hepatosteatosis without affecting fructose-1-phosphate (F1P) and cytosolic acetyl-CoA. Activation of mucosal-regenerative gp130 signalling, administration of the YAP-induced matricellular protein CCN1 or expression of the antimicrobial peptide Reg3b (beta) peptide counteract fructose-induced barrier deterioration, which depends on endoplasmic-reticulum stress and subsequent endotoxaemia. Endotoxin engages TLR4 to trigger TNF production by liver macrophages, thereby inducing lipogenic enzymes that convert F1P and acetyl-CoA to FA in both mouse and human hepatocytes.

Background & Aims Chronic alcohol consumption is a leading risk factor for the development of hepatocellular carcinoma (HCC), which is associated with a marked increase in hepatic expression of pro-inflammatory IL-17A and its receptor IL-17RA. Methods Genetic deletion and pharmacological blocking were used to characterize the role of IL-17A/IL-17RA signaling in the pathogenesis of HCC in mouse models and human specimens. Results We demonstrate that the global deletion of the Il-17ra gene suppressed HCC in alcohol-fed diethylnitrosamine-challenged Il-17ra–/– and major urinary protein-urokinase-type plasminogen activator/Il-17ra–/– mice compared with wild-type mice. When the cell-specific role of IL-17RA signaling was examined, the development of HCC was decreased in both alcohol-fed Il-17raΔMΦ and Il-17raΔHep mice devoid of IL-17RA in myeloid cells and hepatocytes, but not in Il-17raΔHSC mice (deficient in IL-17RA in hepatic stellate cells). Deletion of Il-17ra in myeloid cells ameliorated tumorigenesis via suppression of pro-tumorigenic/inflammatory and pro-fibrogenic responses in alcohol-fed Il-17raΔMΦ mice. Remarkably, despite a normal inflammatory response, alcohol-fed Il-17raΔHep mice developed the fewest tumors (compared with Il-17raΔMΦ mice), with reduced steatosis and fibrosis. Steatotic IL-17RA-deficient hepatocytes downregulated the expression of Cxcl1 and other chemokines, exhibited a striking defect in tumor necrosis factor (TNF)/TNF receptor 1-dependent caspase-2-SREBP1/2-DHCR7-mediated cholesterol synthesis, and upregulated the production of antioxidant vitamin D3. The pharmacological blocking of IL-17A/Th-17 cells using anti-IL-12/IL-23 antibodies suppressed the progression of HCC (by 70%) in alcohol-fed mice, indicating that targeting IL-17 signaling might provide novel strategies for the treatment of alcohol-induced HCC. Conclusions Overall, IL-17A is a tumor-promoting cytokine, which critically regulates alcohol-induced hepatic steatosis, inflammation, fibrosis, and HCC. Lay summary IL-17A is a tumor-promoting cytokine, which critically regulates inflammatory responses in macrophages (Kupffer cells and bone-marrow-derived monocytes) and cholesterol synthesis in steatotic hepatocytes in an experimental model of alcohol-induced HCC. Therefore, IL-17A may be a potential therapeutic target for patients with alcohol-induced HCC.

Alcoholic liver disease (ALD) is a leading cause of cirrhosis in the United States, which is characterized by extensive deposition of extracellular matrix proteins and formation of a fibrous scar. Hepatic stellate cells (HSCs) are the major source of collagen type 1 producing myofibroblasts in ALD fibrosis. However, the mechanism of alcohol‐induced activation of human and mouse HSCs is not fully understood. We compared the gene‐expression profiles of primary cultured human HSCs (hHSCs) isolated from patients with ALD (n = 3) or without underlying liver disease (n = 4) using RNA‐sequencing analysis. Furthermore, the gene‐expression profile of ALD hHSCs was compared with that of alcohol‐activated mHSCs (isolated from intragastric alcohol‐fed mice) or CCl4‐activated mouse HSCs (mHSCs). Comparative transcriptome analysis revealed that ALD hHSCs, in addition to alcohol‐activated and CCl4‐activated mHSCs, share the expression of common HSC activation (Col1a1 [collagen type I alpha 1 chain], Acta1 [actin alpha 1, skeletal muscle], PAI1 [plasminogen activator inhibitor‐1], TIMP1 [tissue inhibitor of metalloproteinase 1], and LOXL2 [lysyl oxidase homolog 2]), indicating that a common mechanism underlies the activation of human and mouse HSCs. Furthermore, alcohol‐activated mHSCs most closely recapitulate the gene‐expression profile of ALD hHSCs. We identified the genes that are similarly and uniquely up‐regulated in primary cultured alcohol‐activated hHSCs and freshly isolated mHSCs, which include CSF1R (macrophage colony‐stimulating factor 1 receptor), PLEK (pleckstrin), LAPTM5 (lysosmal‐associated transmembrane protein 5), CD74 (class I transactivator, the invariant chain), CD53, MMP9 (matrix metallopeptidase 9), CD14, CTSS (cathepsin S), TYROBP (TYRO protein tyrosine kinase‐binding protein), and ITGB2 (integrin beta‐2), and other genes (compared with CCl4‐activated mHSCs). Conclusion: We identified genes in alcohol‐activated mHSCs from intragastric alcohol‐fed mice that are largely consistent with the gene‐expression profile of primary cultured hHSCs from patients with ALD. These genes are unique to alcohol‐induced HSC activation in two species, and therefore may become targets or readout for antifibrotic therapy in experimental models of ALD., We identified genes in alcohol‐activated mHSCs from IG alcohol‐fed mice that are largely consistent with the gene‐expression profile of primary cultured hHSCs from patients with ALD. These genes are unique to alcohol‐induced HSC activation in two species, and therefore may become targets or readout for antifibrotic therapy in experimental models of ALD.

Background & AimsThe extrahepatic bile duct is the primary tissue initially affected by the cholangiopathy biliary atresia. Biliary atresia affects neonates exclusively and current animal models suggest that the developing bile duct is uniquely susceptible to damage. In this study, we aimed to define the anatomical and functional differences between the neonatal and adult mouse extrahepatic bile ducts.MethodsWe studied mouse passaged cholangiocytes, mouse BALB/c neonatal and adult primary cholangiocytes and isolated extrahepatic bile ducts, and a collagen reporter mouse. Methods included transmission electron microscopy, lectin staining, immunostaining, rhodamine uptake assays, bile acid toxicity assays, and in vitro modeling of the matrix.ResultsThe cholangiocyte monolayer of the neonatal extrahepatic bile duct was immature, lacking the uniform apical glycocalyx and mature cell-cell junctions typical of adult cholangiocytes. Functional studies showed that the glycocalyx protected against bile acid injury and that neonatal cholangiocyte monolayers were more permeable than adult monolayers. In adult ducts, the submucosal space was filled with collagen I, elastin, hyaluronic acid, and proteoglycans. In contrast, the neonatal submucosa had little collagen I and elastin, although both increased rapidly after birth. In vitro modeling suggested that the composition of the neonatal submucosa relative to the adult submucosa led to increased diffusion of bile. A Col-GFP reporter mouse showed that cells in the neonatal but not adult submucosa were actively producing collagen.ConclusionWe identified four key differences between the neonatal and adult extrahepatic bile duct. We showed that these features may have functional implications, suggesting the neonatal extrahepatic bile ducts are particularly susceptible to injury and fibrosis.Lay SummaryBiliary atresia is a disease that affects newborns and is characterized by extrahepatic bile duct injury and obstruction with resulting liver injury. We identify four key differences between the epithelial and submucosal layers of the neonatal and adult extrahepatic bile duct and show that these may render the neonatal duct particularly susceptible to injury.HighlightsThe apical glycocalyx is thin and patchy in neonatal compared to adult cholangiocytesNeonatal cholangiocytes have immature cell-cell junctions and increased permeabilityThe neonatal submucosal space has minimal collagen I or elastinThe neonatal submucosal space contains many actively collagen-secreting cellsGraphical abstract

We investigated the role of mesothelin (Msln) and thymocyte differentiation antigen 1 (Thy1) in the activation of fibroblasts across multiple organs and demonstrated that Msln(−/−) mice are protected from cholestatic fibrosis caused by Mdr2 (multidrug resistance gene 2) deficiency, bleomycin-induced lung fibrosis, and UUO (unilateral urinary obstruction)-induced kidney fibrosis. On the contrary, Thy1(−/−) mice are more susceptible to fibrosis, suggesting that a Msln–Thy1 signaling complex is critical for tissue fibroblast activation. A similar mechanism was observed in human activated portal fibroblasts (aPFs). Targeting of human MSLN(+) aPFs with two anti-MSLN immunotoxins killed fibroblasts engineered to express human mesothelin and reduced collagen deposition in livers of bile duct ligation (BDL)–injured mice. We provide evidence that antimesothelin-based therapy may be a strategy for treatment of parenchymal organ fibrosis.

We investigated the role of mesothelin (Msln) and thymocyte differentiation antigen 1 (Thy1) in the activation of fibroblasts across multiple organs and demonstrated that Msln-/- mice are protected from cholestatic fibrosis caused by Mdr2 (multidrug resistance gene 2) deficiency, bleomycin-induced lung fibrosis, and UUO (unilateral urinary obstruction)-induced kidney fibrosis. On the contrary, Thy1-/- mice are more susceptible to fibrosis, suggesting that a Msln-Thy1 signaling complex is critical for tissue fibroblast activation. A similar mechanism was observed in human activated portal fibroblasts (aPFs). Targeting of human MSLN+ aPFs with two anti-MSLN immunotoxins killed fibroblasts engineered to express human mesothelin and reduced collagen deposition in livers of bile duct ligation (BDL)-injured mice. We provide evidence that antimesothelin-based therapy may be a strategy for treatment of parenchymal organ fibrosis.

Interleukin 17A (IL-17A)-producing T helper 17 (Th17) cells were identified as a subset of T helper cells that play a critical role in host defense against bacterial and fungal pathogens. Th17 cells differentiate from Th0 naïve T-cells in response to transforming growth factor β1 (TGF-β1) and IL-6, the cytokines which also drive development of liver fibrosis, require activation of transcription factor retinoic acid receptor-related orphan nuclear receptor gamma t (RORγt). IL-17A signals through the ubiquitously expressed receptor IL-17RA. Expression of IL-17RA is upregulated in patients with hepatitis B virus/hepatitis C virus (HBV/HCV) infections, nonalcoholic steatohepatitis (NASH), alcohol-associated liver disease (AALD), hepatocellular carcinoma (HCC), and experimental models of chronic toxic liver injury. The role of IL-17 signaling in the pathogenesis of NASH- and AALD-induced metabolic liver injury and HCC will be the focus of this review. The role of IL-17A–IL-17RA axis in mediation of the cross-talk between metabolically injured hepatic macrophages, hepatocytes, and fibrogenic myofibroblasts will be discussed.

Hepatocytes have important roles in liver iron homeostasis, abnormalities in which are tightly associated with liver steatosis and fibrosis. Here, we show that non-alcoholic fatty liver disease (NAFLD) and steatohepatitis (NASH) are characterized by iron-deficient hepatocytes and iron overload in hepatic stellate cells (HSCs). Iron deficiency enhances hepatocyte lipogenesis and insulin resistance through HIF2α-ATF4 signaling. Elevated secretion of iron-containing hepatocyte extracellular vesicles (EVs), which are normally cleared by Kupffer cells, accounts for hepatocyte iron deficiency and HSC iron overload in NAFLD/NASH livers. Iron accumulation results in overproduction of reactive oxygen species that promote HSC fibrogenic activation. Conversely, blocking hepatocyte EV secretion or depleting EV iron cargo restores liver iron homeostasis, concomitant with mitigation of NAFLD/NASH-associated liver steatosis and fibrosis. Taken together, these studies show that iron distribution disorders contribute to the development of liver metabolic diseases.

Peroxisome proliferator-activated receptor delta (PPARδ) agonists have been shown to exert beneficial effects in liver disease and reduce total bile acid levels. The mechanism(s) whereby PPARδ agonism reduces bile acid levels are, however, unknown, and therefore the aim of the present study was to investigate the molecular pathways responsible for reducing bile acid synthesis in hepatocytes, following treatment with the selective PPARδ agonist, seladelpar. We show that administration of seladelpar to WT mice repressed the liver expression of cholesterol 7 alpha-hydroxylase (Cyp7a1), the rate-limiting enzyme for bile acid synthesis, and decreased plasma 7α-hydroxy-4-cholesten-3-one (C4), a freely diffusible metabolite downstream of Cyp7a1. In primary mouse hepatocytes, seladelpar significantly reduced the expression of Cyp7a1 independent of the nuclear bile acid receptor, Farnesoid X receptor. In addition, seladelpar upregulated fibroblast growth factor 21 (Fgf21) in mouse liver, serum, and in cultured hepatocytes. We demonstrate that recombinant Fgf21 protein activated the c-Jun N-terminal kinase (JNK) signaling pathway and repressed Cyp7a1 gene expression in primary hepatocytes. The suppressive effect of seladelpar on Cyp7a1 expression was blocked by a JNK inhibitor as well as in the absence of Fgf21, indicating that Fgf21 plays an indispensable role in PPARδ-mediated downregulation of Cyp7a1. Finally, reduction of CYP7A1 expression by seladelpar was confirmed in primary human hepatocytes. In conclusion, we show that seladelpar reduces bile acid synthesis via an FGF21-dependent mechanism that signals at least partially through JNK to repress CYP7A1.

Nonalcoholic steatohepatitis (NASH) is a liver disease associated with significant morbidity. Kupffer cells (KCs) produce endogenous miR-690 and, via exosome secretion, shuttle this miRNA to other liver cells, such as hepatocytes, recruited hepatic macrophages (RHMs), and hepatic stellate cells (HSCs). miR-690 directly inhibits fibrogenesis in HSCs, inflammation in RHMs, and de novo lipogenesis in hepatocytes. When an miR-690 mimic is administered to NASH mice in vivo, all the features of the NASH phenotype are robustly inhibited. During the development of NASH, KCs become miR-690 deficient, and miR-690 levels are markedly lower in mouse and human NASH livers than in controls. KC-specific KO of miR-690 promotes NASH pathogenesis. A primary target of miR-690 is NADK mRNA, and NADK levels are inversely proportional to the cellular miR-690 content. These studies show that KCs play a central role in the etiology of NASH and raise the possibility that miR-690 could emerge as a therapeutic for this condition.

Hepatic fibrosis results from chronic injury and inflammation in the liver and leads to cirrhosis, liver failure, and portal hypertension. Understanding the molecular mechanisms underlying hepatic fibrosis has advanced the prospect of developing therapies for regression of the disease. Resolution of fibrosis requires a reduction of proinflammatory and fibrogenic cytokines, a decrease in extracellular matrix (ECM) protein production, an increase in collagenase activity, and finally, a disappearance of activated myofibroblasts. Exosomes are nanovesicles of endocytic origin secreted by most cell types. They epigenetically reprogram and alter the phenotype of their recipient cells and hold great promise for the reversal of fibrosis. Recent studies have shown that exosomes function as conduits for intercellular transfer and contain all the necessary components to induce resolution of fibrosis, including the ability to (1) inhibit macrophage activation and cytokine secretion, (2) remodel ECM production and decrease fibrous scars, and (3) inactivate hepatic stellate cells, a major myofibroblast population. Here, we discuss the research involving the regression of hepatic fibrosis. We focus on the newly discovered roles of exosomes during fibrogenesis and as a therapy for fibrosis reversal. We also emphasize the novel discoveries of exosome-based antifibrotic treatments in vitro and in vivo.

With an increasing number of patients with degenerative hepatic diseases, such as liver fibrosis, and a limited supply of donor organs, there is an unmet need for therapies that can repair or regenerate damaged liver tissue. Treatment with macrophages that are capable of phagocytosis and anti-inflammatory activities such as secretion of matrix metalloproteinases (MMPs) provide an attractive cellular therapy approach. Human induced pluripotent stem cells (iPSCs) are capable of efficiently generating a large-scale, homogenous population of human macrophages using fully defined feeder- and serum-free differentiation protocol. Human iPSC-macrophages exhibit classical surface cell markers and phagocytic activity similar to peripheral blood-derived macrophages. Moreover, gene and cytokine expression analysis reveal that these macrophages can be efficiently polarized to pro-inflammatory M1 or anti-inflammatory M2 phenotypes in presence of LPS + IFN-γ and IL-4 + IL-13, respectively. M1 macrophages express high level of CD80, TNF-α, and IL-6 while M2 macrophages show elevated expression of CD206, CCL17, and CCL22. Here, we demonstrate that treatment of liver fibrosis with both human iPSC-derived macrophage populations and especially M2 subtype significantly reduces fibrogenic gene expression and disease associated histological markers including Sirius Red, αSMA and desmin in immunodeficient Rag2−/−γc−/− mice model, making this approach a promising cell-based avenue to ameliorate fibrosis.

Cholangiopathies caused by biliary epithelial cell (BEC) injury represent a leading cause of liver failure. No effective pharmacologic therapies exist, and the underlying mechanisms remain obscure. We aimed to explore the mechanisms of bile duct repair after targeted BEC injury. Injection of intermedilysin into BEC-specific human CD59 (hCD59) transgenic mice induced acute and specific BEC death, representing a model to study the early signals that drive bile duct repair. Acute BEC injury induced cholestasis followed by CCR2(+) monocyte recruitment and BEC proliferation. Using microdissection and next-generation RNA-Seq, we identified 5 genes, including Mapk8ip2, Cdkn1a, Itgb6, Rgs4, and Ccl2, that were most upregulated in proliferating BECs after acute injury. Immunohistochemical analyses confirmed robust upregulation of integrin αvβ6 (ITGβ6) expression in this BEC injury model, after bile duct ligation, and in patients with chronic cholangiopathies. Deletion of the Itgb6 gene attenuated BEC proliferation after acute bile duct injury. Macrophage depletion or Ccr2 deficiency impaired ITGβ6 expression and BEC proliferation. In vitro experiments revealed that bile acid–activated monocytes promoted BEC proliferation through ITGβ6. Our data suggest that BEC injury induces cholestasis, monocyte recruitment, and induction of ITGβ6, which work together to promote BEC proliferation and therefore represent potential therapeutic targets for cholangiopathies.

Although hepatocellular cancer (HCC) usually occurs in the setting of liver fibrosis, the causal relationship between liver fibrosis and HCC is unclear. in vivo and in vitro models of HCC involving Col(r/r) mice (that produce a collagenase-resistant type I collagen) or wild-type (WT) mice were used to assess the relationship between type I collagen, liver fibrosis, and experimental HCC. HCC was either chemically induced in WT and Col(r/r) mice or Hepa 1-6 cells were engrafted into WT and Col(r/r) livers. The effect of hepatic stellate cells (HSCs) from WT and Col(r/r) mice on the growth of Hepa 1-6 cells was studied by using multicellular tumor spheroids and xenografts. Collagen type I deposition and fibrosis were increased in Col(r/r) mice, but they developed fewer and smaller tumors. Hepa 1-6 cells had reduced tumor growth in the livers of Col(r/r) mice. Although Col(r/r) HSCs exhibited a more activated phenotype, Hepa 1-6 growth and malignancy were suppressed in multicellular tumor spheroids and in xenografts containing Col(r/r) HSCs. Treatment with vitronectin, which mimics the presence of degraded collagen fragments, converted the Col(r/r) phenotype into a WT phenotype. Although Col(r/r) mice have increased liver fibrosis, they exhibited decreased HCC in several models. Thus, increased liver type I collagen does not produce increased experimental HCC.

Liver fibrosis is a clinically significant finding that has major impacts on patient morbidity and mortality. The mechanism of fibrosis involves many different cellular pathways, but the major cell type involved appears to be hepatic stellate cells. Many liver diseases, including Hepatitis B, C, and fatty liver disease cause ongoing hepatocellular damage leading to liver fibrosis. No matter the cause of liver disease, liver-related mortality increases exponentially with increasing fibrosis. The progression to cirrhosis brings more dramatic mortality and higher incidence of hepatocellular carcinoma. Fibrosis can also affect outcomes following liver transplantation in adult and pediatric patients and require retransplantation. Drugs exist to treat Hepatitis B and C that reverse fibrosis in patients with those viral diseases, but there are currently no therapies to directly treat liver fibrosis. Several mouse models of chronic liver diseases have been successfully reversed using novel drug targets with current therapies focusing mostly on prevention of myofibroblast activation. Further research in these areas could lead to development of drugs to treat fibrosis, which will have invaluable impact on patient survival. This article is categorized under: Metabolic Diseases > Molecular and Cellular Physiology.