Macrophages in necrotic liver lesion repair: opportunities for therapeutical applications.

Healthy livers contain 80% of body resident macrophages known as Kupffer cells. In diseased livers, the number of Kupffer cells usually drops but is compensated by infiltration of monocyte-derived macrophages, some of which can differentiate into Kupffer-like cells. Early studies suggest that Kupffer cells play important roles in both promoting liver injury and liver regeneration. Yet, the distinction between the functionalities of resident and infiltrating macrophages is not always made. By using more specific macrophage markers and targeted cell depletion and single-cell RNA sequencing, recent studies revealed several subsets of monocyte-derived macrophages that play important functions in inducing liver damage and inflammation as well as in liver repair and regeneration. In this review, we discuss the different roles that hepatic macrophages play in promoting necrotic liver lesion resolution and dead cell clearance, as well as the targeting of these cells as potential tools for the development of novel therapies for acute liver failure and acute-on-chronic liver failure.

Role of Macrophages in Liver Fibrosis.

Liver fibrosis, which ultimately leads to liver cirrhosis and hepatocellular carcinoma, is a major health burden worldwide. The progression of liver fibrosis is the result of the wound-healing response of liver to repeated injury. Hepatic macrophages are cells with high heterogeneity and plasticity and include tissue-resident macrophages termed Kupffer cells, and recruited macrophages derived from circulating monocytes, spleen and peritoneal cavity. Studies have shown that hepatic macrophages play roles in the initiation and progression of liver fibrosis by releasing inflammatory cytokines/chemokines and pro-fibrogenic factors. Furthermore, the development of liver fibrosis has been shown to be reversible. Hepatic macrophages have been shown to alternately regulate both the regression and turnover of liver fibrosis by changing their phenotypes during the dynamic progression of liver fibrosis. In this review, we summarize the role of hepatic macrophages in the progression and regression of liver fibrosis.

Dynamics of cellular plasticity in non-alcoholic steatohepatitis (NASH).

Non-alcoholic steatohepatitis (NASH) is a pathogenic stage of the broader non-alcoholic fatty liver disease (NAFLD). Histological presentation of NASH includes hepatocyte ballooning, macrophage polarization, ductular reaction, and hepatic stellate cell (HSCs) activation. At a cellular level, a heterogenous population of cells such as hepatocytes, macrophages, cholangiocytes, and HSCs undergo dramatic intra-cellular changes in response to extracellular triggers, which are termed "cellular plasticity. This dynamic switch in the cellular structure and function of hepatic parenchymal and non-parenchymal cells and their crosstalk culminates in the perpetuation of inflammation and fibrosis in NASH. This review presents an overview of our current understanding of cellular plasticity in NASH and its molecular mechanisms, along with possible targeting to develop cell-specific NASH therapies.

Phenotypic characterization of liver tissue heterogeneity through a next-generation 3D single-cell atlas.

Three-dimensional (3D) geometrical models are potent tools for quantifying complex tissue features and exploring structure-function relationships. However, these models are generally incomplete due to experimental limitations in acquiring multiple (> 4) fluorescent channels in thick tissue sections simultaneously. Indeed, predictive geometrical and functional models of the liver have been restricted to few tissue and cellular components, excluding important cellular populations such as hepatic stellate cells (HSCs) and Kupffer cells (KCs). Here, we combined deep-tissue immunostaining, multiphoton microscopy, deep-learning techniques, and 3D image processing to computationally expand the number of simultaneously reconstructed tissue structures. We then generated a spatial single-cell atlas of hepatic architecture (Hep3D), including all main tissue and cellular components at different stages of post-natal development in mice. We used Hep3D to quantitatively study 1) hepatic morphodynamics from early post-natal development to adulthood, and 2) the effect on the liver's overall structure when changing the hepatic environment after removing KCs. In addition to a complete description of bile canaliculi and sinusoidal network remodeling, our analysis uncovered unexpected spatiotemporal patterns of non-parenchymal cells and hepatocytes differing in size, number of nuclei, and DNA content. Surprisingly, we found that the specific depletion of KCs results in morphological changes in hepatocytes and HSCs. These findings reveal novel characteristics of liver heterogeneity and have important implications for both the structural organization of liver tissue and its function. Our next-gen 3D single-cell atlas is a powerful tool to understand liver tissue architecture, opening up avenues for in-depth investigations into tissue structure across both normal and pathological conditions.

The pivotal role of dysregulated autophagy in the progression of non-alcoholic fatty liver disease.

Non-alcoholic fatty liver disease (NAFLD) is a clinicopathologic syndrome characterized by excessive fat deposition in hepatocytes and a major cause of end-stage liver disease. Autophagy is a metabolic pathway responsible for degrading cytoplasmic products and damaged organelles, playing a pivotal role in maintaining the homeostasis and functionality of hepatocytes. Recent studies have shown that pharmacological intervention to activate or restore autophagy provides benefits for liver function recovery by promoting the clearance of lipid droplets (LDs) in hepatocytes, decreasing the production of pro-inflammatory factors, and inhibiting activated hepatic stellate cells (HSCs), thus improving liver fibrosis and slowing down the progression of NAFLD. This article summarizes the physiological process of autophagy, elucidates the close relationship between NAFLD and autophagy, and discusses the effects of drugs on autophagy and signaling pathways from the perspectives of hepatocytes, kupffer cells (KCs), and HSCs to provide assistance in the clinical management of NAFLD.

CX3CR1 deficiency promotes resolution of hepatic ischemia-reperfusion injury by regulating homeostatic function of liver infiltrating macrophages.

Hepatic ischemia-reperfusion injury(HIRI) remains to be an unsolved risk factor that contributes to organ failure after liver surgery. Our clinical retrospective study showed that lower donor liver CX3-C chemokine receptor-1(CX3CR1) mRNA expression level were correlated with upregulated pro-resolved macrophage receptor MERTK, as well as promoted restoration efficiency of allograft injury in liver transplant. To further characterize roles of CX3CR1 in regulating resolution of HIRI, we employed murine liver partial warm ischemia-reperfusion model by Wt & Cx3cr1-/- mice and the reperfusion time was prolonged from 6 h to 4-7 days. Kupffer cells(KCs) were depleted by clodronate liposome(CL) in advance to focus on infiltrating macrophages, and repopulation kinetics were determined by FACS, IF and RNA-Seq. CX3CR1 antagonist AZD8797 was injected i.p. to interrogate potential pharmacological therapeutic strategies. In vitro primary bone marrow macrophages(BMMs) culture by LXR agonist DMHCA, as well as molecular and functional studies, were undertaken to dissect roles of CX3CR1 in modulating macrophages cytobiological development and resolutive functions. We observed that deficiency or pharmacological inhibition of CX3CR1 facilitated HIRI resolution via promoted macrophages migration in CCR1/CCR5 manner, as well as enhanced MerTK-mediated efferocytosis. Our study demonstrated the critical roles of CX3CR1 in progression of HIRI and identified it as a potential therapeutic target in clinical liver transplantation.

Loss of Toll-like receptor 9 protects from hepatocellular carcinoma in murine models of chronic liver disease.

BACKGROUND & AIMS: Toll-like receptor 9 (Tlr9) is a pathogen recognition receptor detecting unmethylated DNA derivatives of pathogens and damaged host cells. It is therefore an important modulator of innate immunity. Here we investigated the role of Tlr9 in fibrogenesis and progression of hepatocellular carcinoma in chronic liver disease. MATERIALS AND METHODS: We treated mice with a constitutive deletion of Tlr9 (Tlr9-/-) with DEN/CCl4 for 24 weeks. As a second model, we used hepatocyte-specific Nemo knockout (NemoΔhepa) mice and generated double knockout (NemoΔhepaTlr9-/-) animals. RESULTS: We show that Tlr9 is in the liver primarily expressed in Kupffer cells, suggesting a key role of Tlr9 in intercellular communication during hepatic injury. Tlr9 deletion resulted in reduced liver fibrosis as well as tumor burden. We observed down-regulation of hepatic stellate cell activation and consequently decreased collagen production in both models. Tlr9 deletion was associated with decreased apoptosis and compensatory proliferation of hepatocytes, modulating the initiation and progression of hepatocarcinogenesis. These findings were accompanied by a decrease in interferon-ß and an increase in chemokines having an anti-tumoral effect. CONCLUSIONS: Our data define Tlr9 as an important receptor involved in fibrogenesis, but also in the initiation and progression of hepatocellular carcinoma during chronic liver diseases.

Concomitant NAFLD Facilitates Liver Metastases and PD-1-Refractory by Recruiting MDSCs via CXCL5/CXCR2 in Colorectal Cancer.

BACKGROUND & AIMS: Both nonalcoholic fatty liver disease (NAFLD) and colorectal cancer (CRC) are prevalent worldwide. The effects of concomitant NAFLD on the risk of colorectal liver metastasis (CRLM) and its mechanisms have not been definitively elucidated. METHODS: We observed the effect of concomitant NAFLD on CRLM in the mouse model and explored the underlying mechanisms of specific myeloid-derived suppressor cells (MDSCs) recruitment and then tested the therapeutic application based on the mechanisms. Finally we validated our findings in the clinical samples. RESULTS: Here we prove that in different mouse models, NAFLD induces F4/80+ Kupffer cells to secret chemokine CXCL5 and then recruits CXCR2+ MDSCs to promote the growth of CRLM. CRLM with NAFLD background is refractory to the anti-PD-1 monoclonal antibody treatment, but when combined with Reparixin, an inhibitor of CXCR1/2, dual therapy cures the established CRLM in mice with NAFLD. Our clinical studies also indicate that fatty liver diseases increase the infiltration of CXCR2+ MDSCs, as well as the hazard of liver metastases in CRC patients. CONCLUSIONS: Collectively, our findings highlight the significance of selective CXCR2+/CD11b+/Gr-1+ subset myeloid cells in favoring the development of CRLM with NAFLD background and identify a pharmaceutical medicine that is already available for the clinical trials and potential treatment.

6-Formylindolo[3,2-b]carbazole, a potent ligand for the aryl hydrocarbon receptor, attenuates concanavalin-induced hepatitis by limiting T-cell activation and infiltration of proinflammatory CD11b+ Kupffer cells.

FICZ (6-formylindolo[3,2-b]carbazole) is a potent aryl hydrocarbon receptor agonist that has a poorly understood function in the regulation of inflammation. In this study, we investigated the effect of aryl hydrocarbon receptor activation by FICZ in a murine model of autoimmune hepatitis induced by concanavalin A. High-throughput sequencing techniques such as single-cell RNA sequencing and assay for transposase accessible chromatin sequencing were used to explore the mechanisms through which FICZ induces its effects. FICZ treatment attenuated concanavalin A-induced hepatitis, evidenced by decreased T-cell infiltration, decreased circulating alanine transaminase levels, and suppression of proinflammatory cytokines. Concanavalin A revealed an increase in natural killer T cells, T cells, and mature B cells upon concanavalin A injection while FICZ treatment reversed the presence of these subsets. Surprisingly, concanavalin A depleted a subset of CD55+ B cells, while FICZ partially protected this subset. The immune cells showed significant dysregulation in the gene expression profiles, including diverse expression of migratory markers such as CCL4, CCL5, and CXCL2 and critical regulatory markers such as Junb. Assay for transposase accessible chromatin sequencing showed more accessible chromatin in the CD3e promoter in the concanavalin A-only group as compared to the naive and concanavalin A-exposed, FICZ-treated group. While there was overall more accessible chromatin of the Adgre1 (F4/80) promoter in the FICZ-treated group, we observed less open chromatin in the Itgam (CD11b) promoter in Kupffer cells, supporting the ability of FICZ to reduce the infiltration of proinflammatory cytokine producing CD11b+ Kupffer cells. Taken together, these data demonstrate that aryl hydrocarbon receptor activation by FICZ suppresses liver injury through the limitation of CD3+ T-cell activation and CD11b+ Kupffer cell infiltration.

O papel da oxigenação hiperbárica na estrutura do fígado e baço após ligadura das veias hepáticas: estudo em ratos / The role of hyperbaric oxygenation in the liver and spleen structure after hepatic vein ligation: study in rats

OBJETIVO: Avaliação morfológica do fígado e baço de ratos submetidos à oxigenoterapia hiperbárica após a ligadura das veias hepáticas. MÉTODO: Foram utilizados 30 animais machos adultos da espécie Holtzman, distribuídos aleatoriamente em dois grupos de 15 animais cada, assim designados: grupo 1 - ligadura das veias hepáticas; grupo 2 - ligadura das veias hepáticas associada à oxigenoterapia hiperbárica. Todos os animais foram submetidos à anestesia geral por meio de solução contendo cloridrato de cetamina (40 mg/ml) e cloridrato de meperidina (10 mg/ml) na dose de 50 mg/kg/peso, laparotomia mediana e ligadura das veias hepáticas. A oxigenoterapia hiperbárica foi aplicada nos animais do grupo 2, a partir da oitava hora do pós-operatório, por 120 minutos, sendo 90 minutos sob pressão de 2,5 atmosferas e 15 minutos no início e final da terapêutica, para promover a compressão e descompressão gradativa no período de 20 dias consecutivos. No 21° dia de pós-operatório, os animais foram mortos por inalação de éter e submetidos à laparotomia e extirpação dos fígados e baços para exame histológico. Foram comparados os resultados da histologia hepática e esplênica aplicando-se o teste exato de Fisher, considerando-se a diferença significante de P < 0,05. RESULTADOS: Os exames histológicos dos fígados e baços dos animais dos grupos 1 e 2 mostraram as seguintes alterações: presença de trombose nas veias hepática, porta e centro-lobular em cinco (33,3 por cento) animais do grupo 1 e ausência no grupo 2; presença de necrose dos hepatócitos caracterizada como acentuada em sete animais (46,7 por cento) e leve em oito (53,3 por cento) animais do grupo 1, enquanto que, em todos os animais do grupo 2, esta alteração foi caracterizada como leve; presença de células de Kupffer muito proeminentes e hipertrofiadas em 14 (93,3 por cento) animais do grupo 1 e pouco proeminentes e hipertrofiadas em todos os animais do grupo 2; congestão da polpa vermelha considerada acentuada em seis (40 por cento) e moderada em nove (60 por cento) animais do grupo 1 e em todos os animais do grupo 2; hemossiderose moderada ou acentuada em 14 (93,3 por cento) animais do grupo 1 e leve em todos os animais do grupo 2. As análises estatísticas realizadas entre os dois grupos mostraram diferenças significativas em todas a variáveis estudadas (P < 0,05)...

El hígado como órgano inmunológico / The liver and immune organ

La prevalencia de enfermedades hepáticas a nivel mundial registra cifras alarmantes. Solo la infección por malaria afecta a 500 millones de personas por año a nivel mundial. Enfermedades de etiología viral como hepatitis B y C, contribuyen al aumento de la casuística y por su caracter de patologías de tipo crónico evolucionan a formas severas como la fibrosis o los procesos neoplásicos. La relevancia del hígado como órgano central en la maquinaria metabólica del organismo y como clave partícipe de la respuesta inflamatoria sistémica, indican la necesidad de preservar sus capacidades funcionales.