Luteolin-7-diglucuronide, a novel PTP1B inhibitor, ameliorates hepatic stellate cell activation and liver fibrosis in mice.

Liver fibrosis, one of the leading causes of morbidity and mortality worldwide, lacks effective therapy. The activation of hepatic stellate cells (HSCs) is the dominant event in hepatic fibrogenesis. Luteolin-7-diglucuronide (L7DG) is the major flavonoid extracted from Perilla frutescens and Verbena officinalis. Their beneficial effects in the treatment of liver diseases were well documented. In this study we investigated the anti-fibrotic activities of L7DG and the potential mechanisms. We established TGF-ß1-activated mouse primary hepatic stellate cells (pHSCs) and human HSC line LX-2 as in vitro liver fibrosis models. Co-treatment with L7DG (5, 20, 50 µM) dose-dependently decreased TGF-ß1-induced expression of fibrotic markers collagen 1, α-SMA and fibronectin. In liver fibrosis mouse models induced by CCl4 challenge alone or in combination with HFHC diet, administration of L7DG (40, 150 mg·kg-1·d-1, i.g., for 4 or 8 weeks) dose-dependently attenuated hepatic histopathological injury and collagen accumulation, decreased expression of fibrogenic genes. By conducting target prediction, molecular docking and enzyme activity detection, we identified L7DG as a potent inhibitor of protein tyrosine phosphatase 1B (PTP1B) with an IC50 value of 2.10 µM. Further studies revealed that L7DG inhibited PTP1B activity, up-regulated AMPK phosphorylation and subsequently inhibited HSC activation. This study demonstrates that the phytochemical L7DG may be a potential therapeutic candidate for the treatment of liver fibrosis.

Sequential system based on ferritin delivery system and cell therapy for modulating the pathological microenvironment and promoting recovery.

The vicious crosstalk among capillarization of hepatic sinusoidal endothelial cells (LSECs), activation of hepatic stellate cells (aHSCs), and hepatocyte damage poses a significant impediment to the successful treatment of liver fibrosis. In this study, we propose a sequential combination therapy aimed at disrupting the malignant crosstalk and reshaping the benign microenvironment while repairing damaged hepatocytes to achieve effective treatment of liver fibrosis. Firstly, H-subunit apoferrin (Ferritin) was adopted to load platycodonin D (PLD) and MnO2, forming ferritin@MnO2/PLD (FMP) nanoparticles, which exploited the high affinity of ferritin for the highly expressed transferrin receptor 1 (TfR1) to achieve the precise targeted delivery of FMP in the liver. Upon PLD intervention, restoration of the fenestration pores in capillarized LSECs was facilitated by modulating the phosphatidyl inositol 3-kinase/protein kinase B (PI3K/AKT) and Kruppel Like Factor 2 (KLF2) signaling pathways both in vitro and in vivo, enabling efficient entry of FMP into the Disse space. Subsequently, FMP NPs effectively inhibited HSC activation by modulating the TLR2/TLR4/NF-κB-p65 signaling pathway. Moreover, FMP NPs efficiently scavenged reactive oxygen species (ROS) and mitigated the expression of inflammatory mediators, thereby reshaping the microenvironment to support hepatocyte repair. Finally, administration of bone marrow mesenchymal stem cells (BMMSCs) was employed to promote the regeneration and functional recovery of damaged hepatocytes. In conclusion, the combined sequential therapy involving FMP and BMMSCs effectively attenuated liver fibrosis induced by CCl4 administration, resulting in significant amelioration of the fibrotic condition. The therapeutic strategy outlined in this study underscores the significance of disrupting the deleterious cellular interactions and remodeling the microenvironment, thereby presenting a promising avenue for clinical intervention in liver fibrosis.

[Research progress on induced hepatic stellate cell ferroptosis in prevention and treatment of liver fibrosis].

Liver fibrosis is a key pathological stage in the progression of chronic liver disease. If the disease is mistreated, it can further deteriorate into liver failure, which seriously affects the quality of life of patients and brings heavy medical costs. Hepatic stellate cell(HSC) activation triggers extracellular matrix(ECM) deposition, which plays an important driving role in liver fibrosis, and ferroptosis is an effective strategy to clear or reverse the activation of HSCs into a deactivated phenotype. Therefore, inhibiting the activation and proliferation of HSCs by regulating ferroptosis is the key to the treatment of this disease, so as to derive the prospect of inducing ferroptosis of HSCs(including RNA-binding proteins, non-coding RNA, chemicals, and active components of traditional Chinese medicine) to intervene in liver fibrosis. On this basis, this paper started from the activation of HSCs to induce ECM deposition and focused on summarizing the mechanism of inducing HSC ferroptosis in delaying the progression of liver fibrosis, so as to continuously enrich the clinical practice of liver fibrosis and provide a reference for subsequent basic research.

The Upregulation of Leucine-Rich Repeat Containing 1 Expression Activates Hepatic Stellate Cells and Promotes Liver Fibrosis by Stabilizing Phosphorylated Smad2/3.

Liver fibrosis poses a significant global health risk due to its association with hepatocellular carcinoma (HCC) and the lack of effective treatments. Thus, the need to discover additional novel therapeutic targets to attenuate liver diseases is urgent. Leucine-rich repeat containing 1 (LRRC1) reportedly promotes HCC development. Previously, we found that LRRC1 was significantly upregulated in rat fibrotic liver according to the transcriptome sequencing data. Herein, in the current work, we aimed to explore the role of LRRC1 in liver fibrosis and the underlying mechanisms involved. LRRC1 expression was positively correlated with liver fibrosis severity and significantly elevated in both human and murine fibrotic liver tissues. LRRC1 knockdown or overexpression inhibited or enhanced the proliferation, migration, and expression of fibrogenic genes in the human hepatic stellate cell line LX-2. More importantly, LRRC1 inhibition in vivo significantly alleviated CCl4-induced liver fibrosis by reducing collagen accumulation and hepatic stellate cells' (HSCs) activation in mice. Mechanistically, LRRC1 promoted HSC activation and liver fibrogenesis by preventing the ubiquitin-mediated degradation of phosphorylated mothers against decapentaplegic homolog (Smad) 2/3 (p-Smad2/3), thereby activating the TGF-ß1/Smad pathway. Collectively, these results clarify a novel role for LRRC1 as a regulator of liver fibrosis and indicate that LRRC1 is a promising target for antifibrotic therapies.

Antler thymosin ß10 reduces liver fibrosis via inhibiting TGF-ß1/SMAD pathway.

Hepatic stellate cell (HSC) activation is a crucial step in the development of liver fibrosis. Previous studies have shown that antler stem cells (AnSCs) inhibited HSC activation, suggesting that this may be achieved through secreting or releasing peptides. This study aimed to investigate whether AnSC-derived peptides (AnSC-P) could reduce liver fibrosis. The results showed that AnSC-P effectively reduced liver fibrosis in rats. Furthermore, we found that thymosin ß10 (Tß-10) was rich in AnSC-P, which may be the main component of AnSC-P contributing to the reduction in liver fibrosis. A further study showed that Tß-10 reduced liver fibrosis in rats, with a reduction in HYP and MDA levels in the liver tissues, a decrease in the serum levels of ALP, ALT, AST, and TBIL and an increase in TP and ALB. Moreover, Tß-10 decreased the expression levels of the genes related to the TGF-ß/SMAD signaling pathway in vivo. In addition, Tß-10 also inhibited TGF-ß1-induced HSC activation and decreased the expression levels of the TGF-ß/SMAD signaling pathway-related genes in HSCs in vitro. In conclusion, antler Tß-10 is a potential drug candidate for the treatment of liver fibrosis, the effect of which may be achieved via inhibition of the TGFß/SMAD signaling pathway.

Ginsenoside Rg3 promotes hepatic stellate cell ferroptosis by epigenetically regulating ACSL4 to suppress liver fibrosis progression.

BACKGROUND: Ginsenoside Rg3 (G-Rg3), extracted from Panax notoginseng, possesses hepatoprotective properties. Hepatic stellate cells (HSCs) activation is responsible for liver fibrosis. Recent studies have reported the suppressive effects of G-Rg3 on HSC activation and proliferation. Ferroptosis is a novel iron regulated cell death. ACSL4, a key indicator of ferroptosis, is commonly methylated in various diseases. PURPOSE: However, the role of ACSL4 methylation-mediated HSC ferroptosis in G-Rg3 inhibition of hepatic fibrosis needs to be explored. METHODS: Effects of G-Rg3 on inhibiting fibrosis were evaluated in vivo and in vitro. The impact of G-Rg3 on HSC ferroptosis was assessed in vitro. Furthermore, the expression of ACSL4, ACSL4 methylation and microRNA-6945-3p (miR-6945-3p) levels were determined. RESULTS: G-Rg3 significantly alleviated CCl4-induced liver fibrosis, accompanied by collagen downregulation. In vitro, G-Rg3 contributed to HSC inactivation, leading to decreased collagen production. G-Rg3 induced HSC ferroptosis, characterized by increased iron accumulation, depletion of glutathione, malondialdehyde levels, and generation of lipid reactive oxygen species. Moreover, G-Rg3 promoted ACSL4 demethylation and restored its expression. Notably, DNMT3B counteracted the effect of G-Rg3-mediated inhibition of ACSL4 methylation and was targeted by miR-6945-3p. Further investigations revealed that G-Rg3 suppressed ACSL4 methylation through miR-6945-3p-mediated DNMT3B inhibition. Consistent with this, miR-6945-3p inhibition reversed G-Rg3-induced ACSL4 expression and HSC ferroptosis. CONCLUSION: G-Rg3 inhibits ACSL4 methylation by miR-6945-3p-mediated DNMT3B inhibition, thereby promoting HSC ferroptosis and mitigating liver fibrosis.

Runx2 activates hepatic stellate cells to promote liver fibrosis via transcriptionally regulating Itgav expression.

BACKGROUNDS AND AIMS: As a central event during liver fibrosis, hepatic stellate cells (HSC) have been thought to be a potential therapeutic target for liver fibrosis. Previous studies have shown that runt-related transcription factor 2 (Runx2) is associated with the development of non-alcoholic fatty liver disease, while its specific role in HSC activation and hepatic fibrosis remains elusive. APPROACH AND RESULTS: In this study, we found that Runx2 expression was significantly upregulated in human liver fibrosis with different aetiologies. Runx2 expression was also gradually elevated in mouse liver during fibrosis, and Runx2 was mainly expressed in the activated HSC. Knockdown of Runx2 in HSC markedly alleviated CCl4 -induced, 3,5-diethoxycarbonyl-1,4-dihydrocollidine-induced or methionine-choline deficient (MCD)-induced liver fibrosis, while hepatic overexpression of Runx2 via HBAAV-Runx2 or VA-Lip-Runx2 injection exacerbated CCl4 -induced liver fibrosis. In vitro analysis demonstrated that Runx2 promoted HSC activation and proliferation, whereas Runx2 knockdown in HSC suppressed these effects. RNA-seq and Runx2 ChIP-seq analysis demonstrated that Runx2 could promote integrin alpha-V (Itgav) expression by binding to its promoter. Blockade of Itgav attenuated Runx2-induced HSC activation and liver fibrosis. Additionally, we found that cytokines (TGF-ß1, PDGF, EGF) promote the expression and nuclear translocation of Runx2 through protein kinase A (PKA) in HSC. CONCLUSIONS: Runx2 is critical for HSC activation via transcriptionally regulating Itgav expression during liver fibrosis, and may be a promising therapeutic target for liver fibrosis.

Attenuation of hepatic fibrosis by p-Coumaric acid via modulation of NLRP3 inflammasome activation in C57BL/6 mice.

A prolonged high-fat and high-sucrose (HFHS) diet induces hepatic inflammation and mediates hepatic stellate cell (HSC) activation, which result in hepatic fibrosis. Aberrant activation of the innate immune system components, such as the NOD-like receptor protein 3 (NLRP3) inflammasome, has been implicated in HSC activation and hepatic fibrosis. We have previously shown that p-coumaric acid (PCA)-enriched peanut sprout extracts exert anti-inflammatory effects. However, it is unknown whether PCA reduces hepatic fibrosis by modulating innate immunity and HSC activation. To test this hypothesis, C57BL/6 male mice were randomly assigned to three groups and fed low-fat (LF) diet (11% calories from fat), high-fat (HF) diet (60% calories from fat, 0.2% cholesterol) with sucrose drink (20% sucrose, HFHS), or HFHS diet with PCA treatment (HFHS+PCA, 50 mg/kg body weight, intraperitoneally) for 13 weeks. The results showed that PCA treatment (1) partly improved systemic insulin sensitivity without altering adiposity, (2) attenuated hepatic signaling pathways associated with NLRP3 inflammasome activation, including toll-like receptor 4 (TLR4)/nuclear factor kappa B (NFκB), and endoplasmic reticulum/oxidative stress, and (3) reduced circulating interleukin (IL)-1ß levels. More importantly, PCA ameliorated hepatic fibrosis compared to that in the HFHS group, and the anti-fibrogenic effects of PCA were confirmed in vitro in transforming growth factor ß (TGFß) treated-LX-2 HSCs. The role of PCA in decreased NLRP3 activation and caspase-1 cleavage was recapitulated in primary bone marrow‒derived macrophages. These findings indicate that PCA contributes to the prevention of HFHS diet‒mediated liver fibrosis, partly by attenuating the activation of the NLRP3 inflammasome.

Effects of specific knockdown of ribosomal protein S5 in hepatic stellate cells on liver fibrosis / 药学实践与服务

Objective To observe the effect of specific knockdown of hepatic stellate cells (HSC) ribosomal protein S5 (RPS5) on liver fibrosis in rats. Methods The glial fibrillary acidic protein (GFAP) promoter-driven RPS5 shRNA adenovirus was established, and AdGFa2-shRPS5 and its control AdGFa2 shNC were used to transfect primary rat HSCs and hepatocytes, respectively. RPS5 was determined by Western-blot and Real Time PCR, α-SMA and type I collagen expression; the rat liver fibrosis model was established by dimethyl nitrosamine (DMN) and bile duct ligation (BDL), and intrahepatic HSC was specifically knocked down by tail vein injection of adenovirus of RPS5 levels. The pathological changes of liver tissue sections were analyzed by HE staining; the content of hydroxyproline, sections of Sirius red and Masson staining were used to evaluate collagen deposition; immunohistochemical staining was used to detect the expression of α-SMA and RPS5. Results AdGFa2-shRPS5 specifically knocked down the expression level of RPS5 in HSC and increased the expression of α-SMA and type I collagen in vitro. The in vivo results showed that in two animal models of chronic liver injury, specific knockdown of RPS5 expression in HSCs promoted HSC activation, increased the deposition of extracellular matrix, and promoted liver fibrosis. Conclusion RPS5 is essential for HSC activation and liver fibrosis, which could be a potential target for the treatment of liver fibrosis.

Long non-coding RNA in Non-alcoholic fatty liver disease.

Nonalcoholic fatty liver disease (NAFLD) is a chronic disease of worldwide impact. The disease process begins with steatosis, i.e., fat accumulation in the liver, and proceeds to nonalcoholic steatohepatitis (NASH). Liver biopsy is the gold standard for NASH diagnosis, but the procedure is invasive, expensive, error prone and poses considerable risk. Unfortunately, there are currently no precise FDA-approved therapies for NAFLD, the only options being lifestyle change and symptomatic treatment. Recently, much research has focused on the identification of molecular mechanisms that could be translated into novel diagnostics and therapeutics. With the advent of high throughput genomics and transcriptomics, noncoding RNAs, including long non-coding RNAs (lncRNAs) have been identified as key players of NAFLD pathogenesis and have accordingly attracted much attention as potential diagnostics and therapeutics. In this chapter, we reviewed various lncRNAs and their functions at different stages of NAFLD. We also highlighted how these unique molecules can be developed as stage-specific non-invasive diagnostic biomarkers for NAFLD.

Early Growth Response Factor 1 in Aging Hematopoietic Stem Cells and Leukemia.

Aging is associated with various hematological disorders and a higher risk of myeloproliferative disorders. An aged hematopoietic system can be characterized by decreased immune function and increased myeloid cell production. Hematopoietic stem cells (HSCs) regulate the production of blood cells throughout life. The self-renewal and regenerative potential of HSCs determine the quality and quantity of the peripheral blood cells. External signals from the microenvironment under different conditions determine the fate of the HSCs to proliferate, self-renew, differentiate, or remain quiescent. HSCs respond impromptu to a vast array of extracellular signaling cascades such as cytokines, growth factors, or nutrients, which are crucial in the regulation of HSCs. Early growth response factor 1 (EGR1) is one of the key transcription factors controlling HSC proliferation and their localization in the bone marrow (BM) niche. Downregulation of Egr1 activates and recruits HSCs for their proliferation and differentiation to produce mature blood cells. Increased expression of Egr1 is implicated in immuno-aging of HSCs. However, dysregulation of Egr1 is associated with hematological malignancies such as acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), and chronic myelogenous leukemia (CML). Here, we summarize the current understanding of the role of EGR1 in the regulation of HSC functionality and the manifestation of leukemia. We also discuss the alternative strategies to rejuvenate the aged HSCs by targeting EGR1 in different settings.

MiR-22-3p and miR-29a-3p synergistically inhibit hepatic stellate cell activation by targeting AKT3.

Hepatic fibrosis (HF) is a worldwide health problem for which there is no medically effective drug treatment at present, and which is characterized by activation of hepatic stellate cells (HSCs) and excessive extracellular matrix (ECM) deposition. The HF model in cholestatic rats by ligating the common bile duct was induced and the differentially expressed miRNAs in the liver tissues were analyzed by microarray, which showed that miR-22-3p and miR-29a-3p were significantly downregulated in bile-duct ligation (BDL) rat liver compared with the sham control. The synergistic anti-HF activity and molecular mechanism of miR-22-3p and miR-29a-3p by targeting AKT serine/threonine kinase 3 (AKT3) in HSCs were explored. The expression levels of miR-22-3p and miR-29a-3p were downregulated in activated LX-2 and human primary normal hepatic fibroblasts (NFs), whereas AKT3 was found to be upregulated in BDL rat liver and activated LX-2 cells. The proliferation, colony-forming, and migration ability of LX-2 were inhibited synergistically by miR-22-3p and miR-29a-3p. In addition, cellular senescence was induced and the expressions of the LX-2 fibrosis markers COL1A1 and α-SMA were inhibited by miR-22-3p and miR-29a-3p synergistically. Subsequently, these two miRNAs binding to the 3'UTR of AKT3 mRNA was predicted and evidenced by the luciferase reporter assay. Furthermore, the proliferation, migration, colony-forming ability, and the expression levels of COL1A1 and α-SMA were promoted and cellular senescence was inhibited by AKT3 in LX-2 cells. Thus, miR-22-3p/miR-29a-3p/AKT3 regulates the activation of HSCs, providing a new avenue in the study and treatment of HF.

Hesperetin derivative decreases CCl4 -induced hepatic fibrosis by Ptch1-dependent mechanisms.

Hepatic fibrosis (HF), a continuous wound-healing response of the liver to repeated injuries, is characterized by abnormal extracellular matrix (ECM) accumulation. Hepatic stellate cells (HSCs) are considered a major cell type for ECM production. However, recent evidence indicates the lack of effective treatments for HF. Hesperetin, a Traditional Chinese Medicine monomer, has been isolated from the fruit peel of Citrusaurantium L. (Rutaceae). Growing evidence suggests the partial function of hesperetin in HF treatment. A hesperetin derivative (HD) was synthesized in our laboratory to increase the bioavailability and the water solubility of hesperetin. In this study, we detected the functions of HD in a mouse model of CCl4 -induced HF and transforming growth factor-ß1-stimulated HSC-T6 cells, in vivo and in vitro. HD reduced histological damage and CCl4 -induced HF. Moreover, HD interference was associated with the activation of indicators in HSC-T6 cells, showing that HD is involved in HSCs activation in HF. Mechanistically, the Hedgehog pathway is involved in the HD treatment of HF, and HD may attenuate the aberrant expression of patched1. In conclusion, the studies indicate that HD may function as a potential antifibrotic Traditional Chinese Medicine monomer in HF therapy.

JAM-A is a multifaceted regulator in hepatic fibrogenesis, supporting LSEC integrity and stellate cell quiescence.

BACKGROUND AND AIMS: Leukocyte infiltration is a hallmark of hepatic inflammation. The Junctional Adhesion Molecule A (JAM-A) is a crucial regulator of leukocyte extravasation and is upregulated in human viral fibrosis. Reduced shear stress within hepatic sinusoids and the specific phenotype of liver sinusoidal endothelial cells (LSEC) cumulate in differing adhesion characteristics during liver fibrosis. The aim of this study was to define the functional role of cell-specific adhesion molecule JAM-A during hepatic fibrogenesis. METHODS: Complete, conditional (intestinal epithelial; endothelial) and bone marrow chimeric Jam-a knockout animals and corresponding C57Bl/6 wild-type animals were treated with carbon tetrachloride (CCl4 , 6 weeks). For functional analyses of JAM-A, comprehensive in vivo studies, co-culture models and flow-based adhesion assays were performed. RESULTS: Complete and bone marrow-derived Jam-a-/- animals showed aggravated fibrosis with increased non-sinusoidal, perivascular accumulation of CD11b+ F4/80+ monocyte-derived macrophages in contrast to wild-type mice. Despite being associated with disturbed epithelial barrier function, an intestinal epithelial Jam-a knockout did not affect fibrogenesis. In endothelial-specific Jam-a-/- animals, liver fibrosis was aggravated alongside sinusoid capillarization and hepatic stellate cell (HSC) activation. HSC activation is induced via Jam-a-/- LSEC-derived secretion of soluble factors. Sinusoid CD31 expression and hedgehog gene signalling were increased, but leukocyte infiltration and adhesion to LSECs remained unaffected. CONCLUSIONS: Our models decipher cell-specific JAM-A to exert crucial functions during hepatic fibrogenesis. JAM-A on bone marrow-derived cells regulates non-sinusoidal vascular immune cell recruitment, while endothelial JAM-A controls liver sinusoid capillarization and HSC quiescence.

miRNA-130b-5p promotes hepatic stellate cell activation and the development of liver fibrosis by suppressing SIRT4 expression.

Liver fibrosis is a progressive disease accompanied by the deposition of extracellular matrix (ECM). Numerous reports have demonstrated that alterations in the expression of microRNAs (miRNAs) are related to liver disease. However, the effect of individual miRNAs on liver fibrosis has not been studied. Hepatic stellate cells (HSCs), being responsible for producing ECM, exert an important influence on liver fibrosis. Then, microarray analysis of non-activated and activated HSCs induced by transforming growth factor ß1 (TGF-ß1) showed that miR-130b-5p expression was strongly up-regulated during HSC activation. Moreover, the progression of liver fibrosis had a close connection with the expression of miR-130b-5p in different liver fibrosis mouse models. Then, we identified that there were specific binding sites between miR-130b-5p and the 3' UTR of Sirtuin 4 (SIRT4) via a luciferase reporter assay. Knockdown of miR-130b-5p increased SIRT4 expression and ameliorated liver fibrosis in mice transfected with antagomiR-130b-5p oligos. In general, our results suggested that miR-130b-5p promoted HSC activation by targeting SIRT4, which participates in the AMPK/TGF-ß/Smad2/3 signalling pathway. Hence, regulating miR-130b-5p maybe serve as a crucial therapeutic treatment for hepatic fibrosis.

RUNX1 regulates SMAD1 by transcriptionally activating the expression of USP9X, regulating the activation of hepatic stellate cells and liver fibrosis.

Liver fibrosis (LF) is a common pathological process with high morbidity and mortality. Runt-related transcription factor 1 (RUNX1) is a transcription factor that could cause nephropathy and renal fibrosis, but its role in LF is unclear. Therefore, this study aimed to investigate the role RUNX1 in LF. Briefly, hepatic fibrosis was detected by Sirius Red staining. Transcript levels were quantified by qPCR, and proteins were assessed by western blotting or immunofluorescence. Cell viability and cell migration were measured by CCK8 assays and wound healing assays, respectively. The binding of RUNX1 and ubiquitin-specific protease 9X (USP9X) promoter was validated by ChIP assays and luciferase report assays, while the binding of USP9X and SMAD1 was confirmed by co-immunoprecipitation (Co-IP). Our studies found that the expression of RUNX1 was upregulated in LF mice, and RUNX1 knockdown alleviated CCl4-induced LF. RUNX1 silencing reduced the viability and migration of HSCs. Besides, RUNX1, as a transcription factor, bound to the promoter of USP9X and regulated the expression of USP9X. USP9X is a deubiquitination enzyme and was found to be up-regulated in LF mice. USP9X silencing reduced the viability and migration of HSCs, thereby inhibiting LF. Further studies showed that USP9X could stabilize downstream Smad1 expression. Furthermore, we also found that RUNX1 regulated the expression of SMAD1 by transcriptionally activating the expression of USP9X, thereby regulating the activation of hepatic stellate cells and liver fibrosis.

Hepatic Stellate Cell Activation and Inactivation in NASH-Fibrosis-Roles as Putative Treatment Targets?

Hepatic fibrosis is the primary predictor of mortality in patients with non-alcoholic steatohepatitis (NASH). In this process, the activated hepatic stellate cells (HSCs) constitute the principal cells responsible for the deposition of a fibrous extracellular matrix, thereby driving the hepatic scarring. HSC activation, migration, and proliferation are controlled by a complex signaling network involving growth factors, lipotoxicity, inflammation, and cellular stress. Conversely, the clearance of activated HSCs is a prerequisite for the resolution of the extracellular fibrosis. Hence, pathways regulating the fate of the HSCs may represent attractive therapeutic targets for the treatment and prevention of NASH-associated hepatic fibrosis. However, the development of anti-fibrotic drugs for NASH patients has not yet resulted in clinically approved therapeutics, underscoring the complex biology and challenges involved when targeting the intricate cellular signaling mechanisms. This narrative review investigated the mechanisms of activation and inactivation of HSCs with a focus on NASH-associated hepatic fibrosis. Presenting an updated overview, this review highlights key cellular pathways with potential value for the development of future treatment modalities.

Resolvin D1 attenuates CCl4 Induced Liver Fibrosis by Inhibiting Autophagy-Mediated HSC activation via AKT/mTOR Pathway.

Resolvin D1 (RvD1) was previously reported to relieve inflammation and liver damage in several liver diseases, but its potential role in liver fibrosis remains elusive. The aim of our study was to investigate the effects and underlying mechanisms of RvD1 in hepatic autophagy in liver fibrosis. In vivo, male C57BL/6 mice were intraperitoneally injected with 20% carbon tetrachloride (CCl4, 5 ml/kg) twice weekly for 6 weeks to establish liver fibrosis model. RvD1 (100 ng or 300 ng/mouse) was added daily in the last 2 weeks of the modeling period. In vitro, lipopolysaccharide (LPS)-activated LX-2 cells were co-treated with increasing concentrations (2.5-10 nM) of RvD1. The degree of liver injury was measured by detecting serum AST and ALT contents and H&E staining. Hepatic fibrosis was assessed by masson's trichrome staining and metavir scoring. The qRT-PCR, western blot, immunohistochemistry, and immunofluorescence were applied to liver tissues or LPS-activated LX-2 cells to explore the protective effects of RvD1 in liver fibrosis. Our findings reported that RvD1 significantly attenuated CCl4 induced liver injury and fibrosis by decreasing plasma AST and ALT levels, reducing collagen I and α-SMA accumulation and other pro-fibrotic genes (CTGF, TIMP-1 and Vimentin) expressions in mouse liver, restoring damaged histological architecture and improving hepatic fibrosis scores. In vitro, RvD1 also repressed the LPS induced LX-2 cells activation and proliferation. These significant improvements mainly attributed to the inhibiting effect of RvD1 on autophagy in the process of hepatic stellate cell (HSC) activation, as demonstrated by decreased ratio of LC3-II/I and elevated p62 after RvD1 treatment. In addition, using AZD5363 (an AKT inhibitor that activates autophagy) and AZD8055 (an mTOR inhibitor, another autophagy activator), we further verified that RvD1 suppressed autophagy-mediated HSC activation and alleviated CCl4 induced liver fibrosis partly through AKT/mTOR pathway. Overall, these results demonstrate that RvD1 treatment is expected to become a novel therapeutic strategy against liver fibrosis.

Macrophages Orchestrate Hematopoietic Programs and Regulate HSC Function During Inflammatory Stress.

The bone marrow contains distinct cell types that work in coordination to generate blood and immune cells, and it is the primary residence of hematopoietic stem cells (HSCs) and more committed multipotent progenitors (MPPs). Even at homeostasis the bone marrow is a dynamic environment where billions of cells are generated daily to replenish short-lived immune cells and produce the blood factors and cells essential for hemostasis and oxygenation. In response to injury or infection, the marrow rapidly adapts to produce specific cell types that are in high demand revealing key insight to the inflammatory nature of "demand-adapted" hematopoiesis. Here we focus on the role that resident and monocyte-derived macrophages play in driving these hematopoietic programs and how macrophages impact HSCs and downstream MPPs. Macrophages are exquisite sensors of inflammation and possess the capacity to adapt to the environment, both promoting and restraining inflammation. Thus, macrophages hold great potential for manipulating hematopoietic output and as potential therapeutic targets in a variety of disease states where macrophage dysfunction contributes to or is necessary for disease. We highlight essential features of bone marrow macrophages and discuss open questions regarding macrophage function, their role in orchestrating demand-adapted hematopoiesis, and mechanisms whereby they regulate HSC function.

Physalin D attenuates hepatic stellate cell activation and liver fibrosis by blocking TGF-ß/Smad and YAP signaling.

BACKGROUND: Hepatic fibrosis is considered integral to the progression of chronic liver diseases, as it leads to the development of cirrhosis and hepatocellular carcinoma. The activation of hepatic stellate cells (HSCs) is the dominant event in hepatic fibrogenesis. The transforming growth factor-ß1 (TGF-ß1) and Yes-associated protein (YAP) pathways play a pivotal role in HSC activation, hepatic fibrosis and cirrhosis progression. Therefore, targeting the TGF-ß/Smad and YAP signaling pathways is a promising strategy for antifibrotic therapy. PURPOSE: The present study investigated the protective effects of Physalin D (PD), a withanolide isolated from Physalis species (Solanaceae), against liver fibrosis and further elucidated the mechanisms involved in vitro and in vivo. STUDY DESIGN/METHODS: We conducted a series of experiments using carbon tetrachloride (CCl4)- and bile duct ligation (BDL)-induced fibrotic mice and cultured LX-2 cells. Serum markers of liver injury, and the morphology, histology and fibrosis of liver tissue were investigated. Western blot assays and quantitative real-time PCR were used to investigate the mechanisms underlying the antifibrotic effects of PD. RESULT: PD decreased TGF-ß1-induced COL1A1 promoter activity. PD inhibited TGF-ß1-induced expression of Collagen I and α-smooth muscle actin (α-SMA) in human hepatic stellate LX-2 cells. PD significantly ameliorated hepatic injury, including transaminase activities, histology, collagen deposition and α-SMA, in CCl4- or BDL-induced mice. Moreover, PD markedly decreased the expression of phosphorylated Smad2/3 in vitro and in vivo. Furthermore, PD significantly decreased YAP protein levels, and YAP knockdown did not further enhance the effects of PD, namely α-SMA inhibition, Collagen I expression and YAP target gene expression in LX-2 cells. CONCLUSION: These results clearly show that PD ameliorated experimental liver fibrosis by inhibiting the TGF-ß/Smad and YAP signaling pathways, indicating that PD has the potential to effectively treat liver fibrosis.