MANF brakes TLR4 signaling by competitively binding S100A8 with S100A9 to regulate macrophage phenotypes in hepatic fibrosis

The mesencephalic astrocyte-derived neurotrophic factor (MANF) has been recently identified as a neurotrophic factor, but its role in hepatic fibrosis is unknown. Here, we found that MANF was upregulated in the fibrotic liver tissues of the patients with chronic liver diseases and of mice treated with CCl4. MANF deficiency in either hepatocytes or hepatic mono-macrophages, particularly in hepatic mono-macrophages, clearly exacerbated hepatic fibrosis. Myeloid-specific MANF knockout increased the population of hepatic Ly6Chigh macrophages and promoted HSCs activation. Furthermore, MANF-sufficient macrophages (from WT mice) transfusion ameliorated CCl4-induced hepatic fibrosis in myeloid cells-specific MANF knockout (MKO) mice. Mechanistically, MANF interacted with S100A8 to competitively block S100A8/A9 heterodimer formation and inhibited S100A8/A9-mediated TLR4-NF-κB signal activation. Pharmacologically, systemic administration of recombinant human MANF significantly alleviated CCl4-induced hepatic fibrosis in both WT and hepatocytes-specific MANF knockout (HKO) mice. This study reveals a mechanism by which MANF targets S100A8/A9-TLR4 as a "brake" on the upstream of NF-κB pathway, which exerts an impact on macrophage differentiation and shed light on hepatic fibrosis treatment.

Remodeling of the liver fibrosis microenvironment based on nilotinib-loaded multicatalytic nanozymes with boosted antifibrogenic activity

Liver fibrosis is a reversible pathological process caused by chronic liver damage and a major risk factor for hepatocellular carcinoma (HCC). Hepatic stellate cell (HSC) activation is considered the main target for liver fibrosis therapy. However, the efficiency of this strategy is limited due to the complex microenvironment of liver fibrosis, including excessive extracellular matrix (ECM) deposition and hypoxia-induced imbalanced ECM metabolism. Herein, nilotinib (NIL)-loaded hyaluronic acid (HA)-coated Ag@Pt nanotriangular nanozymes (APNH NTs) were developed to inhibit HSCs activation and remodel the microenvironment of liver fibrosis. APNH NTs efficiently eliminated intrahepatic reactive oxygen species (ROS) due to their inherent superoxide dismutase (SOD) and catalase (CAT) activities, thereby downregulating the expression of NADPH oxidase-4 (NOX-4) and inhibiting HSCs activation. Simultaneously, the oxygen produced by the APNH NTs further alleviated the hypoxic microenvironment. Importantly, the released NIL promoted collagen depletion by suppressing the expression of tissue inhibitor of metalloproteinase-1 (TIMP-1), thus synergistically remodeling the microenvironment of liver fibrosis. Notably, an in vivo study in CCl4-induced mice revealed that APNH NTs exhibited significant antifibrogenic effects without obvious long-term toxicity. Taken together, the data from this work suggest that treatment with the synthesized APNH NTs provides an enlightening strategy for remodeling the microenvironment of liver fibrosis with boosted antifibrogenic activity.

Effects of YTHDF2 on proliferation and migration of activated hepatic stellate cells / 中国药理学通报

Aim To investigate the effect of YTHDF2 on the proliferation and migration of activated hepatic stellate cells(HSCs). Methods 5 jjLg • L

Ligustroflavone ameliorates CCl / 中国天然药物

Liver fibrosis is a pathological process characterized by excess deposition of extracellular matrix (ECM) that are mainly derived from activated hepatic stellate cells. Previous studies suggested that ligustroflavone (LF) was an ingredient of Ligustrum lucidum Ait. with activities of anti-inflammation and anti-oxidation. In this study, we investigated whether LF had any effect on liver fibrosis. In our study, we established a mouse model of carbon tetrachloride (CCl

Role of microRNAs in progress of liver Hbrosis / 中国药理学通报

Liver fibrosis is one of the main diseases with high morbidity and mortality worldwide. It is the common pathological results of several chronic irritant disease such as viral hepatitis ，alcohol abuse, autoimmune diseases, metabolic diseases and cholestatic liver diseases and will further develop into cirrhosis ，liver failure, portal hypertension and even death. The excessive accumulation of extracellular matrix (ECM) that leads to disorder of liver structure is the main factor in the development of liver fibrosis. MicroRNAs are a class of 2225 nt endogenous noncoding small RNAs. Sufficient studies have shown that the abnormal expression of microRNAs is closely related to the progression of liver fibrosis. In this review, we summarize the regulatory effects of microRNAs discovered in recent years on the activation ，proliferation apoptosis and senescence of HSCs in liver fibrosisand the underlying mechanisms, putting forward the prospect.

miR-140-3p Knockdown Suppresses Cell Proliferation and Fibrogenesis in Hepatic Stellate Cells via PTEN-Mediated AKT/mTOR Signaling

PURPOSE: Liver fibrosis is a major cause of morbidity and mortality and the outcome of various chronic liver diseases. Activation of hepatic stellate cells (HSCs) is the key event in liver fibrosis. Studies have confirmed that miR-140-3p plays a potential regulatory effect on HSC activation. However, whether miR-140-3p mediates the liver fibrosis remains unknown. MATERIALS AND METHODS: Expression of miR-140-3p was detected by real-time quantitative PCR (qPCR). Cell proliferation was measured by MTT, while cell apoptosis rate was determined via flow cytometry. Western blot assay was used to detect the expression of cleaved PARP. The fibrogenic effect was evaluated by expression of α-smooth muscle actin and desmin. Functional experiments were performed in transforming growth factor β1 (TGF-β1)-induced HSC-T6 cells with transfection of anti-miR-140-3p and/or siPTEN. Target binding between miR-140-3p and PTEN was predicted by the TargetScan database and identified using luciferase reporter assay and RNA immunoprecipitation. RESULTS: TGF-β1 induced the activation of HSC-T6 cells, and miR-140-3p expression varied according to HSC-T6 cell activation status. Knockdown of miR-140-3p reduced cell proliferation and the expressions of α-SMA and desmin, as well as increased apoptosis, in TGF-β1-induced HSC-T6 cells, which could be blocked by PTEN silencing. Additionally, inactivation of the AKT/mTOR signaling pathway stimulated by miR-140-3p knockdown was abolished when silencing PTEN expression. PTEN was negatively regulated by miR-140-3p via direct binding in HSC-T6 cells. CONCLUSION: miR-140-3p is an important mediator in HSC-T6 cell activation, and miR-140-3p knockdown suppresses cell proliferation and fibrogenesis in TGF-β1-induced HSC-T6 cells, indicating that miR-140-3p may be a potential novel molecular target for liver fibrosis.

PD-L1 Expression on Circulating CD34+ Hematopoietic Stem Cells Closely Correlated with T-cell Apoptosis in Chronic Hepatitis C Infected Patients

BACKGROUND AND OBJECTIVES: Lack of understanding of the interplay between hematopoietic stem cells (HSCs) and the immune system has severely hampered stem cell research. Programmed death-1 (PD-L1) has been reported on parenchymal cells in patients with chronically inflamed livers and found to play an essential role in T cell homeostasis regulation. However, the bidirectional interaction between HSCs and lymphocytes remains elusive. Here, we aimed to get more insight into circulating CD34+ HSCs PD-L1 expression and T cell apoptosis in chronic HCV infected patients. METHODS: CD34+ HSCs were isolated and purified by immunomagnetic separation. PD-L1 expression was analyzed by quantitative PCR and flow cytometry. Furthermore, co-culture experiments between CD34+ HSCs and T-lymphocytes were established. T-cell lymphocyte apoptosis in peripheral blood and in cultures was detected. RESULTS: CD34+ HSCs constitutively express low levels of PD-L1. Its expression is up-regulated in chronic HCV infected patients. Moreover, PD-L1 expression on circulating CD34+ HSCs enhanced T cell apoptosis in peripheral blood and co-culture. CONCLUSION: Our results suggest novel bidirectional interplay between HSCs and lymphocytes mediated by PD-L1 expression on CD34+ HSCs. PD-L1 expression correlated with T-cell lymphocyte apoptosis. This may contribute to immunomodulatory properties of HSCs which improves its use for allogeneic transplantation.

Elevated expression of DNMT1 is associated with increased expansion and proliferation of hematopoietic stem cells co-cultured with human MSCs

BACKGROUND: Mesenchymal stem cells (MSCs) play an important role in hematopoietic stem cell (HSC) maintenance, proliferation, and apoptosis. DNA methyltransferase 1 (DNMT1) is considered an essential factor in the maintenance of HSCs in mammalian cells. Therefore, this study was conducted to evaluate the mRNA expression level of DNMT1 during cord blood (CB)-HSC ex vivo expansion with MSCs. METHODS: Ex vivo cultures of CB-HSCs were performed in three culture conditions for 7 days: cytokines, cytokines with MSCs, and only MSCs. Total and viable cell numbers were counted after 5 and 7 days using trypan blue stain, and the stem cell percentage was then evaluated by flow cytometry. Moreover, in vitro colony-forming unit assay was carried out to detect clonogenic potential of HSCs at days 0 and 7 using MethoCult H4434. Finally, DNMT1 mRNA expression level was evaluated by real-time polymerase chain reaction. RESULTS: Maximum CB-CD34⁺ cell expansion was observed on day 7 in all the three cultures. After 7 days, ex vivo expansion of CB-CD34⁺ cells indicated a significant decrease in DNMT1 expression in the cytokine cultures, whereas in the two co-culture conditions DNMT1 expression was increased. A significant difference between the number of CD34⁺ and CD34⁻ cells in the cytokine co-culture system was observed. CONCLUSION: These data indicated that an elevated expression of DNMT1 is associated with increased expansion and proliferation of HSCs co-cultured with human MSCs. Hence, DNMT1 may be a potential factor in the maintenance of expanded HSCs co-cultured with human MSCs.

Study on Inhibitory Effect of Lycianthes biflora Polysaccharide on the Proliferation of Hepatic Stellate HSCs-T6 Cells in Rats and Its Mechanism / 中国药房

OBJECTIVE:To study inhibitory effect of Lycianthes biflora polysaccharide on the proliferation of hepatic stellate HSCs-T6 cells in rats and its mechanism. METHODS:After treated with 0(blank control),50,100,200μg/ml L. biflora polysac-charide for 24 and 48 h,the activity of hepatic stellate HSCs-T6 cells in rats was determined by MTT assay and inhibitory rate of cell proliferation was calculated;the content of Hyp in supernatant was detected by immunohistochemical assay. After 48 h,the ex-pression of cellular α-smooth muscle actin(α-SMA)was measured by immunohistochemical assay;both transforming growth fac-tor β1(TGF-β1)and Smad3 were measured by Western blot assay. RESULTS:Compared to blank control,50,100 and 200 μg/ml L. biflora polysaccharide could inhibit the proliferation of HSCs-T6 cells;cell inhibitory rates were 39.84%-69.31% and 45.16%-82.93% respectively after treated for 24 and 48 h,which were positively associated with time and concentration. The con-tents of Hyp in supernatant were 178.36-93.25 μg/ml and 131.94-68.74 μg/ml respectively after treated with different concentrations of PRP for 24 and 48 h,which were negatively associated with time and concentration. The protein level of TGF-β1 and Smad3 de-creased after treated with L. biflora polysaccharide for 48 h(P<0.01). CONCLUSIONS:L. biflora polysaccharide can inhibit the proliferation of hepatic stellate HSCs-T6 cells in rats,and its mechanism is associated with the inactivation of endogenous TGF-βpathway for reducing collagen production.

Effect of TFB on TGF-β1 signaling pathway in HSC of liver fibrosis rats / 中国药理学通报

Aim To investigate the proliferation of HSCs stimulated by exogenous TGF-β1 (transforming growth factor betal),observe the effect of TFB(total flavonoids of Bidens Bipinnata L.)on smad2/7,typeⅠcollagen mRNA and protein expression of HSCs and study the protective effect and molecular mechanism of TFB on hepatic fibrosis.Methods HSCs were isolated with collagenase Ⅳ perfusion in situ and density gradient centrifugation. The effect of TFB on cell proliferation was observed by MTT colormetric assay. The auto-secretion of TGF-β1 and synthesis of type Ⅰ collagen were measured by enzyme-linked immuneadsordent assay (ELISA).Moreover,the expression of smad2/7, typeⅠcollagen mRNA and protein was measured by semi-quantitative RT-PCR and Western blot methods respectively.Results TFB could markedly inhibit the proliferation of HSCs of liver fibrosis rats stimulated by TGF-β1 and production of TGF-β1 and type Ⅰ collagen.In addition,TFB treatment could significantly down-regulate smad2 and type Ⅰ collagen mRNA expression and up-regulated smad7 mRNA expression of HSCs Smad2 protein expression of HSCs stimulated by TGF-β1 was also down-regulated by TFB.Conclusion TFB has the protective effect against hepatic fibrosis by inhibiting the activation of TGF-β1 signaling pathway and suppressing the HSC proliferation.

Effect Of Salidroside on Apoptosis of Rat Hepatic Stellate Cells Stimulated by Acetaldehyde / 中药新药与临床药理

Objective To investigate the effect of Salidroside in inducing apoptosis of rat hepatic stellate cells (HSC) stimulated by acetaldehyde and to observe the changes of c- Jnk N- terminal kinase (JNK) activity.Methods HSC stimulated by acetaldehyde were cultured in vitro and were treated with different concentrations of Salidroside.Apoptotic rate was analyzed by flow cytometry and the activity of phosphorylating JNK was measured by Western blot method.Results Salidroside in different concentrations (1.0,1.5,2.0 mg/mL) suppressed the activity of JNK in a dose- effect manner.Average light density was 35.8? 3.4,24.9? 2.7 and 3.4? 0.9 in Salidroside groups, which differed from that in acetaldehyde group( 48.6? 4.8; P

THE EXPRESSION OF P75 IN HEPATIC STELLATE CELLS OF FIBROTIC HUMAN AND RAT LIVER / 解剖学报

Objective To study the expression site and the role of low-affinity nerve growth factor receptor(P75) in hepatic stellate cells(HSCs) of fibrotic human and rat liver.Methods Left lobe of liver was removed from CCl-4-induced rats fibrosis model and human fibrotic tissue sample was taken from liver puncture.The tissue specimen was routinely paraffin embedded and expression of P75 was detected by histochemical staining using polyclonal antibody against P75.Additionally,HSCs was cultivated and P75 was immunochemically assayed for P75 binding activity. Results P75 was seen abundantly on cell membrane of the cultured HSCs.In the fibrotic tissue of rat liver as well as human fibrotic liver P75 was also positively expressed.Similarly,P75 was found mainly a hepatocye surface in histochemical staining.P75 was not seen by histochemical staining in the normal tissue of human and rat liver.Conclusion P75 used as a new target for the treatment of hepatic fibrosis.