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Background and aims: Myofibroblasts, the primary producers of extracellular matrix, primarily originate from hepatic stellate cells (HSCs), and their activation plays a pivotal role in liver fibrosis. This study aimed to investigate the function of CXC motif ligand 14 (CXCL14) in the progression of liver fibrosis. Approach and results: CXCL14 knockdown significantly reduced the extent of liver fibrosis. Using Ingenuity pathway analysis and qRT‐PCR, activating transcription factor 3 (ATF3) was identified as a key upstream regulator of CXCL14 expression. Mechanistically, ATF3 was shown to bind to the CXCL14 promoter, promoting its transactivation by TGF‐β in HSCs. Notably, both global CXCL14 deletion (CXCL14−/−) and HSC/myofibroblast‐specific CXCL14 knockdown significantly attenuated liver fibrosis in mice. RNA‐seq comparisons between CXCL14−/− and WT mice highlighted Jak2 as the most significantly downregulated gene, implicating its role in the antifibrotic effects of CXCL14 suppression on HSC inactivation. Moreover, Jak2 overexpression reversed the inhibition of liver fibrosis caused by CXCL14 knockdown in vivo. Conclusions: These findings unveil a novel ATF3/CXCL14/Jak2 signalling axis in liver fibrosis, presenting potential therapeutic targets for the disease. [ABSTRACT FROM AUTHOR]

Introduction: Liver fibrosis is primarily driven by the activation of hepatic stellate cells (HSCs), which involves various epigenetic modifications. Objectives: N6‐methyladenosine (m6A), the most prevalent RNA modification in eukaryotic cells, influences numerous physiological and pathological processes. Nevertheless, the role of insulin‐like growth factor 2 mRNA‐binding protein 3 (IGF2BP3), a reader gene mediating m6A modifications, in liver fibrosis remains unclear. Methods and results: This study demonstrated that IGF2BP3 knockout reduces liver fibrosis by promoting HSC ferroptosis (FPT) and inactivating HSCs. Multi‐omics analysis revealed that HSC‐specific IGF2BP3 knockout decreased m6A content in Jagged1 (Jag1), a key component of the Notch signalling pathway. Furthermore, IGF2BP3 deficiency significantly reduced the expression of hairy and enhancer of split‐1 (Hes1), a transcription factor in the Notch/Jag1 signalling pathway, with mRNA levels declining to 35%–62% and protein levels to 28%–35%. Additionally, it suppressed glutathione peroxidase 4 (GPX4) (decreased to approximately 31%–38%), a negative regulator of FPT, thereby facilitating HSC FPT progression and reducing profibrotic gene expression. Conclusion: These findings uncover a novel IGF2BP3/Notch/Jag1 signalling pathway involving HSC FPT, suggesting promising targets for ameliorating liver fibrosis. Key Points/Highlights: IGF2BP3 deficiency inactivates Jag1 signalling.IGF2BP3 deficiency‐mediated m6A modifications promote HSC ferroptosis.IGF2BP3 inhibition facilitates ferroptosis in HSCs via the Hes1/GPX4 axis.IGF2BP3 deficiency inactivates Jag1/Notch1/3/Hes1 signalling pathway inactivation, leading to the decrease in GPX4, which contributes to HSC ferroptosis. [ABSTRACT FROM AUTHOR]

It is known that ribonucleotide reductase M2 (RRM2) could be induced by hepatitis B virus (HBV) via DNA damage response. However, whether RRM2 is a potential biomarker for diagnosing and monitoring liver fibrosis in chronic hepatitis B (CHB) patients is still unclear. In this study, CHB patients from GSE84044 (a transcriptome data from GEO data set) were downloaded and RRM2 was selected as a hub gene. Interestingly, a positive correlation was found between serum RRM2 and liver fibrosis stage. The similar results were found in CHB patients with normal alanine aminotransferase (ALT). Notably, RRM2 could effectively differentiate preliminary fibrosis from advanced fibrosis in CHB patients with/without normal ALT. In addition, RRM2 had a better performance in diagnosing liver fibrosis than two commonly used noninvasive methods (aspartate aminotransferase‐to‐platelet ratio index and fibrosis index based on the four factors), two classic fibrotic biomarkers (hyaluronic acid and type IV collagen) as well as Mac‐2 binding protein glycosylation isomer, a known serum fibrosis marker. Moreover, CHB patients with high RRM2, who were associated with advanced fibrosis, had higher expressions of immune checkpoints. Overall, serum RRM2 may be a promising biomarker for diagnosing and monitoring liver fibrosis in CHB patients. [ABSTRACT FROM AUTHOR]

We aimed to develop and validate a novel combined score to improve the assessment of liver fibrosis progression in patients with chronic hepatitis B (CHB). In this study, a total of 331 CHB patients from three cohorts who underwent liver biopsy were enrolled, and the Scheuer system was used for liver fibrosis classification. The combined score was derived by principal component analysis of key differentially expressed genes. For significant liver fibrosis (≥S2), the areas under the receiver operating characteristics curves (AUROCs) of the combined score were 0.838, 0.842, and 0.881 in the three cohorts, respectively. And for advanced liver fibrosis (≥S3), the AUROCs were 0.794, 0.801, and 0.901, respectively. Compared with the results of AUROCs for aspartate aminotransferase≥to≥platelet ratio (APRI) and fibrosis index based on four factors (FIB‐4) in the validation cohorts, better clinical diagnostic value for assessing the progression of liver fibrosis was found in the combined score. Additionally, univariate ordered logistic regression analysis indicated that the combined score could serve as a more superior and stable risk factor than APRI and FIB‐4 in the assessment of liver fibrosis. For CHB patients with normal alanine aminotransferase (ALT), our results further emphasized the diagnostic value of the combined score for significant fibrosis (≥S2) and advanced fibrosis (≥S3). Moreover, it was found that patients with the high combined score, who were associated with the advanced fibrosis stage, had higher levels of drug sensitivity and immune checkpoint expression. In conclusion, the novel combined score could serve as a potential biomarker and contribute to improving the assessment of fibrosis stage in CHB patients. [ABSTRACT FROM AUTHOR]

Kaempferol, a natural flavonoid molecule, has demonstrated anti-inflammatory, antimicrobial and antioxidant activities. Recent studies have shown the beneficial effects of kaempferol on liver fibrosis. Notch pathway has been reported to be involved in the aberrant activation of hepatic stellate cells (HSCs). However, whether Notch pathway plays a key role in the anti-fibrotic effects of kaempferol is largely unknown. In this study, kaempferol significantly suppressed liver fibrosis in CCl4 mice, with reduced collagen deposition as well as restored liver function. In vitro , kaempferol enhanced the suppression of HSC activation, with a decrease in α -SMA as well as collagen level. It was found that Notch pathway played an important role in kaempferol-reduced the activation of HSCs. Jag1, a ligand of Notch pathway, was obviously inhibited by kaempferol. Overexpression of Jag1 effectively abolished kaempferol-induced HSC inactivation. Furthermore, Jag1 was demonstrated as a target of microRNA-26b-5p (miR-26b-5p). Interestingly, miR-26b-5p inhibitor prevented HSC activation inhibition caused by kaempferol. Further studies indicated that kaempferol inhibited Notch pathway via miR-26b-5p and Jag1, leading to HSC inactivation. Collectively, we demonstrate that kaempferol could inhibit HSC activation, at least in part, via miR-26b-5p-mediated Jag1 axis and Notch pathway. Kaempferol may serve as a promising drug in the application of treating liver fibrosis. [ABSTRACT FROM AUTHOR]

BACKGROUND: Long non-coding RNA-growth arrest specific transcript 5 (lncRNA-GAS5) plays a suppressive role in activated hepatic stellate cells (HSCs). LncRNAs could circulate in the blood in a cell-free form and serve as promising biomarkers for various human diseases. Herein, we investigated the feasibility of using serum GAS5 as a biomarker for liver fibrosis in chronic hepatitis B (CHB) patients and whether promoter methylation was responsible for GAS5 down-regulation. METHODS: Serum GAS5 levels were quantified using quantitative real-time PCR in CHB patients and healthy controls. GAS5 promoter methylation was examined in LX-2 cells and cirrhotic tissues. RESULTS: Compared with the sera from healthy controls, lower GAS5 levels were found in the sera from CHB patients. Receiver operating characteristic curve analysis indicated that serum GAS5 had a significant diagnostic value for liver fibrosis in CHB patients. Serum GAS5 negatively correlated with HAI scores as well as ALT values in CHB patients. GAS5 was additionally reduced in cirrhotic tissues, associated with its hypermethylation promoter. In LX-2 cells, transforming growth factor- β 1 treatment led to a reduction in GAS5 expression and an increase in promoter methylation. Hypermethylation of GAS5 was blocked down by DNA methyltransferase (DNMT) inhibitor and restored GAS5 inhibited HSC activation including proliferation and collagen production. Further studies confirmed that GAS5 methylation was mediated by DNMT1. CONCLUSION: We demonstrate that epigenetically-regulated serum GAS5 could serve as a potential biomarker in CHB patients. Loss of GAS5 is associated with DNMT1-mediated promoter methylation. [ABSTRACT FROM AUTHOR]

Liver fibrosis is primarily driven by the activation of hepatic stellate cells (HSCs), a process associated with ferroptosis. Ginsenoside Rb1 (GRb1), a major active component extracted from Panax ginseng, inhibits HSC activation. However, the potential role of GRb1 in mediating HSC ferroptosis remains unclear. This study examined the effect of GRb1 on liver fibrosis both in vivo and in vitro , using CCl 4 -induced liver fibrosis mouse model and primary HSCs, LX-2 cells. The findings revealed that GRb1 effectively inactivated HSCs in vitro , reducing alpha-smooth muscle actin (α-SMA) and type I collagen (Col1A1) levels. Moreover, GRb1 significantly alleviated CCl 4 -induced liver fibrosis in vivo. From a mechanistic standpoint, the ferroptosis pathway appeared to be central to the antifibrotic effects of GRb1. Specifically, GRb1 promoted HSC ferroptosis both in vivo and in vitro , characterized by increased glutathione depletion, malondialdehyde production, iron overload, and accumulation of reactive oxygen species (ROS). Intriguingly, GRb1 increased Beclin 1 (BECN1) levels and decreased the System Xc-key subunit SLC7A11. Further experiments showed that BECN1 silencing inhibited GRb1-induced effects on HSC ferroptosis and mitigated the reduction of SLC7A11 caused by GRb1. Moreover, BECN1 could directly interact with SLC7A11, initiating HSC ferroptosis. In conclusion, the suppression of BECN1 counteracted the effects of GRb1 on HSC inactivation both in vivo and in vitro. Overall, this study highlights the novel role of GRb1 in inducing HSC ferroptosis and promoting HSC inactivation, at least partly through its modulation of BECN1 and SLC7A11. [Display omitted] • GRb1 effectively inactivated HSC and alleviated CCl 4 -induced liver fibrosis. • GRb1 promotes HSC inactivation via ferroptosis pathway, and this is a first report. • BECN1 is the key medium in the ferroptosis process induced by GRb1. • GRb1-induced HSC ferroptosis is via BECN1/SLC7A11 axis. [ABSTRACT FROM AUTHOR]

Thymoquinone (TQ), the main active constituent of Nigella sativa seeds, has been shown to play a role in antioxidation, anti‐inflammation, and antitumor. Recent studies have demonstrated that TQ contributes to the suppression of liver fibrosis. Abnormal activated epithelial‐mesenchymal transition (EMT) promotes the activation of hepatic stellate cells (HSCs). However, whether the antifibrotic effects of TQ occur through inhibiting EMT is largely unknown. In this study, it was found that TQ ameliorated liver fibrosis and collagen accumulation in carbon tetrachloride (CCl4) mice. In vitro, TQ inhibited HSC activation including reduced proliferation, α‐smooth muscle actin, and collagen. In addition, TQ markedly suppressed the EMT process, with enhanced E‐cadherin and reduced desmin. Notably, snail family transcriptional repressor 1 (Snai1), the EMT master transcription factor, was obviously inhibited by TQ in vivo and in vitro. Further studies demonstrated that Snai1 was a target of microRNA‐30a (miR‐30a), which was upregulated by TQ. Interestingly, the effects of TQ on HSC activation and EMT were almost inhibited by miR‐30a inhibitor. Collectively, we demonstrate that TQ inhibits HSC activation, at least in part, via regulation of miR‐30a and Snai1. TQ upregulates miR‐30a expression, resulting in a reduced Snai1 level as well as EMT process inactivation, which contributes to the inhibition of HSC activation. TQ may be a potential therapeutic agent for liver fibrosis. [ABSTRACT FROM AUTHOR]

Circular RNAs (circRNAs), often dysregulated in a variety of human diseases, participate in the initiation and development of cancers. Recently, circMTO1 (a circRNA derived from MTO1 gene), identified as a tumor suppressor, has been shown to contribute to the suppression of hepatocellular carcinoma. The present study aimed to explore the clinical significance and roles of circMTO1 in liver fibrosis. Here, we found that serum circMTO1 was significantly down‐regulated in chronic hepatitis B (CHB) patients. Interestingly, serum circMTO1 was negatively correlated with fibrosis stages as well as HAI scores. Receiver operating characteristic curve analysis revealed that serum circMTO1 may serve as a diagnostic biomarker for liver fibrosis in CHB patients. Notably, overexpression of circMTO1 led to the suppression of transforming growth factor‐β1‐induced hepatic stellate cells (HSCs) activation. Bioinformatic analysis and luciferase activity assays indicated that circMTO1 was a target of mircoRNA‐17‐5p (miR‐17‐5p). Data from RNA pull‐down assay further confirmed that circMTO1 interacted with miR‐17‐5p. The inhibitory effects of circMTO1 on HSC activation were suppressed by miR‐17‐5p mimics. Further studies showed that Smad7 was a target of miR‐17‐5p. Moreover, circMTO1‐inhibited HSC activation was also blocked down by loss of Smad7. Taken together, we demonstrate that circMTO1 inhibits liver fibrosis via regulation of miR‐17‐5p and Smad7, and serum circMTO1 may be a novel promising biomarker of liver fibrosis. [ABSTRACT FROM AUTHOR]

Sustained liver fibrosis may lead to cirrhosis. Activated hepatic stellate cells (HSCs) are crucial for liver fibrosis development. Ferroptosis, a newly iron-dependent regulated cell death, has been demonstrated to be involved in HSC inactivation. Ginsenoside Rh2 (GRh2), a natural bioactive product derived from ginseng, has been shown to promote HSC inactivation. However, the effect of GRh2 on HSC ferroptosis remains unclear. We explored the effects of GRh2 on liver fibrosis in vivo and in vitro. RNA-sequence analysis was performed in HSCs after GRh2 treatment. The crosstalk between ferroptotic HSCs and macrophages was also explored. GRh2 alleviated liver fibrosis in vivo. In vitro , GRh2 reduced HSC proliferation and activation via ferroptosis, with increased intracellular iron, reactive oxygen species, malondialdehyde and glutathione depletion. The expression of SLC7A11, a negative regulator of ferroptosis, was obviously reduced by GRh2. Interestingly, interferon regulatory factor 1 (IRF1), a transcription factor, was predicted to bind the promoter region of SCL7A11. The interaction between IRF1 and SCL7A11 was further confirmed by the results of chromatin immunoprecipitation and luciferase reporter assays. Furthermore, loss of IRF1 led to an increase in SCL7A11, which contributed to the suppression of HSC ferroptosis and the enhancement of HSC activation in GRh2-treated HSCs. Further studies revealed that GRh2-induced HSC ferroptosis contributed to the inhibition of macrophage recruitment via regulation of inflammation-related genes. Moreover, GRh2 caused a reduction in liver inflammation in vivo. Collectively, GRh2 up-regulates IRF1 expression, resulting in the suppression of SLC7A11, which contributes to HSC ferroptosis and inactivation. GRh2 ameliorates liver fibrosis through enhancing HSC ferroptosis and inhibiting liver inflammation. GRh2 may be a promising drug for treating liver fibrosis. [Display omitted] [ABSTRACT FROM AUTHOR]

Serum long non-coding RNAs (lnc RNAs) are emerging as promising biomarkers for various human diseases. The aim of this study was to investigate the feasibility of using serum long intergenic non-coding RNA-p21 (linc RNA-p21) as a biomarker for chronic hepatitis B patients. Serum linc RNA-p21 levels were quantified using real-time PCR in 417 CHB patients and 363 healthy controls. The promoter methylation level of linc RNA-p21 was detected using bisulphite-sequencing analysis in primary hepatic stellate cells ( HSCs). Sera from hepatitis B-infected patients contained lower levels of linc RNA-p21 than sera from healthy controls. Serum linc RNA-p21 levels negatively correlated with stages of liver fibrosis in infected patients. Receiver operating characteristic ( ROC) curve analyses suggested that serum linc RNA-p21 had a significant diagnostic value for liver fibrosis in these patients. It yielded an area under the curve of ROC of 0.854 with 100% sensitivity and 70% specificity in discriminating liver fibrosis from healthy controls. There was additionally a negative correlation between serum linc RNA-p21 level and the markers of liver fibrosis including α- SMA and Col1A1. However, there was no correlation of serum linc RNA-p21 level with the markers of viral replication, liver inflammatory activity, and liver function. Notably, during primary HSCs culture, loss of linc RNA-p21 expression was associated with promoter methylation. Serum linc RNA-p21 could serve as a potential biomarker of liver fibrosis in CHB patients. Down-regulation of linc RNA-p21 in liver fibrosis may be associated with promoter methylation. [ABSTRACT FROM AUTHOR]

Background/Aims: Emerging evidence shows that microRNAs (miRNAs) play a crucial role in the regulation of activation, proliferation and apoptosis of hepatic stellate cells (HSCs). Previous studies have indicated that miR-125a-5p is correlated with hepatitis B virus replication and disease progression. However, little is known about the biological role and underlying mechanism of miR-125a-5p in liver fibrosis. Methods: We analyzed the level of miR-125a-5p in carbon tetrachloride-induced liver fibrosis and activated HSCs. We analyzed the effects of miR-125a-5p down-regulation on HSC activation and proliferation. We also analyzed the binding of miR-125a-5p to the 3'-untranslated region of factor inhibiting hypoxia-inducible factor 1 (FIH1) mRNA. Results: Up-regulation of miR-125a-5p was observed in the liver tissues of fibrotic mice and activated HSCs. Down-regulation of miR-125a-5p prevented the activation and proliferation of HSCs. FIH1, a negative modulator of hypoxia inducible factor 1, was confirmed to be a target of miR-125a-5p using the luciferase reporter assay. Further studies demonstrated that miR-125a-5p prompted the activation and proliferation of HSCs, at least in part, by down-regulating FIH1. Conclusion: Our findings shed new light on miRNAs as a promising therapeutic target in liver fibrosis. [ABSTRACT FROM AUTHOR]

Long non-coding RNAs are involved in various biological processes and diseases. The biological role of long intergenic non-coding RNA-p21 (lincRNA-p21) in liver fibrosis remains unknown before this study. In this study, we observed marked reduction of lincRNA-p21 expression in mice liver fibrosis models and human cirrhotic liver. Over-expression of lincRNA-p21 suppressed activation of hepatic stellate cells (HSCs) in vitro. Lentivirus-mediated lincRNA-p21 transfer into mice decreased the severity of liver fibrosis in vivo. Additionally, lincRNA-p21 reversed the activation of HSCs to their quiescent phenotype. The mRNA levels of lincRNA-p21 and p21 were positively correlated. Our results show that over-expression of lincRNA-p21 promotes up-regulation of p21 at both the mRNA and protein levels. Furthermore, lincRNA-p21 inhibited cell-cycle progression and proliferation of primary HSCs through enhancement of p21 expression. Compared with healthy subjects, serum lincRNA-p21 levels were significantly lower in patients with liver cirrhosis, especially those with decompensation. These findings collectively indicate that lincRNA-p21 is a mediator of HSC activation, supporting its utility as a novel therapeutic target for liver fibrosis. [ABSTRACT FROM AUTHOR]

In this study, we aimed at assessing a role of Alu -mediated p21 transcriptional regulator (APTR) in hepatofibrogenesis. APTR was upregulated in fibrotic liver samples and activated hepatic stellate cells (HSCs). Knockdown of APTR inhibited the activation of HSCs in vitro and mitigated the accumulation of collagen in vivo . Importantly, APTR silencing could abrogate TGF-β 1 -induced upregulation of α-SMA in HSCs. In addition, inhibition of cell cycle and cell proliferation by APTR knockdown was attenuated by p21 siRNA1 in primary HSCs. Finally, serum APTR levels were increased in patients with liver cirrhosis, indicating a potential biomarker for liver cirrhosis. Collectively, evidence is proposed for a new biological role of APTR in hepatofibrogenesis. [ABSTRACT FROM AUTHOR]

Background: In the development of liver fibrosis, activated hepatic stellate cells (HSCs) contribute to the synthesis and deposition of extracellular matrix (ECM) proteins. HSC activation is considered as a central driver of liver fibrosis. Recently, microRNAs (miRNAs) have been reported to act as key regulators in HSC activation.Purpose: Pinostilbene hydrate (PSH), a methylated derivative of resveratrol, has demonstrated anti-inflammatory, antioxidant and anti-tumour activities. However, the effects of PSH on HSC activation remain unclear.Methods: The effects of PSH on HSC activation were examined. Moreover, the roles of WNT inhibitory factor 1 (WIF1) and miR-17-5p in the effects of PSH on HSC activation were examined.Results: PSH induced a significant reduction in HSC proliferation. PSH also effectively inhibited HSC activation, with reduced α-SMA and collagen expression. Notably, it was found that Wnt/β-catenin signalling was involved in the effects of PSH on HSC activation. PSH resulted in Wnt/β-catenin signalling inactivation, with a reduction in TCF activity as well as β-catenin nuclear translocation. Further studies showed that PSH inhibited Wnt/β-catenin signalling via regulation of WIF1 and miR-17-5p. Reduced HSC activation caused by PSH could be restored by loss of WIF1 or miR-17-5p mimics. Luciferase reporter assays further confirmed that WIF1 was a target of miR-17-5p.Conclusion: PSH has a significant protective effect against HSC activation. In addition, we demonstrate that PSH enhances WIF1 expression and inhibits Wnt/β-catenin signalling via miR-17-5p, contributing to the suppression of HSC activation. [ABSTRACT FROM AUTHOR]

Background: Liquiritigenin (LQ), an aglycone of liquiritin in licorice, has demonstrated antioxidant, anti-inflammatory and anti-tumor activities. Previously, LQ was found to inhibit liver fibrosis progression.Purpose: Phosphatase and tensin homolog (PTEN) has been reported to act as a negative regulator of hepatic stellate cell (HSC) activation. However, the roles of PTEN in the effects of LQ on liver fibrosis have not been identified to date.Methods: The effects of LQ on liver fibrosis in carbon tetrachloride (CCl4) mice as well as primary HSCs were examined. Moreover, the roles of PTEN and microRNA-181b (miR-181b) in the effects of LQ on liver fibrosis were examined.Results: LQ markedly ameliorated CCl4-induced liver fibrosis, with a reduction in collagen deposition as well as α-SMA level. Moreover, LQ induced an increase in PTEN and effectively inhibited HSC activation including cell proliferation, α-SMA and collagen expression, which was similar with curcumin (a positive control). Notably, loss of PTEN blocked down the effects of LQ on HSC activation. PTEN was confirmed as a target of miR-181b and miR-181b-mediated PTEN was involved in the effects of LQ on liver fibrosis. LQ led to a significant reduction in miR-181b expression. LQ-inhibited HSC activation could be restored by over-expression of miR-181b. Further studies demonstrated that LQ down-regulated miR-181b level via Sp1. Collectively, we demonstrate that LQ inhibits liver fibrosis, at least in part, via regulation of miR-181b and PTEN.Conclusion: LQ down-regulates miR-181b level, leading to the restoration of PTEN expression, which contributes to the suppression of HSC activation. LQ may be a potential candidate drug against liver fibrosis. [ABSTRACT FROM AUTHOR]