Kidney function mediates the association of per- and poly-fluoroalkyl substances (PFAS) and heavy metals with hepatic fibrosis risk.

Heavy metals and per- and polyfluoroalkyl substances (PFAS) are significantly associated with the risk of hepatic fibrosis. However, the potential mediating effect of kidney function in the relationship between heavy metals, PFAS, and hepatic fibrosis risk remains unexplored. This research gap limits the development of hepatic fibrosis prevention and treatment strategies. To address this, this study conducts a cross-sectional analysis based on data from 10,870 participants in NHANES 2005-2018 to explore the relationship between heavy metals, PFAS, and the risk of hepatic fibrosis, as well as the mediating effect of kidney function. Participants with a Fibrosis-4 index <1.45 are defined as not having hepatic fibrosis in this study. Results from generalized linear regression models and weighted quantile sum regression models indicate that both individual and combined exposures to heavy metals and PFAS are positively associated with the risk of hepatic fibrosis. Nonlinear exposure-response functions suggest that there may be a threshold for the relationship between heavy metals (except mercury) and PFAS with the risk of hepatic fibrosis. Furthermore, heavy metals and PFAS increase the risk of kidney function impairment. After stratification by kidney function stage, the relationship between heavy metals (except lead) and proteinuria is not significant, while PFAS show a significant negative association with proteinuria. The decline in kidney function has a significant mediating effect in the relationship between heavy metals and PFAS and the risk of hepatic fibrosis, with mediation effect proportions all above 20%. The findings suggest that individual or combined exposure to heavy metals and PFAS does not increase the risk of hepatic fibrosis until a certain threshold is reached, and the mediating role of declining kidney function is very important. These results highlight the need to consider kidney function in the context of hepatic fibrosis risk assessment and management.

New peripherally-restricted CB1 receptor antagonists, PMG-505-010 and -013 ameliorate obesity-associated NAFLD and fibrosis.

The endocannabinoid system plays a crucial role in metabolic regulation, prompting the investigation of cannabinoid type 1 receptor (CB1R) antagonists for obesity and its complications like non-alcoholic fatty liver disease (NAFLD). Concerns over psychiatric side effects led to the development of peripheral CB1R antagonists that circumvent the blood-brain barrier (BBB). In this study, we synthesized PMG-505-010 and PMG-505-013 as peripherally restricted CB1 receptor antagonists by modifying rimonabant to minimize BBB penetration. Physicochemical analysis confirmed their reduced lipophilicity and increased polarity compared to rimonabant, indicating limited brain exposure. Molecular docking studies revealed similar binding modes to rimonabant at CB1R, characterized by robust hydrophobic interactions. Functionally, they acted as CB1R antagonists and inverse agonists, effectively reversing CP55,940-induced cAMP inhibition. In a murine model of obesity-related NAFLD, PMG-505-010 and -013 improved metabolic profiles, including fasting blood glucose levels and dyslipidemia. They also ameliorated hepatic injury, steatosis, and inflammation, evidenced by reduced liver enzymes, lipid peroxidation, hepatic lipid levels, and inflammatory cytokine levels. Notably, these compounds inhibited hepatic fibrosis by reducing extracellular matrix (ECM) deposition and altering fibrosis-related gene and protein expressions. In conclusion, PMG-505-010 and PMG-505-013 hold promise for treating obesity-related liver diseases, including NAFLD and fibrosis, through selective peripheral CB1R targeting, potentially avoiding CNS-related side effects seen with earlier CB1R antagonists.

Schistosoma japonicum infection-mediated downregulation of lncRNA Malat1 contributes to schistosomiasis hepatic fibrosis by the Malat1/miR-96/Smad7 pathway.

BACKGROUND: Schistosoma japonicum infection causes hepatic fibrosis, a primary cause of morbidity and mortality associated with the disease, and effective treatments are still lacking. Long non-coding RNAs (lncRNAs) have been implicated in the pathogenic process of various tissue fibroses. However, the role of lncRNAs in schistosomiasis hepatic fibrosis (HF) is poorly understood. Understanding the role of lncRNAs in schistosomiasis HF will enhance knowledge of disease processes and aid in the discovery of therapeutic targets and diagnostic biomarkers. METHODS: Differentially expressed lncRNA profiles in primary hepatic stellate cells (HSCs) of mice infected with S. japonicum were identified using high-throughput lncRNA sequencing. Primary HSCs were isolated from infected mice using collagenase digestion and density-gradient centrifugation, cultured in DMEM with 10% fetal bovine serum. Dual-luciferase reporter assays, nuclear cytoplasm fractionation and RIP assays were employed to assess the relationship between Malat1 and miRNA-96. Malat1 lentivirus and ASO-Malat1 were constructed for forced expression and downregulated expression of Malat1. The Malat1-KO mouse was constructed by CRISPR/Cas9 technology. Pathological features of the liver were evaluated by hematoxylin-eosin (HE), Masson's trichrome staining and immunohistochemistry (IHC). The expression levels of fibrosis-related genes were determined by quantitative real-time PCR (qRT-PCR) and Western blot. RESULTS: A total of 1561 differentially expressed lncRNAs were identified between infected and uninfected primary HSCs. Among the top altered lncRNAs, the downregulated Malat1 was observed in infected HSCs and verified by qPCR. Treatment of infected mice with praziquantel (PZQ) significantly increased the Malat1 expression. Elevated Malat1 expression in infected primary HSC reduced the expressions of profibrogenic genes, whereas Malat1 knockdown had the opposite effect. Moreover, Malat1 was found to interact with miR-96, a profibrotic miRNA, by targeting Smad7. Forced Malat1 expression reduced miR-96 levels in infected primary HSCs, attenuating fibrogenesis and showing negative correlation between Malat1 expression and the expression levels of miR-96 and profibrogenic genes α-SMA and Col1α1. Notably, in Malat1-KO mice, knockout of Malat1 aggravates schistosomiasis HF, while restored Malat1 expression in the infected HSCs reduced the expression of profibrogenic genes. CONCLUSIONS: We demonstrate that lncRNA is involved in regulation of schistosomiasis HF. Elevated lncRNA Malat1 expression in infected HSCs reduces fibrosis via the Malat1/miR-96/Smad7 pathway, thus providing a novel therapeutic target for schistosomiasis HF. Furthermore, Malat1 expression is sensitive to PZQ treatment, thus offering a potential biomarker for assessing the response to chemotherapy.

Protective effect of MP-40 mitigates BDL-induced hepatic fibrosis by inhibiting the NLRP3-mediated pyroptosis.

Background: Hepatic fibrosis and its associated consequences continue to pose a substantial global health challenge. Developing novel approaches to hepatic fibrosis management and prevention is critically necessary. Radix Paeoniae Alba (RPA) is widely used in Traditional Chinese Medicine (TCM) to treat various diseases. Our earlier research found that a bioactive component of RPA had a dose-dependent effect on anti-allergic asthma. RPA reduces allergic asthma by slowing the hepatic wind, according to "Treatise on Febrile Diseases". However, this bioactive fraction's pharmacological effects and mechanisms on the liver are unknown. Aim: This study examined the bioactive fraction MP-40, the methanol extract of RPA (MRPA), on bile duct ligation (BDL) for its anti-hepatic fibrosis activity and potential mechanisms. Methods: First, the effectiveness of MP-40 in treating BDL-induced hepatic fibrosis in mice and rats was evaluated through survival rates, ALT, AST HYP, and pathological changes. Molecular assays were performed using in vitro cultures of HSC-T6 activation. The expression of α-SMA and Collagen I evaluated fibro-tropic factors with HSC activation. Furthermore, the levels of pyroptosis were assessed by examining the expression of the pyroptosis-related proteins, including NLRP3, Cleaved Caspase-1, GSDMD-N, and 1L-1ß. Additionally, the effective constituents of MP-40 were identified by extraction, separation, and identification. Finally, PF and TGG, as the delegate compounds of MP-40, were tested to confirm their inhibition effects on HSC-T6 activation. Results: The findings demonstrated that MP-40 and MRPA could lower ALT, AST, and HYP levels, boost survival rates, and reduce liver damage in BDL mice and rats. Furthermore, MP-40 outperforms MRPA. MP-40 was proven to drastically diminish fibrotic α-SMA and Collagen I. The expression of pyroptosis-related proteins NLRP3, Cleaved Caspase-1, TGF-ß1, GSDMD-N, and 1L-1ß decreased. MP-40 inhibited the synthesis of pyroptosis-related proteins more effectively than MCC950 (an NLRP3-specific inhibitor). Monoterpene glycosides and tannins were shown to be the most potent MP-40 components. Finally, the delegate compounds MP-40, PF, and TGG were shown to have substantial inhibitory effects on HSC-T6 activation. Conclusion: The results proved that MP-40 alleviates BDL-induced cholestatic hepatic fibrosis by inhibiting NLRP3-mediated pyroptosis. PF and TGG play a role in treating BDL-induced cholestatic hepatic fibrosis in MP-40.

Epidemiological and transcriptome data identify association between iron overload and metabolic dysfunction-associated steatotic liver disease and hepatic fibrosis.

The primary objective of this study was to examine the association between iron overload (IO), metabolic dysfunction-associated steatotic liver disease (MASLD), and hepatic fibrosis. We hypothesized that there is a significant association. Data from the NHANES (2017-2020) were analyzed to explore IO's impact on MASLD and hepatic fibrosis in U.S. adults. We assessed serum ferritin, controlled attenuation parameter (CAP), liver stiffness measurement (LSM), and various covariates. Gene expression data were sourced from the FerrDb V2 and GEO databases. Differential gene expression analysis, Protein-Protein Interaction (PPI) Network construction, and Gene Ontology (GO) and KEGG pathway enrichment analyses were performed. The study verified the link between MASLD, hepatic fibrosis, and iron overload hub genes. This study of 5927 participants, averaging 46.78 years of age, revealed significant correlations between serum ferritin and CAP, LSM, after adjusting for covariates. Threshold effect analysis indicated nonlinear associations between serum ferritin and CAP, LSM, with distinct patterns observed by age and gender. Moreover, the area under the ROC curve for serum ferritin with MASLD and hepatic fibrosis was 0.8272 and 0.8376, respectively, demonstrating its performance in assessing these conditions. Additionally, molecular analyses identified potential hub genes associated with iron overload and MASLD, and hepatic fibrosis, revealing the underlying mechanisms. Our study findings reveal an association between iron overload, MASLD, and hepatic fibrosis. Additionally, the hub genes may be implicated in iron overload and subsequently contribute to the progression of MASLD and hepatic fibrosis. These findings support precision nutrition strategies.

The impact of traditional Chinese medicine and dietary compounds on modulating gut microbiota in hepatic fibrosis: A review.

Traditional Chinese medicine (TCM) and dietary compounds have a profound influence on the regulation of gut microbiota (GM) in hepatic fibrosis (HF). Certain substances found in both food and herbs that are edible and medicinal, such as dietary fiber, polyphenols, and polysaccharides, can generate beneficial metabolites like short-chain fatty acids (SCFAs), bile acids (BAs), and tryptophan (Trp). These compounds contribute to regulate the GM, reduce levels of endotoxins in the liver, and alleviate fibrosis and inflammation in the liver. Furthermore, they enhance the composition and functionality of GM, promoting the growth of beneficial bacteria while inhibiting the proliferation of harmful bacteria. These mechanisms mitigate the inflammatory response in the intestines and maintain the integrity of the intestinal barrier. The purpose of this review is to analyze how the GM regulates the pathogenesis of HF, evaluate the regulatory effect of TCM and dietary compounds on the intestinal microflora, with a particular emphasis on modulating flora structure, enhancing gut barrier function, and addressing associated pathogenic factors, thereby provide new insights for the treatment of HF.

Relationships between tumor CD147 expression, tumor-infiltrating lymphocytes, and oncostatin M in hepatocellular carcinoma.

In hepatocellular carcinoma (HCC), CD147 expression contributes to tumor malignancy; however, its relationship with the tumor-immune microenvironment (TIME) remains unclear. This study aimed to elucidate the clinicopathological characteristics associated with CD147 expression in HCC and investigate its association with the TIME, specifically its association with tumor-infiltrating lymphocytes (TILs) and oncostatin M (OSM). Using 397 HCC specimens from patients undergoing curative-intent resection, we assessed CD147 expression in tumor cells and quantified OSM-positive cells and various TILs (CD8+, CD4+, FOXP3+, and CD20+ cells) in the TIME. Using tissue microarrays, these assessments were performed through immunohistochemical analysis. We investigated the associations between CD147 expression status, the density of OSM-positive cells, and the densities of various TILs. High CD147 expression, found in 332 specimens (83.6%), was associated with advanced clinical stage (P = 0.029), fibrosis (P = 0.036), and higher densities of FOXP3+ cells (P = 0.0039), CD4+ cells (P = 0.0012), and OSM-positive cells (P = 0.0017). In CD147-high tumors, OSM-positive cell density was associated with all assessed TIL subsets (CD8+, CD4+, FOXP3+, and CD20+ cells; all Ps < 0.001), whereas in CD147-low tumors, OSM-positive cell density was associated only with FOXP3+ cells (P = 0.0004). In HCC, CD147 expression is associated with an immunosuppressive TIME, characterized by increased FOXP3+ regulatory T cells and a correlation with OSM-positive cells. These results elucidate the potential mechanisms through which CD147 facilitates tumor-immune evasion, suggesting the CD147 - OSM axis as a promising target for therapeutic intervention in HCC.

The processing mechanism of vinegar-processed Curcumae Rhizome enhances anti hepatic fibrotic effects through regulation of PI3K/Akt/mTOR signaling pathway.

BACKGROUND: Hepatic fibrosis, a chronic pathological condition resulting from various forms of persistent liver injury, in the later stage, it can evolve into cirrhosis and even liver cancer. Curcumae Rhizoma (CR), traditionally recognized for its properties in line qi break blood, eliminate accumulation and relieve pain. According to traditional Chinese medicine (TCM) principles, vinegar-processing enhances CR's ability to enter the liver meridian and act on the blood level, potentially augmenting its therapeutic effects on hepatic diseases. Therefore, vinegar-processed Curcumae Rhizoma (VCR) is frequently employed in treating liver fibrosis and related hepatic conditions. However, the underlying mechanisms of vinegar processing in enhancing its therapeutic efficacy remain unclear. METHODS: The anti-liver fibrosis effects of CR and VCR were verified at individual and cellular levels. Subsequently, HPLC-Q-TOFMS and pharmacokinetic analysis were utilized to elucidate the potential bioactive substances underlying the enhanced anti-fibrotic efficacy of VCR. Building upon these findings, network pharmacology and metabolomics were integrated to screen for key effect components and regulatory pathways. Finally, the mechanisms of action were further analyzed and validated at the tissue and cellular levels through Western blotting (WB) and molecular docking studies. RESULTS: Both CR and VCR exhibited therapeutic effects against hepatic fibrosis, with VCR demonstrating enhanced efficacy after vinegar processing. 6 sesquiterpenes including furanodiene and curdione, showed significant alterations in plasma exposure and hepatic distribution post-processing. VCR significantly improved pathological liver conditions, lipid accumulation, and fibrosis severity. Additionally, VCR markedly reduced the expression of α-SMA in the liver and attenuated the elevations in liver function markers such as ALT and AST. Combined network pharmacology, metabolomics, and hepatic tissue WB analysis revealed that the reduced phosphorylation of the PI3K/Akt/mTOR pathway is a critical mechanism in VCR's anti-fibrotic effects. Experiments on LX-2 cells demonstrated that four sesquiterpenes, including furanodiene and curdione, effectively inhibited the proliferation of activated hepatic stellate cells (HSCs). Furanodiene, in particular, promoted apoptosis in activated HSCs by reducing phosphorylation levels of the PI3K/Akt/mTOR pathway proteins, increasing BAX expression, and activating downstream caspase-3 to achieve the effect of anti-liver fibrosis. CONCLUSION: Vinegar-processing significantly increases the plasma exposure and hepatic distribution of components such as furanodiene in VCR, enhancing anti-fibrotic efficacy by downregulating the phosphorylation levels of the PI3K/Akt/mTOR pathway and promoting HSC apoptosis. This study provides a comprehensive explanation of the vinegar-processing mechanism and its role in enhancing the anti-fibrotic effects of VCR, offering insights for its clinical application in liver fibrosis treatment and reference for the mechanistic study of other vinegar-processed herbal medicines.

Bie Jia Jian pill ameliorates BDL-induced cholestatic hepatic fibrosis in rats by regulating intestinal microbial composition and TMAO-mediated PI3K/AKT signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: As a compound of traditional Chinese medicine (TCM), Bie Jia Jian pill (BJJP) is extensively used to treat the clinical chronic liver disease. Nevertheless, the specific mechanism through which BJJP affects hepatic fibrosis (HF) remains unknown. AIM OF THE STUDY: To explore the role and potential mechanism of BJJP involved in treating HF. MATERIALS AND METHODS: HF model of Sprague-Dawley (SD) rats was induced by a bile duct ligation (BDL). The function of BJJP involved in the intestinal microbiota (IM) and its metabolites in BDL-induced HF rats were explored through the 16S rRNA sequencing and untargeted metabolomics technologies. Network pharmacology was used to forecast mechanism underlying BJJP's anti-HF effects, which were validated in BDL-induced rats and trimethylamine N-oxide (TMAO)-induced LX-2 and HSC-T6 cells. RESULTS: BJJP effectively ameliorated pathological liver damage, inflammation, and fibrosis of the BDL-induced HF rats. BJJP regulated IM diversity and composition and interfered with trimethylamine (TMA)-flavin monooxygenase 3 (FMO3)-TMAO process. In vitro, BJJP significantly inhibited the TMAO-induced activation of hepatic stellate cells (HSCs) (rat HSC cell line, HSC-T6; human HSC cell line, LX-2). Network pharmacology results demonstrated that PI3K/AKT signal pathway is crucially involved in BJJP treatment of HF. Further research revealed that BJJP inhibited the PI3K/AKT signal pathway in BDL-induced HF rats. Moreover, TMAO activated the PI3K/AKT pathway, whereas BJJP suppressed TMAO-induced activation. Subsequent intervention with 740Y-P (the PI3K agonist) successfully neutralized the repression effect on PI3K/AKT signal pathway by BJJP. CONCLUSION: These results clearly show that BJJP attenuates HF by regulating the IM, as well as inhibiting PI3K/AKT pathway mediated by TMAO.

Exogenous S1P via S1P receptor 2 induces CTGF expression through Src-RhoA-ROCK-YAP pathway in hepatic stellate cells.

BACKGROUND: Hepatic fibrosis, a prevalent chronic liver condition, involves excessive extracellular matrix production associated with aberrant wound healing. Hepatic stellate cells (HSCs) play a pivotal role in liver fibrosis, activated by inflammatory factors such as sphingosine 1-phosphate (S1P). Despite S1P's involvement in fibrosis, its specific role and downstream pathway in HSCs remain controversial. METHODS: In this study, we investigated the regulatory role of S1P/S1P receptor (S1PR) in Hippo-YAP activation in both LX-2 cell lines and primary HSCs. Real-time PCR, western blot, pharmacological inhibitors, siRNAs, and Rho activity assays were adopted to address the molecular mechanisms of S1P mediated YAP activation. RESULTS: Serum and exogenous S1P significantly increased the expression of YAP target genes in HSCs. Pharmacologic inhibitors and siRNA-mediated knockdowns of S1P receptors showed S1P receptor 2 (S1PR2) as the primary mediator for S1P-induced CTGF expression in HSCs. Results using siRNA-mediated knockdown, Verteporfin, and Phospho-Tag immunoblots showed that S1P-S1PR2 signaling effectively suppressed the Hippo kinases cascade, thereby activating YAP. Furthermore, S1P increased RhoA activities in cells and ROCK inhibitors effectively blocked CTGF induction. Cytoskeletal-perturbing reagents were shown to greatly modulate CTGF induction, suggesting the important role of actin cytoskeleton in S1P-induced YAP activation. Exogeneous S1P treatment was enough to increase the expression of COL1A1 and α-SMA, that were blocked by YAP specific inhibitor. CONCLUSIONS: Our data demonstrate that S1P/S1PR2-Src-RhoA-ROCK axis leads to Hippo-YAP activation, resulting in the up-regulation of CTGF, COL1A1 and α-SMA expression in HSCs. Therefore, S1PR2 may represent a potential therapeutic target for hepatic fibrosis.

Monoexponential and advanced diffusion-weighted imaging for hepatic fibrosis staging based on high inter-examiner reliability.

OBJECTIVES: To determine the diagnostic efficiencies of multiple diffusion-weighted imaging (DWI) techniques for hepatic fibrosis (HF) staging under the premise of high inter-examiner reliability. METHODS: Participants with biopsy-confirmed HF were recruited and divided into the early HF (EHF) and advanced HF (AHF) groups; healthy volunteers (HVs) served as controls. Two examiners analyzed intravoxel incoherent motion (IVIM) using the IVIM-DWI and diffusion kurtosis imaging (DKI) models. Intravoxel incoherent motion-DWI, DKI, and diffusion tensor imaging parameters with intraclass correlation coefficients (ICCs) of ≥0.6 were used to create regression models: HVs vs. EHF and EHF vs. AHF. RESULTS: We enrolled 48 HVs, 59 EHF patients, and 38 AHF patients. Mean, radial, and axial kurtosis; fractional anisotropy; mean, radial, and axial diffusivity; and α exhibited excellent reliability (ICCs: 0.80-0.98). Fractional anisotropy of kurtosis, f, and apparent diffusion coefficient showed good reliability (ICCs: 0.69-0.92). The real (0.58-0.67), pseudo- (0.27-0.76), and distributed diffusion coefficients (0.58-0.67) showed low reliability. In the HVs versus (vs.) EHF model, α (p=0.008) and ADC (p=0.011) presented statistical differences (area under curve [AUC]: 0.710). In the EHF vs. AHF model, α (p=0.04) and distributed diffusion coefficient (p=0.02) presented significant differences (AUC: 0.758). CONCLUSION: Under the premise of high inter-examiner reliability, DWI and IVIM-derived stretched-exponential model parameters may help stage HF.

Exploring hepatic fibrosis screening via deep learning analysis of tongue images.

Background: Tongue inspection, an essential diagnostic method in Traditional Chinese Medicine (TCM), has the potential for early-stage disease screening. This study aimed to evaluate the effectiveness of deep learning-based analysis of tongue images for hepatic fibrosis screening. Methods: A total of 1083 tongue images were collected from 741 patients and divided into training, validation, and test sets. DenseNet-201, a convolutional neural network, was employed to train the AI model using these tongue images. The predictive performance of AI was assessed and compared with that of FIB-4, using real-time two-dimensional shear wave elastography as the reference standard. Results: The proposed AI model achieved an accuracy of 0.845 (95% CI: 0.79-0.90) and 0.814 (95% CI: 0.76-0.87) in the validation and test sets, respectively, with negative predictive values (NPVs) exceeding 90% in both sets. The AI model outperformed FIB-4 in all aspects, and when combined with FIB-4, the NPV reached 94.4%. Conclusion: Tongue inspection, with the assistance of AI, could serve as a first-line screening method for hepatic fibrosis.

Advancements in Plant-Based Therapeutics for Hepatic Fibrosis: Molecular Mechanisms and Nanoparticulate Drug Delivery Systems.

Chronic liver injuries often lead to hepatic fibrosis, a condition characterized by excessive extracellular matrix accumulation and abnormal connective tissue hyperplasia. Without effective treatment, hepatic fibrosis can progress to cirrhosis or hepatocellular carcinoma. Current treatments, including liver transplantation, are limited by donor shortages and high costs. As such, there is an urgent need for effective therapeutic strategies. This review focuses on the potential of plant-based therapeutics, particularly polyphenols, phenolic acids, and flavonoids, in treating hepatic fibrosis. These compounds have demonstrated anti-fibrotic activities through various signaling pathways, including TGF-ß/Smad, AMPK/mTOR, Wnt/ß-catenin, NF-κB, PI3K/AKT/mTOR, and hedgehog pathways. Additionally, this review highlights the advancements in nanoparticulate drug delivery systems that enhance the pharmacokinetics, bioavailability, and therapeutic efficacy of these bioactive compounds. Methodologically, this review synthesizes findings from recent studies, providing a comprehensive analysis of the mechanisms and benefits of these plant-based treatments. The integration of novel drug delivery systems with plant-based therapeutics holds significant promise for developing effective treatments for hepatic fibrosis.

Carthamus tinctorius L. inhibits hepatic fibrosis and hepatic stellate cell activation by targeting the PI3K/Akt/mTOR pathway.

Hepatic fibrosis (HF) is a process that occurs during the progression of several chronic liver diseases, for which there is a lack of effective treatment options. Carthamus tinctorius L. (CTL) is often used in Chinese or Mongolian medicine to treat liver diseases. However, its mechanism of action remains unclear. In the present study, CTL was used to treat rats with CCl4­induced HF. The histopathological, biochemical and HF markers of the livers of the rats were analyzed, and CTL­infused serum was used to treat hepatic stellate cells (HSCs) in order to detect the relevant markers of HSC activation. Protein expression pathways were detected both in vitro and in vivo. Histopathological results showed that CTL significantly improved CCl4­induced liver injury, reduced aspartate aminotransferase and alanine aminotransferase levels, promoted E­cadherin expression, and decreased α­smooth muscle actin (SMA), SOX9, collagen I and hydroxyproline expression. Moreover, CTL­infused serum was found to decrease α­SMA and collagen I expression in HSCs. Further studies showed that CTL inhibited the activity of the PI3K/Akt/mTOR pathway in the rat livers. Following the administration of the PI3K agonist 740Y­P to HSCs, the inhibitory effect of CTL on the PI3K/Akt//mTOR pathway was blocked. These results suggested that CTL can inhibit HF and HSC activation by inhibiting the PI3K/Akt/mTOR pathway.

Cpd861 Targeting BCL2 to Alleviate Hepatic Fibrosis: Network Pharmacology, Mendelian Randomization, and Molecular Docking Mechanisms.

BACKGROUND: Compound 861 (Cpd861) is a traditional Chinese herbal compound for the treatment of hepatic fibrosis (HF). In the current investigation, Cpd861 has been demonstrated to have an underlying molecular mechanism and material foundation for the treatment of HF through network pharmacology, Mendelian randomization (MR), and molecular docking. METHODS: Public databases were consulted for Cpd861 constituents and HF targets. Protein-protein interactions (PPIs) were established using STRING software, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. To elucidate the causal relationship between potential targets and liver injury, MR was used as a methodological tool. Finally, a molecular docking analysis was conducted between the active compound and the key target. RESULTS: We obtained 174 active ingredients and 113 intersecting genes. Through the PPI network, high-degree targets were identified, namely CTNNB1, ESR1, FOS, MDM2, CCND1, TP53, RELA, and BCL2. As shown by GO and KEGG pathway enrichment analyses, Cpd861 functions through xenobiotic stimulus and oxidative stress-related genes, as well as the PI3K-AKT and non-alcoholic fatty liver disease (NAFLD) signaling pathways. The results of MR showed that MDM2 and BCL2 had a causal relationship with liver injury. Molecular docking results showed that several active compounds in Cpd861 were stably bound to BCL2. CONCLUSION: This study made predictions regarding the efficacious components, as well as potential targets and pathways of Cpd861 in the therapy of HF. This will open up a new perspective for further investigation of the molecular mechanism of Cpd861 in the treatment of HF.

A proteomics-based study of the mechanism of oxymatrine to ameliorate hepatic fibrosis in mice.

OBJECTIVE: This study investigated the protective effect of oxymatrine (OMT) on carbon tetrachloride (CCl4)-induced hepatic fibrosis in mice and explored its possible targets and signaling pathways. METHODS: Male BALB/c mice were randomly divided into blank control, model, positive drug (silymarin), and OMT administration groups, respectively, with 10 mice in each group. Hepatic fibrosis was induced in mice using CCl4 and the corresponding drug intervention was given. After the final administration, ultrasonography tests, blood tests, and analysis of liver differential proteins using tandem mass tag labeling and liquid chromatography-mass spectrometry were performed. RESULTS: OMT intervention ameliorated CCl4-induced hepatic fibrosis in mice, significantly reduced serum alanine aminotransferase and aspartate aminotransferase levels, down-regulated the expression of fibrosis factors, such as type IV collagen IV, laminin, type III procollagen III, and alpha-smooth muscle actin, and improved liver function. The results of the proteomic analysis showed that the intervention of OMT significantly down-regulated 130 out of 440 up-regulated proteins and up-regulated 70 out of 294 down-regulated proteins, primarily involving the transient receptor potential (TRP) signaling pathway, the peroxisome proliferator-activated receptors (PPAR) signaling pathway, and the metabolic pathway of arachidonic acid. The main differential proteins involved were Cyp2c37, SCP-2, and Tbxas1. In addition, OMT intervention significantly reversed the expression of sterol carrier protein-2 (SCP2) and upregulated the expression of peroxisome proliferator-activated receptor gamma, Cyp2c37, and transient receptor potential cation channel subfamily V member 1 proteins. CONCLUSION: OMT inhibited the proliferative capacity of hepatic stellate cells, induced apoptotic properties, and suppressed the development of fibrosis by elevating Cyp2c37/TRP signaling axis activity and upregulating PPAR pathway activity by inhibiting SCP2.

[Analysis of core functional components in Yinchenhao Decoction and their pathways for treating liver fibrosis].

OBJECTIVE: To analyze the core functional component groups (CFCG) in Yinchenhao Decoction (YCHD) and their possible pathways for treating hepatic fibrosis based on network pharmacology. METHODS: PPI data were extracted from DisGeNET, Genecards, CMGRN and PTHGRN to construct a weighted network using Cytoscape 3.9.1. The data of the chemical components in YCHD were obtained from Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), and the potential active components and targets were selected using PreADMET Web server and SwissTargetPrediction. A fusion model was constructed to obtain the functional effect space and evaluate the effective proteins to identify the CFCG followed by GO and KEGG pathway enrichment analyses for all the targets. In cultured human hepatic stellate cells (LX-2 cells), the cytotoxicity of different compounds in YCHD was tested using CCK-8 assay; the effects of these compounds on collagen α1 (Col1a1) mRNA expression and the pathways in 20 ng/mL TGF-ß1-stimulated cells were analyzed using RT-qPCR and Western blotting. RESULTS: A total of 1005 pathogenic genes, 226 potential active components and 1529 potential targets in YCHD and 52 potential targets of CFCG were obtained. Benzyl acetate, vanillic acid, clorius, polydatin, lauric acid and ferulic acid were selected for CCK-8 verification, and they all showed minimal cytotoxicity below the concentration of 200 µmol/L. Clorius, polydatin, lauric acid and ferulic acid all effectively inhibited TGF-ß1-induced LX-2 cell activation. At the concentration of 200 µmol/L, all these 4 components inhibited PI3K, p-PI3K, AKT, p-AKT, ERK, p-ERK, P38 MAPK and p-P38 MAPK expressions in TGF-ß1-induced LX-2 cells. CONCLUSION: The therapeutic effect of YCHD on hepatic fibrosis is probably mediated by its core functional components including benzyl acetate, vanillic acid, clorius, polydatin, lauric acid and ferulic acid, which inhibit the PI3K-AKT and MAPK pathways in hepatic stellate cells.

Catalpol ameliorates liver fibrosis via inhibiting aerobic glycolysis by EphA2/FAK/Src signaling pathway.

BACKGROUND: Hepatic fibrosis is a pathological process in a variety of acute or chronic liver injuries. Catalpol (CAT), an iridoid glycoside found in Rehmannia glutinosa, has several pharmacological properties, including anti-inflammatory, antidiabetic and anti-fibrotic effects. Nevertheless, there is currently no report on whether CAT regulates the aerobic glycolysis of hepatic stellate cells (HSCs) to inhibit liver fibrosis. OBJECTIVE: This study aimed to investigate the protective effects of CAT on hepatic fibrosis and elucidate its underlying mechanisms. METHODS: To explore whether CAT improved liver fibrosis in vivo and in vitro, hepatic fibrosis was induced to mice by intraperitoneally injecting carbon tetrachloride (CCl4). Additionally, LX-2 cells were stimulated with transforming growth factor-ß (TGF-ß) to simulate fibrosis in vitro. Serum markers of liver injury were examined by using an automatic biochemical analyzer. Histopathological staining, Immunofluorescence (IF) staining, Western blot (WB) analysis, co-immunoprecipitation (Co-IP), drug affinity responsive target stability (DARTS), cellular thermal shift assay (CETSA), etc. were employed to identify the targeting between CAT and EphA2 and detect the expression of aerobic glycolysis related proteins, fiber markers and signaling pathways that are responsible for CAT's anti-fibrotic effects of CAT. RESULTS: Results showed that CAT significantly inhibited hepatic injury, fibrogenesis and inflammation in mice treated with CCl4. This was demonstrated by the enhancement of fibrosis markers, liver function indices, and histopathology. In addition, CAT significantly inhibited the activation of HSCs in TGF-ß-induced LX-2 cells, as indicated by decreased proliferation, migration, and expression of collagen I and a-SMA. The study results also suggested that CAT may exert anti-fibrotic effects by inhibiting glycolysis in activated HSCs and in CCl4-treated mice. Mechanistically, CAT directly targets Ephrin type-A receptor 2 (EphA2) to reduce binding with focal adhesion kinases (FAK) and significantly inhibits the FAK/Src pathway. In addition, the pharmacological inhibition of EphA2 cannot further increase the therapeutic effects of CAT on liver fibrosis in vivo and in vitro. CONCLUSION: The study findings generally demonstrated that CAT presented a novel therapeutic method to treat hepatic fibrosis; this method which inhibits the aerobic glycolysis of activated HSCs through the EphA2/FAK/Src signaling pathway.

Hepatoprotective action mechanism and quantification of soyasaponin Bb in Abri Herba by HPLC and network pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: The herb of Abrus cantoniensis Hance (AC) is an important Traditional Chinese Medicine (TCM) and is also used as an herbal tea with hepatoprotective action. Soyasaponin Bb is one of the pharmacodynamic substances of AC for the herb's effective pharmacological activity. This study aims to investigate the anti-fibrotic and hepatoprotective activities of soyasaponin Bb in vivo and in vitro experiments, mechanism by network pharmacology and quantification by HPLC. MATERIALS AND METHODS: High-performance liquid chromatography (HPLC) was applied to evaluate the quality of the herb and determine the contents of soyasaponin Bb from different sources and parts of the AC. In vivo experiments were conducted to induce an acute liver injury model by injecting CCl4 into mice, and an in vitro hepatic fibrosis model was established by cultivating LX-2 cells with TGF-ß1. These models were used to explore the anti-fibrotic and hepatoprotective effects of soyasaponin Bb and its underlying mechanisms. In addition, the potential target genes corresponding to soyasaponin Bb were identified using the Swiss Target Prediction database through network pharmacology methods. Meanwhile, hepatic fibrosis targets were screened using the GeneCards, TTD, and OMIM disease databases. The STING database was used to construct the protein-protein interaction (PPI) network of soyasaponin Bb-hepatic fibrosis. The soyasaponin Bb-hepatic fibrosis disease target-pathway network was constructed using Cytoscape 3.9.1 software. Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed to enrich and analyze the common targets of the drug and the disease, aiming to identify the potential targets and pathways involved in the anti-fibrotic and hepatoprotective effects of soyasaponin Bb. RESULTS: The content of soyasaponin Bb varied across different sources, with the roots containing the highest concentration, up to 0.2480%. In vivo experiments showed that soyasaponin Bb had a protective effect against CCl4-induced acute liver injury, effectively inhibiting the increase in ALT and AST levels and slowing down the hepatocyte inflammatory damage caused by CCl4. Soyasaponin Bb also down-regulated MDA levels and up-regulated SOD levels, indicating a certain antioxidant capacity. In vitro cell experiments showed that soyasaponin Bb could effectively inhibit the proliferation of HSC-LX2 cells induced by TGF-ß1 by regulating the TGF-ß1/α-SMA pathway, significantly down-regulate the protein expression of TGF-ß1 and α-SMA, while also reducing the levels of inflammatory cytokines IL-6 and IL-1ß. Further network pharmacology analysis suggested that soyasaponin Bb can exert anti-fibrosis activity by regulating the IBD signaling pathway, Th17 signaling pathway, Hepatitis B signaling pathway, and JAK-STAT signaling pathway. CONCLUSION: Soyasaponin Bb is primarily distributed in the root of AC, and it has a strong protective effect against CCl4-induced acute liver injury. It can reduce the level of inflammatory factors, relieve inflammation, and exert anti-fibrotic activity by regulating the TGF-ß1/α-SMA pathway. Its potential anti-hepatic fibrosis mechanism has been investigated through network pharmacology.

A New Target for Hepatic Fibrosis Prevention and Treatment: The Warburg Effect.

Hepatic fibrosis is a major public health problem that endangers human wellbeing. In recent years, a number of studies have revealed the important impact of metabolic reprogramming on the occurrence and development of hepatic fibrosis. Among them, the Warburg effect, as an intracellular glucose metabolism reprogramming, can promote the occurrence and development of hepatic fibrosis by promoting the activation of hepatic stellate cells (HSCs) and inducing the polarization of liver macrophages (KC). Understanding the Warburg effect and its important role in the progression of hepatic fibrosis will assist in developing new strategies for the prevention and treatment of hepatic fibrosis. This review focuses on the Warburg effect and the specific mechanism by which it affects the progression of hepatic fibrosis by regulating HSCs activation and KC polarization. In addition, we also summarize and discuss the related experimental drugs and their mechanisms that inhibit the Warburg effect by targeting key proteins of glycolysis in order to improve hepatic fibrosis in the hope of providing more effective strategies for the clinical treatment of hepatic fibrosis.