Ba-Qi-Rougan formula alleviates hepatic fibrosis by suppressing hepatic stellate cell activation via the MSMP/CCR2/PI3K pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: The Ba-Qi-Rougan formula (BQRGF) is a traditional and effective compound prescription from Traditional Chinese Medicine (TCM) utilized in treating hepatic fibrosis (HF). AIM OF THE STUDY: We aimed to evaluate the therapeutic efficacy of BQRGF on HF and explore the underlying mechanisms of action. MATERIALS AND METHODS: UPLC-Q-TOF-MS technology was employed to identify the material basis of BQRGF. Mice with carbon tetrachloride (CCl4)-induced HF received BQRGF at three doses (3.87, 7.74, and 15.48 g/kg per day). We examined serum and liver biochemical indicators and liver histology to assess the therapeutic impact. Primary mouse cells were isolated and utilized for experimental analysis. MSMP expression levels were examined in vitro and in vivo experimental models, including human and mouse tissue. Furthermore, lentivirus and small interfering RNA (siRNA) transfections were employed to manipulate microseminoprotein (MSMP) expression in LO2 cells (human normal liver cells). These manipulated LO2 cells were then co-cultured with LX2 human hepatic stellate cells (HSCs). Through the modulation of MSMP expression in co-cultured cells, administering recombinant MSMP (rMSMP) with or without BQRGF-medicated serum, and using specific pathway inhibitors or agonists in LX2 cells, we elucidated the underlying mechanisms. RESULTS: A total of 48 compounds were identified from BQRGF, with 12 compounds being absorbed into the bloodstream and 9 compounds being absorbed into the liver. Four weeks of BQRGF treatment in the HF mouse model led to significant improvements in biochemical and molecular assays and histopathology, particularly in the medium and high-dose groups. These improvements included a reduction in the level of liver injury and fibrosis-related factors. MSMP levels were elevated in human and mouse fibrotic liver tissues, and this increase was mitigated in HF mice treated with BQRGF. Moreover, primary cells and co-culture studies revealed that BQRGF reduced MSMP expression, decreased the expression of the hepatic stellate cell (HSC) activation markers, and suppressed critical phosphorylated protein levels in the CCR2/PI3K/AKT pathway. These findings were further validated using CCR2/PI3K/AKT signaling inhibitors and agonists in MSMP-activated LX2 cells. CONCLUSIONS: Collectively, our results suggest that BQRGF combats HF by diminishing MSMP levels and inhibiting MSMP-induced HSC activation through the CCR2/PI3K/AKT pathway.

Plant-derived flavonoids are a potential source of drugs for the treatment of liver fibrosis.

Liver fibrosis is a dynamic pathological process that can be triggered by any chronic liver injury. If left unaddressed, it will inevitably progress to the severe outcomes of liver cirrhosis or even hepatocellular carcinoma. In the past few years, the prevalence and fatality of hepatic fibrosis have been steadily rising on a global scale. As a result of its intricate pathogenesis, the quest for pharmacological interventions targeting liver fibrosis has remained a formidable challenge. Currently, no pharmaceuticals are exhibiting substantial clinical efficacy in the management of hepatic fibrosis. Hence, it is of utmost importance to expedite the development of novel therapeutics for the treatment of this condition. Various research studies have revealed the ability of different natural flavonoid compounds to alleviate or reverse hepatic fibrosis through a range of mechanisms, which are related to the regulation of liver inflammation, oxidative stress, synthesis and secretion of fibrosis-related factors, hepatic stellate cells activation, and proliferation, and extracellular matrix synthesis and degradation by these compounds. This review summarizes the progress of research on different sources of natural flavonoids with inhibitory effects on liver fibrosis over the last decades. The anti-fibrotic effects of natural flavonoids have been increasingly studied, making them a potential source of drugs for the treatment of liver fibrosis due to their good efficacy and biosafety.

Danhongqing formula alleviates cholestatic liver fibrosis by downregulating long non-coding RNA H19 derived from cholangiocytes and inhibiting hepatic stellate cell activation.

OBJECTIVE: This study explores the mechanism of action of Danhongqing formula (DHQ), a compound-based Chinese medicine formula, in the treatment of cholestatic liver fibrosis. METHODS: In vivo experiments were conducted using 8-week-old multidrug resistance protein 2 knockout (Mdr2-/-) mice as an animal model of cholestatic liver fibrosis. DHQ was administered orally for 8 weeks, and its impact on cholestatic liver fibrosis was evaluated by assessing liver function, liver histopathology, and the expression of liver fibrosis-related proteins. Real-time polymerase chain reaction, Western blot, immunohistochemistry and other methods were used to observe the effects of DHQ on long non-coding RNA H19 (H19) and signal transducer and activator of transcription 3 (STAT3) phosphorylation in the liver tissue of Mdr2-/- mice. In addition, cholangiocytes and hepatic stellate cells (HSCs) were cultured in vitro to measure the effects of bile acids on cholangiocyte injury and H19 expression. Cholangiocytes overexpressing H19 were constructed, and a conditioned medium containing H19 was collected to measure its effects on STAT3 protein expression and cell activation. The intervention effect of DHQ on these processes was also investigated. HSCs overexpressing H19 were constructed to measure the impact of H19 on cell activation and assess the intervention effect of DHQ. RESULTS: DHQ alleviated liver injury, ductular reaction, and fibrosis in Mdr2-/- mice, and inhibited H19 expression, STAT3 expression and STAT3 phosphorylation. This formula also reduced hydrophobic bile acid-induced cholangiocyte injury and the upregulation of H19, inhibited the activation of HSCs induced by cholangiocyte-derived conditioned medium, and decreased the expression of activation markers in HSCs. The overexpression of H19 in a human HSC line confirmed that H19 promoted STAT3 phosphorylation and HSC activation, and DHQ was able to successfully inhibit these effects. CONCLUSION: DHQ effectively alleviated spontaneous cholestatic liver fibrosis in Mdr2-/- mice by inhibiting H19 upregulation in cholangiocytes and preventing the inhibition of STAT3 phosphorylation in HSC, thereby suppressing cell activation. Please cite this article as: Li M, Zhou Y, Zhu H, Xu LM, Ping J. Danhongqing formula alleviates cholestatic liver fibrosis by downregulating long non-coding RNA H19 derived from cholangiocytes and inhibiting hepatic stellate cell activation. J Integr Med. 2024; 22(2): 188-198.

Lichen pectin-containing polysaccharide from Xanthoria elegans and its ability to effectively protect LX-2 cells from H2O2-induced oxidative damage.

Xanthoria elegans, a drought-tolerant lichen, is the original plant of the traditional Chinese medicine "Shihua" and effectively treats a variety of liver diseases. However, thus far, the hepatoprotective effects of polysaccharides, the most important chemical constituents of X. elegans, have not been determined. The aim of this study was to screen the polysaccharide fraction for hepatoprotective activity by using free radical scavenging assays and a H2O2-induced Lieming Xu-2 cell (LX-2) oxidative damage model and to elucidate the chemical composition of the bioactive polysaccharide fraction. In the present study, three polysaccharide fractions (XEP-50, XEP-70 and XEP-90) were obtained from X. elegans by hot-water extraction, DEAE-cellulose anion exchange chromatography separation and ethanol gradient precipitation. Among the three polysaccharide fractions, XEP-70 exhibited the best antioxidant activity in free radical scavenging capacity and reducing power assays. Structural studies showed that XEP-70 was a pectin-containing heteropolysaccharide fraction that was composed mainly of (1 â†’ 4)-linked and (1 â†’ 4,6)-linked α-D-Glcp, (1 â†’ 4)-linked α-D-GalpA, (1 â†’ 2)-linked, (1 â†’ 6)-linked and (1 â†’ 2,6)-linked α-D-Manp, and (1 â†’ 6)-linked and (1 â†’ 2,6)-linked ß-D-Galf. Furthermore, XEP-70 exhibited effectively protect LX-2 cells against H2O2-induced oxidative damage by enhancing cellular antioxidant capacity by activating the Nrf2/Keap1/ARE signaling pathway. Thus, XEP-70 has good potential to protect hepatic stellate cells against oxidative damage.

Wuling capsule modulates macrophage polarization by inhibiting the TLR4-NF-κB signaling pathway to relieve liver fibrosis.

BACKGROUND AND PURPOSE: Wuling capsule (WL) has good efficacy in the clinical treatment of chronic hepatitis B and liver injury. Liver fibrosis is a common pathological feature of chronic liver disease and may progress to irreversible cirrhosis and liver cancer. Accumulating evidence reveals that modulating macrophage polarization contribute to the therapy of liver fibrosis. However, the effects of WL on modulating macrophage polarization to relive liver fibrosis remain unclear. This study investigated the anti-liver fibrosis effects of WL in carbon tetrachloride (CCl4)-induced liver fibrosis in rats, and the modulation effects and underlying molecular mechanism on macrophage polarization. METHODS: A rat liver fibrosis model was constructed by intraperitoneal injection of 40 % CCl4 olive oil mixture. At 2, 4, 6, and 8 weeks, the histopathological status of the liver was assessed by hematoxylin-eosin (HE) and Masson staining; the liver biochemical indexes were measured in rat liver tissue. The expression levels of inflammatory cytokines in liver tissue were detected by ELISA. The mRNA levels and proteins expression of macrophage markers of different phenotypes, TLR4-NF-κB signaling pathway indicators were detected independently by ELISA, immunofluorescence, RT-PCR and western blotting. RESULTS: In vivo, WL treatment attenuated abnormal changes in weight, organ indices and biochemical indices, alleviated pathological changes, and reduced collagen fiber deposition as well as the expression of α-SMA in liver tissues. Further studies revealed that WL decreased the expression of the macrophage M1 polarization markers inducible nitric oxide synthase (iNOS), TNF-α, IL-6, and CD86, promoted the expression of the M2 macrophage polarization markers IL-10, CD206, and arginase-1 (Arg-1), and inhibited the activation of the TLR4-NF-κB signaling pathway via several key signaling proteins. In vitro, WL significantly suppressed macrophage M1 polarization, and promoted M2 polarization while boosted M1 polarization transform to M2 polarization in LPS-activated RAW264.7 cells. CONCLUSIONS: This study demonstrated that WL modulated macrophage polarization against liver fibrosis mainly by inhibiting the activation of the TLR4-NF-κB signaling pathway.

Dual-Targeting Macrophages and Hepatic Stellate Cells by Modified Albumin Nanoparticles for Liver Cirrhosis Treatment.

Hepatic cirrhosis has become a global public health concern with high mortality and currently lacks effective clinical treatment methods. Activation of hepatic stellate cells (HSCs) and the large number of macrophages infiltrating into the liver play a critical role in the development of liver cirrhosis. This study developed a novel modified nanoparticle system (SRF-CS-PSA NPs) in which Sorafenib (SRF) was encapsulated by palmitic acid-modified albumin (PSA) and further modified with chondroitin sulfate (CS). These modifications enabled the SRF-CS-PSA NPs to effectively target hepatic stellate cells (HSCs) and macrophages. SRF-CS-PSA NPs showed uniform particle size distribution of approximately 120 nm and high loading efficiency of up to 99.5% and can be taken up by HSCs and macrophages via CD44 and SR-A receptors, respectively. In a mouse model of liver cirrhosis, SRF-CS-PSA NPs demonstrated superior targeting and inhibition of HSCs and macrophages, effectively reversing the process of liver cirrhosis. Overall, our study demonstrates the potential of SRF-CS-PSA NPs as a targeted therapy for liver cirrhosis, with promising clinical applications.

Anti-tumor therapy of glycyrrhetinic acid targeted liposome co-delivery of doxorubicin and berberine for hepatocellular carcinoma.

During the development of hepatocellular carcinoma (HCC), hepatic stellate cells undergo activation and transform into cancer-associated fibroblasts (CAFs) due to the influence of tumor cells. The interaction between CAFs and tumor cells can compromise the effectiveness of chemotherapy drugs and promote tumor proliferation, invasion, and metastasis. This study explores the potential of glycyrrhetinic acid (GA)-modified liposomes (lip-GA) as a strategy for co-delivery of berberine (Ber) and doxorubicin (Dox) to treat HCC. The characterizations of liposomes, including particle size, zeta potential, polydispersity index, stability and in vitro drug release, were investigated. The study evaluated the anti-proliferation and anti-migration effects of Dox&Ber@lip-GA on the Huh-7 + LX-2 cell model were through MTT and wound-healing assays. Additionally, the in vivo drug distribution and anti-tumor efficacy were investigated using the H22 + NIH-3T3-bearing mouse model. The results indicated that Dox&Ber@lip-GA exhibited a nanoscale particle size, accumulated specifically in the tumor region, and was efficiently taken up by tumor cells. Compared to other groups, Dox&Ber@lip-GA demonstrated higher cytotoxicity and lower migration rates. Additionally, it significantly reduced the deposition of extracellular matrix (ECM) and inhibited tumor angiogenesis, thereby suppressing tumor growth. In conclusion, Dox&Ber@lip-GA exhibited superior anti-tumor effects both in vitro and in vivo, highlighting its potential as an effective therapeutic strategy for combating HCC.

Predictive toxicological effects of Artemisia absinthium essential oil on hepatic stellate cells.

Medicinal plants are important worldwide, considering their properties for treating diseases; however, few studies have evaluated their toxicological potential. Among them, Artemisia absinthium is frequently used to treat liver diseases, because its essential oil has several popular therapeutic properties. Based on this information, in the present study, we investigated molecular connectors of physiological effects of the Artemisia absinthium essential oil on human hepatic stellate cell line, LX-2, to explore the potential toxicity of the plant on liver cells. LX-2 is a cellular model to investigate mechanisms of liver fibrosis; then, to analyze the essential oil effects LX-2 was cultured under different conditions, treated or not with the essential oil at 0.4 µg/µL for 24 h. Next, fluorescence microscopy analyses, gene expression measurements, and biochemical approaches revealed that the essential oil reduced pro-fibrogenic markers; however, disrupt lipid metabolism, and cause cellular stress, by the activation of cellular detoxification and pro-inflammatory processes. In conclusion, the hepatic stellate cells incubated with the essential oil present an antifibrotic potential, supporting its popular use; however, the combined results suggest that the essential oil of Artemisia absinthium should be used with caution.

Jiawei Taohe Chengqi decoction inhibition of the notch signal pathway affects macrophage reprogramming to inhibit HSCs activation for the treatment of hepatic fibrosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Jiawei Taohe Chengqi Tang (JTCD) is a modified formulation of Traditional Chinese Medicine (TCM) known as Taohe Chengqi Decoction, which has been described in the ancient TCM literature "Treatise on Febrile Diseases". As a formula that can activate blood circulation and eliminate blood stasis and regulate Yin and Yang in traditional Chinese medicine applications, JTCD has been reported to be effective in the treatment of chronic liver disease and hepatic fibrosis (HF). AIM OF STUDY: The current study aimed to evaluate the effectiveness of JTCD in modulating hepatic macrophages by regulating the Notch signal pathway, and to further investigate the mechanisms underlying macrophage reprogramming that leads to HF. MATERIALS AND METHODS: Molecular assays were performed using in vitro cultures of human mononuclear THP-1 cells and human-derived hepatic stellate cells LX-2. CCl4-induced mice were utilized as an in vivo model to simulate HF. RESULTS: Our results demonstrated that JTCD exhibited dual effects by inhibiting hepatic stellate cell (HSCs) activation and modulating the polarisation of macrophages towards the M2 phenotype while decreasing the M1 phenotype. Network pharmacological analyses and molecular docking studies revealed that the Notch signal pathway was significantly enriched and played a crucial role in the therapeutic response of JTCD against HF. Moreover, through the establishment of a co-culture model, we validated that JTCD inhibited the Notch signal pathway in macrophages, leading to alterations in macrophage reprogramming, subsequent inhibition of HSC activation, and ultimately exerting anti-HF effects. CONCLUSION: In conclusion, our findings provide solid evidence for JTCD in treating HF, as it suppresses the Notch signal pathway in macrophages, regulates macrophage reprogramming, and inhibits HSC activation.

Calcitriol Inhibits NaAsO2 Triggered Hepatic Stellate Cells Activation and Extracellular Matrix Oversecretion by Activating Nrf2 Signaling Pathway Through Vitamin D Receptor.

Previous studies, including our own, have demonstrated that arsenic exposure can induce liver fibrosis, while the underlying mechanism remains unclear and there is currently no effective pharmacological intervention available. Recent research has demonstrated that vitamin D supplementation can ameliorate liver fibrosis caused by various etiologies, potentially through modulation of the Nrf2 signaling pathways. However, it remains unclear whether vitamin D intervention can mitigate arsenic-caused liver fibrosis. As is known hepatic stellate cells (HSCs) activation and extracellular matrix (ECM) deposition are pivotal in the pathogenesis of liver fibrosis. In this study, we investigated the intervention effect of calcitriol (a form of active vitamin D) on arsenite-triggered Lx-2 cells (a human hepatic stellate cell line) activation and ECM oversecretion. Additionally, we also elucidated the role and mechanism of Nrf2 antioxidant signaling pathway. Our results demonstrated that calcitriol intervention significantly inhibits Lx-2 cell activation and ECM oversecretion induced by arsenite exposure. Additionally, calcitriol activates Nrf2 and its downstream antioxidant enzyme expression in Lx-2 cells, thereby reducing ROS overproduction caused by arsenite exposure. Further investigation reveals that calcitriol activates the Nrf2 signaling pathway and inhibits arsenite-triggered Lx-2 cell activation and ECM oversecretion by targeting vitamin D receptor (VDR). In conclusion, this study has demonstrated that vitamin D intervention can effectively inhibit HSC activation and ECM oversecretion triggered by arsenite exposure through its antioxidant activity. This provides a novel strategy for targeted nutritional intervention in the treatment of arsenic-induced liver fibrosis.

Protective effect of water extracts of Veronicastrum latifolium (Hemsl.) Yamazaki on carbon tetrachloride-induced liver fibrosis in mice and its effect on intestinal flora.

Liver fibrosis refers to a reversible event of repair and reconstruction following injury due to various etiologies, and its continuous development will lead to cirrhosis and liver cancer. Abnormal alterations in intestinal microbiota can hasten the development of hepatic fibrosis and damage. Veronicastrum latifolium (Hemsl.) Yamazaki (VLY) is a classic drug applied extensively for managing acute and chronic hepatitis, liver cirrhosis and ascites in ethnic minority areas of Guizhou Province, China, which possesses broad-spectrum pharmacological activities. In view of the crucial role of intestinal microbiota in the development of liver fibrosis, the present study attempted to investigate the effects of VLY aqueous extract on ameliorating CCl4-elicited liver fibrosis in mice and on intestinal microbiota and to explore its possible mechanism. Phytochemical analysis showed that VLY water extract contained a variety of components, particularly rich in organic acids and their derivatives, flavonoids, phenolic acids, nucleotides and their derivatives, carbohydrates and other compounds. VLY water extract remarkably alleviated CCl4-induced liver damage and fibrosis in mice, improved liver histology, and improved liver function abnormalities. VLY water extract also inhibited the activation of hepatic stellate cells and invasion of intrahepatic inflammatory cells. Additionally, sequencing the 16 s rDNA gene revealed that VLY water extract changed the intestinal microbiota composition in liver fibrotic mice. It elevated the Firmicutes/Bacteroidota ratio and enriched the relative Lactobacillus richness, which is capable of mitigating fibrosis and inflammation in impaired liver. In summary, through modulation of inflammation and intestinal microbiota, VLY water extract can reduce the CCl4-elicited liver fibrosis.

New Perspectives on Chinese Medicine in Treating Hepatic Fibrosis: Lipid Droplets in Hepatic Stellate Cells.

Hepatic fibrosis (HF) is a wound healing response featuring excessive deposition of the extracellular matrix (ECM) and activation of hepatic stellate cells (HSCs) that occurs during chronic liver injury. As an initial stage of various liver diseases, HF is a reversible pathological process that, if left unchecked, can escalate into cirrhosis, liver failure, and liver cancer. HF is a life-threatening disease presenting morbidity and mortality challenges to healthcare systems worldwide. There is no specific and effective anti-HF therapy, and the toxic side effects of the available drugs also impose a heavy financial burden on patients. Therefore, it is significant to study the pathogenesis of HF and explore effective prevention and treatment measures. Formerly called adipocytes, or fat storage cells, HSCs regulate liver growth, immunity, and inflammation, as well as energy and nutrient homeostasis. HSCs in a quiescent state do not proliferate and store abundant lipid droplets (LDs). Catabolism of LDs is characteristic of the activation of HSCs and morphological transdifferentiation of cells into contractile and proliferative myofibroblasts, resulting in the deposition of ECM and the development of HF. Recent studies have revealed that various Chinese medicines (e.g., Artemisia annua, turmeric, Scutellaria baicalensis Georgi, etc.) are able to effectively reduce the degradation of LDs in HSCs. Therefore, this study takes the modification of LDs in HSCs as an entry point to elaborate on the process of Chinese medicine intervening in the loss of LDs in HSCs and the mechanism of action for the treatment of HF.

Salvianolic acid B inhibits hepatic stellate cell activation and liver fibrosis by targeting PDGFRß.

Liver fibrosis is a reversible pathological process and a wound healing response to liver injury. As an early stage of various liver diseases, liver fibrosis can develop into cirrhosis, liver failure, and even liver cancer if not controlled in time. Salvia miltiorrhiza is a medicinal plant with hepatoprotective effects. Salvianolic acid B (Sal B) is the representative component of S. miltiorrhiza. Many studies have reported the anti-liver fibrosis effects and mechanisms of Sal B. However, the direct anti-fibrotic targets of Sal B have not yet been reported. Platelet-derived growth factor receptor ß (PDGFRß) is one of the most classical targets in liver fibrosis, which is closely related to hepatic stellate cells (HSCs) activated. Previously, we established and applied a PDGFRß affinity chromatography model, and found that Sal B binds well to PDGFRß. Therefore, this study aimed to investigate the direct targets of Sal B against liver fibrosis. We confirmed the binding ability of Sal B to PDGFRß by molecular docking and a surface plasmon resonance biosensor. Our findings indicated that Sal B targeted PDGFRß to inhibit the activation, migration and proliferation of HSCs and suppressed the PDGF-BB-induced PDGFRß signaling pathway. Annexin V-FITC/PI assay showed that Sal B reversed the PDGF-BB-induced decrease in HSC apoptosis rate. In the mouse liver fibrosis model, Sal B inhibited the PDGFRß signaling pathway, HSC activation and reduced inflammatory response, ultimately improved CCl4-induced liver fibrosis. In summary, the direct anti-fibrotic targets of Sal B may be PDGFRß, and this study clarified the anti-liver fibrosis effects and mechanism of Sal B.

[Effect of Isodon ternifolius-medicated serum on hepatic stellate cells based on TLR4/NF-κB/NLRP3 signaling pathway].

The present study aimed to investigate the inhibitory effect and mechanism of Isodon terricolous-medicated serum on lipopolysaccharide(LPS)-induced hepatic stellate cell(HSC) activation. LPS-induced HSCs were divided into a blank control group, an LPS model group, a colchicine-medicated serum group, an LPS + blank serum group, an I. terricolous-medicated serum group, a Toll-like receptor 4(TLR4) blocker group, and a TLR4 blocker + I. terricolous-medicated serum group. HSC proliferation was detected by methyl thiazolyl tetrazolium(MTT) assay. Enzyme-linked immunosorbent assay(ELISA) was used to measure type Ⅰ collagen(COL Ⅰ), COL Ⅲ, transforming growth factor-ß1(TGF-ß1), intercellular adhesion molecule-1(ICAM-1), α-smooth muscle actin(α-SMA), vascular cell adhesion molecule-1(VCAM-1), cysteinyl aspartate-specific proteinase-1(caspase-1), and monocyte chemotactic protein-1(MCP-1). Real-time PCR(RT-PCR) was used to detect mRNA expression of TLR4, IκBα, and NOD-like receptor thermal protein domain associated protein 3(NLRP3), nuclear factor-κB(NF-κB) p65, gasdermin D(GSDMD), and apoptosis-associated speck-like protein containing a CARD(ASC) in HSCs. Western blot(WB) was used to detect the protein levels of TLR4, p-IκBα, NF-κB p65, NLRP3, ASC, and GSDMD in HSCs. The results showed that I. terricolous-medicated serum could inhibit the proliferation activity of HSCs and inhibit the secretion of COL Ⅰ, COL Ⅲ, α-SMA, TGF-ß1, caspase-1, MCP-1, VCAM-1, and ICAM-1 in HSCs. Compared with the LPS model group, the I. terricolous-medicated serum group, the colchicine-medicated serum group, and the TLR4 blocker group showed down-regulated expression of p-IκBα, NLRP3, NF-κB p65, GSDMD, and ASC, and up-regulated expression of IκBα. Compared with the TLR4 blocker group, the TLR4 blocker + I. terricolous-medicated serum group showed decreased expression of TLR4, p-IκBα, NLRP3, NF-κB p65, GSDMD, and ASC, and increased expression of IκBα. In conclusion, I. terricolous-medicated serum down-regulates HSC activation by inhibiting the TLR4/NF-κB/NLRP3 signaling pathway.

Ginsenoside Rh2 promotes hepatic stellate cell ferroptosis and inactivation via regulation of IRF1-inhibited SLC7A11.

BACKGROUND: 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. PURPOSE: 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. METHODS: 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. RESULTS: 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. CONCLUSION: 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.

Calcitriol attenuates liver fibrosis through hepatitis C virus nonstructural protein 3-transactivated protein 1-mediated TGF ß1/Smad3 and NF-κB signaling pathways.

BACKGROUND: Hepatic fibrosis is a serious condition, and the development of hepatic fibrosis can lead to a series of complications. However, the pathogenesis of hepatic fibrosis remains unclear, and effective therapy options are still lacking. Our group identified hepatitis C virus nonstructural protein 3-transactivated protein 1 (NS3TP1) by suppressive subtractive hybridization and bioinformatics analysis, but its role in diseases including hepatic fibrosis remains undefined. Therefore, additional studies on the function of NS3TP1 in hepatic fibrosis are urgently needed to provide new targets for treatment. AIM: To elucidate the mechanism of NS3TP1 in hepatic fibrosis and the regulatory effects of calcitriol on NS3TP1. METHODS: Twenty-four male C57BL/6 mice were randomized and separated into three groups, comprising the normal, fibrosis, and calcitriol treatment groups, and liver fibrosis was modeled by carbon tetrachloride (CCl4). To evaluate the level of hepatic fibrosis in every group, serological and pathological examinations of the liver were conducted. TGF-ß1 was administered to boost the in vitro cultivation of LX-2 cells. NS3TP1, α-smooth muscle actin (α-SMA), collagen I, and collagen III in every group were examined using a Western blot and real-time quantitative polymerase chain reaction. The activity of the transforming growth factor beta 1 (TGFß1)/Smad3 and NF-κB signaling pathways in each group of cells transfected with pcDNA-NS3TP1 or siRNA-NS3TP1 was detected. The statistical analysis of the data was performed using the Student's t test. RESULTS: NS3TP1 promoted the activation, proliferation, and differentiation of hepatic stellate cells (HSCs) and enhanced hepatic fibrosis via the TGFß1/Smad3 and NF-κB signaling pathways, as evidenced by the presence of α-SMA, collagen I, collagen III, p-smad3, and p-p65 in LX-2 cells, which were upregulated after NS3TP1 overexpression and downregulated after NS3TP1 interference. The proliferation of HSCs was lowered after NS3TP1 interference and elevated after NS3TP1 overexpression, as shown by the luciferase assay. NS3TP1 inhibited the apoptosis of HSCs. Moreover, both Smad3 and p65 could bind to NS3TP1, and p65 increased the promoter activity of NS3TP1, while NS3TP1 increased the promoter activity of TGFß1 receptor I, as indicated by coimmunoprecipitation and luciferase assay results. Both in vivo and in vitro, treatment with calcitriol dramatically reduced the expression of NS3TP1. Calcitriol therapy-controlled HSCs activation, proliferation, and differentiation and substantially suppressed CCl4-induced hepatic fibrosis in mice. Furthermore, calcitriol modulated the activities of the above signaling pathways via downregulation of NS3TP1. CONCLUSION: Our results suggest that calcitriol may be employed as an adjuvant therapy for hepatic fibrosis and that NS3TP1 is a unique, prospective therapeutic target in hepatic fibrosis.

Bromelain mitigates liver fibrosis via targeting hepatic stellate cells in vitro and in vivo.

Various therapeutic approaches are conducted for regression of liver fibrosis and prevent possible further carcinogenic transformation. This study was aimed to assess the prospective therapeutic potential of bromelain against thioacetamide (TAA)-induced liver fibrosis using in-vitro and in vivo approaches. In vitro study, HSC-T6 cell line was used to evaluate the effect of bromelain on HSC-T6 cell viability and apoptosis. In vivo, Rats were treated by TAA for 6 weeks for induction of hepatic fibrosis followed by post treatment by different doses of bromelain and silymarin for further 4 weeks to assess the regression of hepatic fibrosis. The in-vitro findings indicated that bromelain hindered the proliferation of HSCs in concentration dependent manner compared with the untreated cells. The in vivo study revealed that treatment of TAA fibrotic rats with different doses of bromelain and silymarin induced a significant restoration in liver function biomarkers, attenuation of oxidative stress, upregulation of total antioxidant capacity and thereby decline of fibrotic biomarkers and improving histopathological and immunohistochemical changes. In conclusion, This study indicates that bromelain can regress TAA induced hepatic fibrosis in rats via inhibiting HSCs activation, α-SMA expression and the ECM deposition in hepatic tissue in addition to its antioxidants pathway, these findings prove the promising therapeutic potential of bromelain as a novel therapeutic approach for chronic hepatic fibrotic diseases.

Sclareol attenuates liver fibrosis through SENP1-mediated VEGFR2 SUMOylation and inhibition of downstream STAT3 signaling.

Liver fibrosis is a key global health care burden. Sclareol, isolated from Salvia sclarea, possesses various biological activities. Its effect on liver fibrosis remains unknown. This study was proposed to evaluate the antifibrotic activity of sclareol (SCL) and explore its underlying mechanisms. Stimulated hepatic stellate cells served as an in vitro liver fibrosis model. The expression of fibrotic markers was assessed by western blot and real-time PCR. Two classical animal models, bile duct-ligated rats and carbon tetrachloride-treated mice, were utilized for the in vivo experiments. The liver function and fibrosis degree were determined by serum biochemical and histopathological analyses. VEGFR2 SUMOylation was analyzed using coimmunoprecipitation assay. Our results indicated that SCL treatment restricted the profibrotic propensity of activated HSCs. In fibrotic rodents, SCL administration alleviated hepatic injury and reduced collagen accumulation. Mechanistic studies indicated that SCL downregulated the protein level of SENP1 and enhanced VEGFR2 SUMOylation in LX-2 cells, which affected its intracellular trafficking. Blockade of the interaction between VEGFR2 and STAT3 was observed, resulting in the suppression of downstream STAT3 phosphorylation. Our findings demonstrated that SCL has therapeutic efficacy against liver fibrosis through mediating VEGFR2 SUMOylation, suggesting that SCL may be a potential candidate compound for its treatment.

Gandouling inhibits hepatic fibrosis in Wilson's disease through Wnt-1/ß-catenin signaling pathway.

ETHNOPHARMACOLOGIC SIGNIFICANCE: Wilson's disease (WD) hepatic fibrosis is the result of chronic liver injury induced by Cu2+ deposition in the liver. Gandouling (GDL) is a hospital preparation of the First Affiliated Hospital of Anhui University of Chinese Medicine. Previous studies have found that GDL can play an anti-inflammatory, anti-oxidation, and promote Cu2+ excretion, which has a clear anti-WD effect. AIM OF THE STUDY: We found that Wnt-1 was significantly up-regulated in the liver tissue of toxic-milk (TX) mouse in the WD gene mutant model, and the monomer components of GDL could combine well with Wnt-1. Therefore, in this work, we used RT-qPCR, Western blot, immunofluorescence, network pharmacology, molecular docking, and related methods to study the effects of GDL on hepatic stellate cell (HSC) activation and Wnt-1/ß-catenin pathway in TX mice to clarify the effect of GDL on WD hepatic fibrosis. RESULTS: GDL could alleviate hepatic fibrosis, improve liver function, and inhibit the activation of HSC in TX mice. Network pharmacology predicted that the Wnt-1/ß-catenin was the target of GDL, and molecular dynamics further revealed that GDL has a good binding ability with Wnt-1 and inhibits the Wnt/ß-catenin signaling pathway through Wnt-1. Furthermore, we found that GDL blocked the Wnt-1/ß-catenin signaling pathway in the liver of TX mice in vivo. In vitro, serum containing GDL blocked the Cu2+ ion-induced Wnt-1/ß-catenin signaling pathway in LX-2 cells. Therefore, GDL blocked the Wnt-1/ß-catenin signaling pathway, inhibited HSC activation, and improved WD hepatic fibrosis by binding to Wnt-1. CONCLUSION: GDL improves hepatic fibrosis in WD model mice by blocking the Wnt-1/ß-catenin signaling pathway, and Wnt-1 may be a new target for the diagnosis and treatment of WD. This reveals a new mechanism of GDL against WD, and promotes the clinical promotion of GDL.

Understanding the Potential Role of Nanotechnology in Liver Fibrosis: A Paradigm in Therapeutics.

The liver is a vital organ that plays a crucial role in the physiological operation of the human body. The liver controls the body's detoxification processes as well as the storage and breakdown of red blood cells, plasma protein and hormone production, and red blood cell destruction; therefore, it is vulnerable to their harmful effects, making it more prone to illness. The most frequent complications of chronic liver conditions include cirrhosis, fatty liver, liver fibrosis, hepatitis, and illnesses brought on by alcohol and drugs. Hepatic fibrosis involves the activation of hepatic stellate cells to cause persistent liver damage through the accumulation of cytosolic matrix proteins. The purpose of this review is to educate a concise discussion of the epidemiology of chronic liver disease, the pathogenesis and pathophysiology of liver fibrosis, the symptoms of liver fibrosis progression and regression, the clinical evaluation of liver fibrosis and the research into nanotechnology-based synthetic and herbal treatments for the liver fibrosis is summarized in this article. The herbal remedies summarized in this review article include epigallocathechin-3-gallate, silymarin, oxymatrine, curcumin, tetrandrine, glycyrrhetinic acid, salvianolic acid, plumbagin, Scutellaria baicalnsis Georgi, astragalosides, hawthorn extract, and andrographolides.