Bicyclol attenuates pulmonary fibrosis with silicosis via both canonical and non-canonical TGF-ß1 signaling pathways.

BACKGROUND: Silicosis is an irreversible fibrotic disease of the lung caused by chronic exposure to silica dust, which manifests as infiltration of inflammatory cells, excessive secretion of pro-inflammatory cytokines, and pulmonary diffuse fibrosis. As the disease progresses, lung function further deteriorates, leading to poorer quality of life of patients. Currently, few effective drugs are available for the treatment of silicosis. Bicyclol (BIC) is a compound widely employed to treat chronic viral hepatitis and drug-induced liver injury. While recent studies have demonstrated anti-fibrosis effects of BIC on multiple organs, including liver, lung, and kidney, its therapeutic benefit against silicosis remains unclear. In this study, we established a rat model of silicosis, with the aim of evaluating the potential therapeutic effects of BIC. METHODS: We constructed a silicotic rat model and administered BIC after injury. The FlexiVent instrument with a forced oscillation system was used to detect the pulmonary function of rats. HE and Masson staining were used to assess the effect of BIC on silica-induced rats. Macrophages-inflammatory model of RAW264.7 cells, fibroblast-myofibroblast transition (FMT) model of NIH-3T3 cells, and epithelial-mesenchymal transition (EMT) model of TC-1 cells were established in vitro. And the levels of inflammatory mediators and fibrosis-related proteins were evaluated in vivo and in vitro after BIC treatment by Western Blot analysis, RT-PCR, ELISA, and flow cytometry experiments. RESULTS: BIC significantly improved static compliance of lung and expiratory and inspiratory capacity of silica-induced rats. Moreover, BIC reduced number of inflammatory cells and cytokines as well as collagen deposition in lungs, leading to delayed fibrosis progression in the silicosis rat model. Further exploration of the underlying molecular mechanisms revealed that BIC suppressed the activation, polarization, and apoptosis of RAW264.7 macrophages induced by SiO2. Additionally, BIC inhibited SiO2-mediated secretion of the inflammatory cytokines IL-1ß, IL-6, TNF-α, and TGF-ß1 in macrophages. BIC inhibited FMT of NIH-3T3 as well as EMT of TC-1 in the in vitro silicosis model, resulting in reduced proliferation and migration capability of NIH-3T3 cells. Further investigation of the cytokines secreted by macrophages revealed suppression of both FMT and EMT by BIC through targeting of TGF-ß1. Notably, BIC blocked the activation of JAK2/STAT3 in NIH-3T3 cells required for FMT while preventing both phosphorylation and nuclear translocation of SMAD2/3 in TC-1 cells necessary for the EMT process. CONCLUSION: The collective data suggest that BIC prevents both FMT and EMT processes, in turn, reducing aberrant collagen deposition. Our findings demonstrate for the first time that BIC ameliorates inflammatory cytokine secretion, in particular, TGF-ß1, and consequently inhibits FMT and EMT via TGF-ß1 canonical and non-canonical pathways, ultimately resulting in reduction of aberrant collagen deposition and slower progression of silicosis, supporting its potential as a novel therapeutic agent.

Upregulation of Hepatic Glutathione S-Transferase Alpha 1 Ameliorates Metabolic Dysfunction-Associated Steatosis by Degrading Fatty Acid Binding Protein 1.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is the most common metabolic disease of the liver, characterized by hepatic steatosis in more than 5% of hepatocytes. However, despite the recent approval of the first drug, resmetirom, for the management of metabolic dysfunction-associated steatohepatitis, decades of target exploration and hundreds of clinical trials have failed, highlighting the urgent need to find new druggable targets for the discovery of innovative drug candidates against MASLD. Here, we found that glutathione S-transferase alpha 1 (GSTA1) expression was negatively associated with lipid droplet accumulation in vitro and in vivo. Overexpression of GSTA1 significantly attenuated oleic acid-induced steatosis in hepatocytes or high-fat diet-induced steatosis in the mouse liver. The hepatoprotective and anti-inflammatory drug bicyclol also attenuated steatosis by upregulating GSTA1 expression. A detailed mechanism showed that GSTA1 directly interacts with fatty acid binding protein 1 (FABP1) and facilitates the degradation of FABP1, thereby inhibiting intracellular triglyceride synthesis by impeding the uptake and transportation of free fatty acids. Conclusion: GSTA1 may be a good target for the discovery of innovative drug candidates as GSTA1 stabilizers or enhancers against MASLD.

Bicyclol mitigates lipopolysaccharide-induced acute lung injury through myeloid differentiation factor 88 inhibition.

Acute lung injury (ALI) remains a significant clinical challenge due to the absence of effective treatment alternatives. This study presents a new method that employs a screening platform focusing on MyD88 affinity, anti-inflammatory properties, and toxicity. This platform was used to evaluate a 300-compound library known for its anti-inflammatory potential. Among the screened compounds, Bicyclol emerged as a standout, exhibiting MyD88 binding and a significant reduction in LPS-stimulated pro-inflammatory factors production in mouse primary peritoneal macrophages. By targeting MyD88, Bicyclol disrupts the MyD88/TLR4 complex and MyD88 polymer formation, thereby mitigating the MAPKs and NF-κB signaling pathways. In vivo experiments further confirmed Bicyclol's efficacy, demonstrating alleviated ALI symptoms, decreased inflammatory cytokines level, and reduced inflammatory cells presence in lung tissues. These findings were associated with a decrease in mortality in LPS-challenged mice. Overall, Bicyclol represents a promising treatment option for ALI by specifically targeting MyD88 and limiting inflammatory responses.

Comprehensive review of dibenzocyclooctadiene lignans from the Schisandra genus: anticancer potential, mechanistic insights and future prospects in oncology.

Cancer remains one of the leading causes of mortality worldwide. The search for novel and effective anticancer agents has been a significant area of research. Dibenzocyclooctadiene lignans (DBCLS), derived from the Schisandra genus plants like: S. chinensis, S. sphenanthera, S. henryi, S. rubriflora, S. grandiflora, S. propinqua, and S. glabra, have been traditionally used in various medicinal systems and are known for their myriad health benefits, including anticancer properties. This comprehensive review aimed to collate and critically analyse the recent literature on the anticancer properties of DBCLS, focusing on their mechanistic approaches against different cancer types. An exhaustive literature search was performed using databases like PubMed/MedLine, Scopus, Web of Science, Embase, TRIP database and Google Scholar from 1980 to 2023. Peer-reviewed articles that elucidated the mechanistic approach of these lignans on cancer cell lines, in vivo models and preliminary clinical studies were included. Studies were assessed for their experimental designs, cancer types studied, and the mechanistic insights provided. The studies demonstrate that the anticancer effects of DBCLS compounds are primarily driven by their ability to trigger apoptosis, arrest the cell cycle, induce oxidative stress, modulate autophagy, and disrupt essential signaling pathways, notably MAPK, PI3K/Akt, and NF-κB. Additionally, these lignans have been shown to amplify the impact of traditional chemotherapy treatments, suggesting their potential role as supportive adjuncts in cancer therapy. Notably, several studies also emphasise their capacity to target cancer stem cells and mitigate multi-drug resistance specifically. DBCLS from the Schisandra genus have showcased significant potential as anticancer agents. Their multi-targeted mechanistic approach makes them promising candidates for further research, potentially leading to developing of new therapeutic strategies in cancer management.

Practical gram-scale synthesis of bicyclol metabolites M2 and M3.

Bicyclol, an innovative hepatoprotective drug, was approved by the Chinese National Medical Products Administration (NMPA) in 2001 to treat Hepatitis B and drug-induced liver injury. Two active metabolites of bicyclol have been identified as M2 and M3. To evaluate the impact on drug safety and efficacy of possible drug-drug interactions (DDIs) associated with these metabolites, a sufficient quantity of these metabolites is required. Herein, we report a concise novel route for the synthesis of M2 and M3 using the Suzuki-Miyaura coupling as the key step. Furthermore, we complete the gram-scale syntheses of M2 and M3.

Investigation on the in vitro metabolism of bicyclol using liver microsomes, hepatocytes and human recombinant cytochrome P450 enzymes.

In vitro metabolism of bicyclol was studied using liver microsomes, hepatocytes and human recombinant cytochrome P450 enzymes. Liquid chromatography-benchtop orbitrap mass spectrometry technique was utilised to identify the metabolites.A total of 19 metabolites, including 5 new metabolites (M2, M3, M4, M5 and M16) were tentatively identified. Among these metabolites, M6&M8 (demethylenation), M9&M10 (demethylation) and M19 (glucuronidation) were the major metabolites.In glutathione (GSH)-supplemented liver microsomes, 5 new GSH conjugates were found and tentatively identified. The formation was assumed to be through demethylenation of methylenedioxyphenyl to form catechol derivatives, which further underwent oxidation to form ortho-quinone intermediates, reacting with GSH to form stable adducts.CYP3A4 and 2C19 were demonstrated to be the major enzymes responsible for the bioactivation of bicyclol.This study provided valuable information on the metabolic fate of bicyclol in liver microsomes and hepatocytes, and the bioactivation pathways were reported for the first time, which would be helpful for us to understand the potential drug-drug interactions and the possible side effect of this drug.

Erratum: Bicyclol attenuates high fat diet-induced non-alcoholic fatty liver disease/non-alcoholic steatohepatitis through modulating multiple pathways in mice.

[This corrects the article DOI: 10.3389/fphar.2023.1157200.].

Bicyclol attenuates high fat diet-induced non-alcoholic fatty liver disease/non-alcoholic steatohepatitis through modulating multiple pathways in mice.

Introduction: The pathological progression of non-alcoholic fatty liver disease (NAFLD) is driven by multiple factors, and non-alcoholic steatohepatitis (NASH) represents its progressive form. In our previous studies, we found that bicyclol had beneficial effects on NAFLD/ NASH. Here we aim to investigate the underlying molecular mechanisms of the bicyclol effect on NAFLD/NASH induced by high-fat diet (HFD) feeding. Methods: A mice model of NAFLD/NASH induced by HFD-feeding for 8 weeks was used. As a pretreatment, bicyclol (200 mg/kg) was given to mice by oral gavage twice daily. Hematoxylin and eosin (H&E) stains were processed to evaluate hepatic steatosis, and hepatic fibrous hyperplasia was assessed by Masson staining. Biochemistry analyses were used to measure serum aminotransferase, serum lipids, and lipids in liver tissues. Proteomics and bioinformatics analyses were performed to identify the signaling pathways and target proteins. Data are available via Proteome X change with identifier PXD040233. The real-time RT-PCR and Western blot analyses were performed to verify the proteomics data. Results: Bicyclol had a markedly protective effect against NAFLD/NASH by suppressing the increase of serum aminotransferase, hepatic lipid accumulation and alleviating histopathological changes in liver tissues. Proteomics analyses showed that bicyclol remarkably restored major pathways related to immunological responses and metabolic processes altered by HFD feeding. Consistent with our previous results, bicyclol significantly inhibited inflammation and oxidative stress pathway related indexes (SAA1, GSTM1 and GSTA1). Furthermore, the beneficial effects of bicyclol were closely associated with the signaling pathways of bile acid metabolism (NPC1, SLCOLA4 and UGT1A1), cytochrome P450-mediated metabolism (CYP2C54, CYP3A11 and CYP3A25), biological processes such as metal ion metabolism (Ceruloplasmin and Metallothionein-1), angiogenesis (ALDH1A1) and immunological responses (IFI204 and IFIT3). Discussion: These findings suggested that bicyclol is a potential preventive agent for NAFLD/NASH by targeting multiple mechanisms in future clinical investigations.

Bicyclol Alleviates Streptozotocin-induced Diabetic Cardiomyopathy By Inhibiting Chronic Inflammation And Oxidative Stress.

PURPOSE: Diabetic cardiomyopathy (DCM) is a common and severe complication of diabetes. Inflammation and oxidative stress play important roles in DCM development. Bicyclol is a hepatoprotective drug in China that exerts anti-inflammatory effects by inhibiting the MAPK and NF-κB pathways to prevent obesity-induced cardiomyopathy. Our purpose was to explore the effect and mechanism of bicyclol on DCM. METHODS: A type 1 diabetes mouse model was established using C57BL/6 mice by intraperitoneal injection of STZ. The therapeutic effect of bicyclol was evaluated in both heart tissues of diabetic mice and high concentration of glucose (HG)-stimulated H9c2 cells. RESULTS: We showed that bicyclol significantly attenuated diabetes-induced cardiac hypertrophy and fibrosis, which is accompanied by the preservation of cardiac function in mice. In addition, bicyclol exhibited anti-inflammatory and anti-oxidative effects both in vitro and in vivo. Furthermore, bicyclol inhibited the hyperglycemia-induced activation of MAPKs and NF-κB pathways, while upregulating the Nrf-2/HO-1 pathway to exhibit protective effects. CONCLUSION: Our data indicate that bicyclol could be a promising cardioprotective agent in the treatment of DCM.

Bicyclol Attenuates Obesity-Induced Cardiomyopathy via Inhibiting NF-κB and MAPK Signaling Pathways.

PURPOSE: Schisandra is a well-known traditional Chinese medicine in East Asia. As a traditional Chinese medicine derivative with Schisandra chinensis as raw material, bicyclol is well known for its significant anti-inflammatory effect. Chronic inflammation plays a significant part in obesity-induced cardiomyopathy. Our purpose was to explore the effect and mechanism of bicyclol on obesity-induced cardiomyopathy. METHODS: Mice fed with a high-fat diet (HFD) and cardiomyocytes stimulated by palmitic acid (PA) were used as models of obesity-related cardiomyopathy in vivo and in vitro, respectively. The therapeutic effect of bicyclol on pathological changes such as myocardial hypertrophy and fibrosis was evaluated by staining cardiac tissue sections. PCR was used to detect inflammatory factors in H9c2 cells and animal heart tissue after bicyclol treatment. Then, we used western blotting to detect the expression levels of the myocardial hypertrophy related protein, myocardial fibrosis related protein, NF-κB and MAPK pathways. RESULTS: Our results indicated that bicyclol treatment significantly alleviates HFD-induced myocardial inflammation, fibrosis, and hypertrophy by inhibiting the MAPK and NF-κB pathways. Similar to animal level results, bicyclol could significantly inhibit PA-induced inflammation and prevent NF-κB and MAPK pathways from being activated. CONCLUSION: Our results showed that bicyclol has potential as a drug to treat obesity-induced cardiomyopathy.

Effects of bicyclol on hepatic sinusoidal obstruction syndrome induced by Gynura segetum.

BACKGROUND: The intake of Gynura segetum, a traditional Chinese medicine, may be induce hepatic sinusoidal obstruction syndrome (HSOS). It has a high mortality rate based on the severity of the disease and the absence of therapeutic effectiveness. Therefore, the current study was designed to investigate the effects of bicyclol on HSOS induced by Gynura segetum and the potential molecular mechanisms. METHODS: Gynura segetum (30 g/kg) was administered for 4 weeks in the model group, while the bicyclol pretreatment group received bicyclol (200 mg/kg) administration. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), cholesterol (CHO), triglyceride (TG), and liver histological assays were detected to assess HSOS. The gene expressions of cytochrome P450 (CYP450) isozymes were quantified by real-time PCR. Moreover, hepatocellular apoptosis was detected using the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay, then apoptosis and autophagy-related markers were determined using Western blot. RESULTS: As a result, bicyclol pretreatment is notably protected against Gynura segetum-induced HSOS, as observed by reducing serum ALT levels, inhibiting the reduction in CHO and TG levels, and alleviating the histopathological changes. Bicyclol pretreatment inhibited the changes in mRNA levels of CYP450 isozymes (including the increase in CYP2a5 and decrease in CYP2b10, 2c29, 2c37, 3a11, and 7b1). In addition, the upregulation of Bcl-2 and the downregulation of LC3-II/LC3-I proteins expression in HSOS were inhibited with bicyclol pretreatment. CONCLUSION: Bicyclol exerted a protective effect against HSOS induced by Gynura segetum, which could be attributed to the regulated expressions of CYP450 isozymes and alleviated the downregulation of autophagy.

Identification of bicyclol metabolites in rat plasma, urine and feces by UPLC-Q-TOF-MS/MS and evaluation of the efficacy and safety of these metabolites based on network pharmacology and molecular docking combined with toxicity prediction.

Bicyclol (BIC) has been widely used to treat drug-induced liver injury (DILI), however, it still has the problems of low solubility and bioavailability. Besides, the metabolic characteristics of BIC remain unclear. In the current study, we identified the metabolite of BIC in rat plasma, urine and feces, and evaluated the efficacy and safety of these metabolites. Based on the fragmentation behavior, we totally identified 11 metabolites and 7 metabolites in plasma, 8 metabolites in urine and 8 metabolites in feces. Notably, M1-M3, M6, M7, M10 and M11 were identified for the first time. M7 was the most abundant metabolite in the rat plasma. The metabolic pathways mainly involved demethylation, dealkylation, hydrolysis, methylation, oxidation and glucuronidation. In addition, the efficacy and safety of BIC's metabolites were evaluated by network pharmacology and molecular docking combined with toxicity prediction. The analysis of network pharmacology indicated that BIC's metabolites against DILI through the MAPK signaling pathway and Hepatitis B pathway. The molecular docking results showed that the binding energy of 5 compounds that docked with "7nuw" and 10 compounds that docked with "4tjz" was lower than BIC. 11 compounds possessed higher solubility and lower toxicity than BIC in prediction. Thus, the identification and evaluation of BIC's metabolites contributed to a better understanding of pharmacological mechanism of BIC and the high-value metabolites of high efficacy, safety and solubility provided a basis for drug development.

Bicyclol alleviates high-fat diet-induced hepatic ER stress- and autophagy-associated non-alcoholic fatty liver disease/non-alcoholic steatohepatitis in mice.

BACKGROUND AND OBJECTIVE: Non-alcoholic fatty liver disease (NAFLD) is a manifestation of the metabolic syndrome in the liver, and non-alcoholic steatohepatitis (NASH) represents its advanced stage. Bicyclol has protective activity against NAFLD in mice; however, the effect of bicyclol on high-fat diet (HFD)-induced NASH and its underlying molecular mechanism remains unknown particularly anti-endoplasmic reticulum (ER) stress and autophagic machinery potentials. Therefore, the present study was performed to investigate the protective effect and underlying mechanisms of bicyclol action on NAFLD/NASH. METHODS: Mice were fed an HFD to induce NAFLD/NASH, and bicyclol was administered as a treatment. Biochemistry and histopathological assays were performed to evaluate the effects of bicyclol on NAFLD/NASH. Moreover, the levels of hepatic ER stress- and autophagy-related markers were determined by western blotting. RESULTS: The present results revealed that bicyclol exerted significant protective effects against HFD-induced NAFLD/NASH. This activity was evidenced by the decrease in elevated serum transaminase and hepatic triglyceride levels, and the attenuation of negative histopathological changes. Bicyclol considerably alleviated hepatic inflammation and apoptosis. The protein expression of ER stress-related markers, including C/EBP homologous protein (CHOP) and glucose-regulated protein 78 (GRP78), was downregulated by the bicyclol treatment in HFD-induced mice. However, the protein expression of autophagy-related markers (LC3 and Beclin 1) was upregulated by the treatment with bicyclol. CONCLUSION: Bicyclol protected HFD-induced NASH, and partly due to its ability of reducing ER stress and promoting autophagy.

Bicyclol ameliorates advanced liver diseases in murine models via inhibiting the IL-6/STAT3 signaling pathway.

Bicyclol, a synthetic hepatoprotective and anti-inflammatory agent approved in China, was widely used to treat various hepatitis accompanied by elevated serum aminotransferases. However, the pharmacological effects and mechanisms of bicyclol on advanced liver diseases, such as fibrosis/cirrhosis and hepatocellular carcinoma (HCC), remain to be explored. Here, we revealed that bicyclol prevents from formatting severe fibrosis, slows the progression of moderate liver fibrosis, accelerates the regression of moderate liver fibrosis, decreases the malignancy of HCC in rat models induced by diethylnitrosamine (DEN), and also blocks steatohepatitis to HCC in mice induced by western diet plus carbon tetrachloride and DEN. The detailed pharmacological mechanism showed that bicyclol alleviates chronic progressive liver diseases by inhibiting the levels of IL-6 and subsequent phosphorylated STAT3. Conclusion: Bicyclol plays significant protective roles in multiply stages of fibrosis/cirrhosis-HCC and nonalcoholic fatty liver disease-related HCC via inhibiting IL-6/STAT3 signaling pathway. Therefore, bicyclol might be a promising therapeutic strategy for treating advanced liver diseases.

Efficacy and safety of bicyclol for treating patients with idiosyncratic acute drug-induced liver injury: A multicenter, randomized, phase II trial.

BACKGROUND AND AIMS: Evidence for using bicyclol in drug-induced liver injury (DILI) is limited. This study aimed to explore the efficacy and safety of bicyclol in acute DILI. METHODS: This was a multicenter, randomized, double-blinded, double-dummy, active-controlled, superiority and phase II trial. Patients with idiosyncratic acute DILI were randomized 1: 1:1 to low-dose bicyclol (25 mg times a day [TID]), high-dose bicyclol (50 mg TID) and polyene phosphatidylcholine (control) groups. The primary endpoint was the decrease from baseline in serum alanine aminotransferase (ALT) levels at post-treatment for 4 weeks. RESULTS: Overall, 241 patients were included in the full analysis set, with 81, 82 and 78 patients in the low-dose bicyclol, high-dose bicyclol, and control groups respectively. ALT levels decreased across groups (-249.2 ± 151.1, -273.6 ± 203.1, and -180.8 ± 218.2 U/L in the low-dose bicyclol, high-dose bicyclol and control groups, respectively; both p < .001, the bicyclol-dependent groups vs. control group). The ALT normalization rates at weeks 1, 2, 4, 6 and 8 were higher in the bicyclol-dependent groups than in the control group (p = .002 at week 1 and all p < .001 at weeks 2, 4, 6 and 8 respectively). The median times to ALT normalization in the low-dose bicyclol, high-dose bicyclol and control groups were 29, 16 and 43 days respectively. Adverse events, serious adverse events and adverse drug reactions were similar across groups. CONCLUSIONS: Bicyclol (25 and 50 mg TID) appeared efficacious and safe for treating idiosyncratic acute DILI, while bicyclol 50 mg TID showed higher efficacy. TRIAL REGISTRATION NUMBER: www. CLINICALTRIALS: gov (registration no. NCT02944552).

Effect of berberine and bicyclol on Chinese patients with nonalcoholic fatty liver disease: a retrospective study.

OBJECTIVES: To compare the effect of berberine and bicyclol on patients with nonalcoholic fatty liver disease (NAFLD). METHODS: Chinese nonalcoholic and non-viral hepatitis patients with a hepatic lipid content > 13% and nonalcoholic fatty liver disease activity score (NAS) ≥ 2 were treated with 500 mg berberine thrice daily, together with dietary modification (low-fat diet) and Tai Chi exercise for 4 months (BT cohort; n = 112), or 25 mg bicyclol thrice daily plus dietary modification and Tai Chi exercise for 4 months (CT cohort, n = 145), or dietary modification and Tai Chi exercise for 4 months (DT cohort, n = 128). RESULTS: Patients in the BT and the CT cohorts had improved anthropometric measurements (weight, height, body mass index, and waist-to-hip ratio), biochemical parameters (blood sugar, lipid profile, and liver functions tests), liver/spleen computed tomography findings, and liver biopsy results after 4 months of intervention (p < 0.05 for all). Bicyclol decreased the NAS in the CT cohort to a significantly greater degree than berberine in the BT cohort (p < 0.0001, q = 3.879). Patients in the DT cohort had reduced body mass index and waist-to-hip ratio (p < 0.05 for both). During the 4-month intervention, patients in the BT cohort had abdominal distension, mild diarrhea, constipation, nausea, and dyspepsia; patients in the CT cohort had dizziness and abdominal distension. CONCLUSIONS: Berberine or bicyclol plus dietary modification and Tai Chi exercise could control NAFLD without serious adverse effects. Dietary modification and Tai Chi exercise alone for 4 months are insufficient for the management of NAFLD. It is possible to reduce body weight by administering berberine or bicyclol.

Combined Use of Bicyclol and Berberine Alleviates Mouse Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD), ranging from simple steatosis to nonalcoholic steatohepatitis (NASH), is a liver disease worldwide without approved therapeutic drugs. Anti-inflammatory and hepatoprotective drug bicyclol and multi-pharmacological active drug berberine, respectively, have shown beneficial effects on NAFLD in murine nutritional models and patients, though the therapeutic mechanisms remain to be illustrated. Here, we investigated the combined effects of bicyclol and berberine on mouse steatosis induced by Western diet (WD), and NASH induced by WD/CCl4. The combined use of these was rather safe and better reduced the levels of transaminase in serum and triglycerides and cholesterol in the liver than their respective monotherapy, accompanied with more significantly attenuating hepatic inflammation, steatosis, and ballooning in mice with steatosis and NASH. The combined therapy also significantly inhibited fibrogenesis, characterized by the decreased hepatic collagen deposition and fibrotic surface. As per mechanism, bicyclol enhanced lipolysis and ß-oxidation through restoring the p62-Nrf2-CES2 signaling axis and p62-Nrf2-PPARα signaling axis, respectively, while berberine suppressed de novo lipogenesis through downregulating the expression of acetyl-CoA carboxylase and fatty acid synthetase, along with enrichment of lipid metabolism-related Bacteroidaceae (family) and Bacteroides (genus). Of note, the combined use of bicyclol and berberine did not influence each other but enhanced the overall therapeutic role in the amelioration of NAFLD. Conclusion: Combined use of bicyclol and berberine might be a new available strategy to treat NAFLD.

Bicyclol Alleviates Atherosclerosis by Manipulating Gut Microbiota.

Atherosclerosis (AS) is associated with high morbidity and mortality, thus imposing a growing burden on modern society. Herb-derived bicyclol (BIC) is a versatile bioactive compound that can be used to treat AS. However, its efficacy in AS is not yet described. Here, it is shown that BIC normalizes gut microflora dysbiosis induced by a high fat diet in Apoe(-/-) mice. Metagenome-wide association study analysis verifies that the modulation on carbohydrate-active enzymes and short-chain fatty acid generating genes in gut flora is among the mechanisms. The gut healthiness, especially the gut immunity and integrity, is restored by BIC intervention, leading to improved systemic immune cell dynamic and liver functions. Accordingly, the endothelial activation, macrophage infiltration, and cholesterol ester accumulation in the aortic arch are alleviated by BIC to lessen the plaque onset. Moreover, it is proved that the therapeutic effect of BIC on AS is transmissible by fecal microbiota transplantation. The current study, for the first time, demonstrates the antiatherosclerotic effects of BIC and shows that its therapeutic value can at least partially be attributed to its manipulation of gut microbiota.

Bicyclol Alleviates Signs of BDL-Induced Cholestasis by Regulating Bile Acids and Autophagy-Mediated HMGB1/p62/Nrf2 Pathway.

Cholestasis is a liver disease characterized by the accumulation of toxic bile salts, bilirubin, and cholesterol, resulting in hepatocellular damage. Recent findings have revealed several key steps of cholestasis liver injury including the toxicity of bile acids and accumulation of proinflammatory mediator. In this study, we investigated the protective effect of bicyclol in cholestasis caused by bile duct ligation (BDL), as well as relevant mechanisms. Bicyclol attenuated liver damage in BDL mice by increasing the levels of hydrophilic bile acid such as α-MCA and ß-MCA, regulating bile acid-related pathways and improving histopathological indexes. High-mobility group box 1 (HMGB1) is an extracellular damage-associated molecular pattern molecule which can be used as biomarkers of cells and host defense. Bicyclol treatment decreased extracellular release of HMGB1. In addition, HMGB1 is also involved in regulating autophagy in response to oxidative stress. Bicyclol promoted the lipidation of LC3 (microtubule-associated protein 1 light chain 3)-Ⅱ to activate autophagy. The nuclear factor, E2-related factor 2 (Nrf2) and its antioxidant downstream genes were also activated. Our results indicate that bicyclol is a promising therapeutic strategy for cholestasis by regulating the bile acids and autophagy-mediated HMGB1/p62/Nrf2 pathway.

Bicyclol ameliorates nonalcoholic fatty liver disease in mice via inhibiting MAPKs and NF-κB signaling pathways.

Bicyclol has been approved as an anti-inflammatory, hepatoprotective drug in China to treat various forms of hepatitis. However, the role of bicyclol in non-alcoholic fatty liver disease (NAFLD) is unknown. In this study, NAFLD model was established by feeding mice with high fat diet (HFD) for 16 weeks, and bicyclol (25 and 50 mg/kg) were orally administered for the last 4 weeks. Although bicyclol treatment did not change the body weight of mice, bicyclol administration significantly improved HFD-induced dyslipidemia, NAFLD activity score, hepatic apoptosis, systemic and hepatic inflammation, and liver fibrosis in the mice. Moreover, bicyclol treatment significantly inhibited HFD-induced activation of MAPKs and NF-κB signaling pathways that may mediate the inflammatory responses. Further in vitro studies showed that bicyclol pretreatment markedly ameliorated PA-induced inflammatory responses in human hepatocyte HL-7702 cells and mouse peritoneal macrophages through inhibiting MAPKs and NF-κB signaling pathways. These data indicated that bicyclol may have the potency to treat NAFLD by reducing inflammation.