anti-NAFLD Effects Research Articles

Polysaccharides from hawthorn fruit alleviate high-fat diet-induced NAFLD in mice by improving gut microbiota dysbiosis and hepatic metabolic disorder.

Hawthorn fruit, renowned as both a functional food and herbal medicine with lipid-lowering effects, is abundant in polysaccharides. However, there is limited research on the effects and mechanisms of hawthorn fruit polysaccharides (HP) in addressing non-alcoholic fatty liver disease (NAFLD). This study aims to investigate the effects of HP on NAFLD both in vivo and in vitro, and to elucidate the underlying mechanisms by which HP exerts its anti-NAFLD activity. NAFLD mice induced by a high-fat diet were employed as the in vivo model, while oleate/palmitate-induced HepG2 cells served as the in vitro model. H&E and Oil Red O staining were employed to examine fat accumulation in hepatocytes. Serum aminotransferase (ALT), aspartate aminotransferase (AST), hepatic malondialdehyde (MDA), superoxide dismutase (SOD), interleukin-1 beta (IL-1β), and interleukin-6 (IL-6) were measured using corresponding ELISA kits. Hepatic metabolomics analysis based on UHPLC-QTOF/MS was utilized to examine the role of HP in improving hepatic metabolic disorders. 16S rRNA sequencing was conducted to explore the effect of HP in alleviating gut microbiota dysbiosis. GC-MS was applied to detect short-chain fatty acids (SCFAs) to clarify the impact of HP in NAFLD mice. HP significantly inhibited weight gain and hepatic fat accumulation in NAFLD mice. The reduction in serum ALT and AST levels indicated that HP mitigated liver function damage, while the decreased MDA levels and increased SOD activity suggested that HP alleviated hepatic oxidative stress. Furthermore, HP diminished the release of inflammatory cytokines such as IL-1β and IL-6 in the liver. HP significantly regulated metabolic pathways related to amino acids, lipids, and vitamins. Key metabolites such as l-tyrosine, urocanic acid, undecanedioic acid, oleamide, vitamin A, and vitamin B7 were restored to near-normal levels under the regulatory effects of HP. Gut microbiota dysbiosis in NAFLD mice was also ameliorated by HP, with genera such as unclassified_f__Lachnospiraceae and Dubosiella being notably affected. Correlation analysis indicated a significant correlation between the regulatory effects of HP on liver metabolism and gut microbiota. Additionally, HP showed no effect in vitro but increased acetic acid level in the gut of NAFLD mice. These findings demonstrate that HP exhibits its anti-NAFLD effects, including alleviating lipid accumulation, liver dysfunction, oxidative stress, and inflammation. Mechanistically, HP primarily improves gut microbiota dysbiosis, thereby elevating intestinal SCFA levels and restoring hepatic metabolic disorders in NAFLD mice.

Unlocking the therapeutic potential of canagliflozin in NAFLD: Insights into AMPK/SIRT1-mediated lipophagy.

This study aimed to explore the possible anti-NAFLD effects of CANA in rats and HepG2 cells, focusing on AMPK/SIRT1-mediated lipophagy. Wistar rats were assigned to four groups: control group, NAFLD group, NAFLD+CANA group, and NAFLD+CANA+chloroquine (CQ) group, where CQ served as autophagy inhibitor. HepG2 cells were also divided into four groups: control group, NAFLD group, NAFLD+CANA group, and NAFLD+CANA+compound C (Comp C) group, where Comp C served as AMPK inhibitor. The histopathological examination showed that CANA alleviated hepatic and intracellular lipid deposition in rats and HepG2 cells. CANA induced lipophagy by increasing LC3-II levels and lowering both p62 and perilipin 2 levels in rats and HepG2 cells, in addition to decreasing mTOR protein expression in rats' livers. These outcomes were associated with upregulation of the lipophagy regulator Rab7 and downregulation of the ER stress-related protein CHOP. CANA enhanced autophagic engulfment of lipid droplets while decreased ER stress and mitochondrial damage in rats' livers, as demonstrated by TEM. In rats, CANA improved hyperglycemia, hyperinsulinemia, dyslipidemia, and obesity. In HepG2 cells, CANA's effects were linked to increased phosphorylated AMPK level and enhanced SIRT1 level and expression. However, blocking lipophagy in rats and AMPK in HepG2 cells markedly weakened CANA's protective effects against NAFLD. CANA ameliorated NAFLD via enhancing AMPK/SIRT1-mediated lipophagy, suggesting its potential as a therapeutic intervention for this metabolic disorder.

Ellagic acid alleviates high-fructose diet-induced non-alcoholic fatty liver disease by modulating liver metabolic profiles and gut microbiota

This study integrated analyses of gut microbiota and metabolomics to investigate the impact of ellagic acid (EA) on non-alcoholic fatty liver disease (NAFLD). Compared to the high-fructose diet (HFruD) group, the EA group exhibited reduced body weight and fat mass, alongside improvements in blood glucose and lipid metabolism. Liver metabolomics analysis revealed that EA increased the abundance of metabolites in pathways related to unsaturated fatty acids, amino acids and bile acids. Furthermore, EA induced alterations in the composition and structure of gut microbiota, notably decreasing bacterial genera enriched by HFruD while promoting beneficial bacteria such as Faecalibaculum. Correlation analysis demonstrated significant associations among NAFLD markers, gut microbiota and liver metabolites influenced by EA. This study provides new insights into the anti-NAFLD effects of EA, suggesting EA as a promising nutraceutical for improving NAFLD.

Integrated lipidomic and transcriptomics to explore the effects of ethyl acetate extract of Herpetospermum pedunculosum on nonalcoholic fatty liver disease in mice

Ethnopharmacological relevanceHerpetospermum pedunculosum (Ser.) C.B. Clarke (HP), a traditional Tibetan medicine used to treat hepatobiliary diseases, was confirmed that lignans-enriched ethyl acetate extract of HP (EAHP) could alleviate the hepatic injury by modern pharmacological evidence. However, the effects and potential mechanisms of EAHP against nonalcoholic fatty liver disease (NAFLD) are still unknown. Aim of the studyTo reveal the effects of EAHP on NAFLD and explore the potential mechanisms from the perspective of lipidomics and transcriptomics. Materials and methodsUPLC‒Q–TOF‒MS analysis was carried out to investigate the chemical components of EAHP. A Choline-deficient, L-amino acid defined, high fat diet (CDAHFD) was used to establish a NAFLD mouse model. The anti-NAFLD effects of various dosages of EAHP were evaluated by biochemical indexes and histological analysis. Hepatic lipidomic and transcriptomic analysis and multiple bioinformatics methods were used to screen biomarkers and signaling pathways. The levels of the corresponding genes were verified by qPCR. Results36 kinds of compounds were identified by UPLC‒Q–TOF‒MS analysis. Oral treatment with EAHP significantly decrease the liver index and the levels of ALT and AST in the serum. The measurements lipid content and Oil Red O staining results suggested that EAHP ameliorated lipid metabolism disorders by reducing the content of TG and LDL-C, increasing HDL-C in the liver. H&E staining and ELISA revealed that EAHP restored hepatic inflammatory infiltration and decrease the levels of IL-1β, IL-6, TNF-α, and increase IL-10 in the serum. Lipidomic analysis showed that EAHP could regulate CDAHFD-induced lipid metabolic disorder. The different lipid metabolites included TG, phosphatidyl choline (PC), diacylglycerol (DG), phosphatidylethanolamine (PE), phosphatidylinositol (PI), ceramide (Cer). Transcriptomic analysis revealed that Bmp8b, Nbl1, Rgma, Sphk1, Thbs1, and Ugt8a were important regulators, which were associated with TGF-β signaling pathway and sphingolipid metabolism. The expressions of above genes detected by were qPCR consistent with transcriptomic data. ConclusionsThe ameliorative effects of EAHP on NAFLD are potentially attributable to the regulation of sphingolipid metabolism and TGF-β signaling pathway, etc., which results in abnormal hepatic lipid metabolism and inflammatory response.

Genetically engineered long-acting Esculentin-2CHa(1−30) fusion protein with potential applicability for the treatment of NAFLD

Esculentin-2CHa(1–30) (‟ESC”) has been reported as a potent anti-diabetic peptide with little toxicity. However, its very short plasma residence time severely limits the therapeutic efficacy. To address this issue, we genetically engineered a fusion protein of tandem trimeric ESC with an albumin binding domain (ABD) and a fusion partner, SUMO (named ‟SUMO-3×ESC-ABD”). The SUMO-3×ESC-ABD, successfully produced from E. coli, showed low cellular and hemolytic toxicity while displaying potent activities for the amelioration of hyperglycemia as well as non-alcoholic fatty liver disease (NAFLD) in vitro. In animal studies, the estimated plasma half-life of SUMO-3×ESC-ABD was markedly longer (427-fold) than that of the ESC peptide. In virtue of the extended plasma residence, the SUMO-3×ESC-ABD could produce significant anti-hyperglycemic effects that lasted for >2 days, while both the ESC or ESC-ABD peptides elicited little effects. Further, twice-weekly treatment for 10 weeks, the SUMO-3×ESC-ABD displayed significant improvement in blood glucose control with a reduction in body weight. Most importantly, a significant improvement in the conditions of NAFLD was observed in the SUMO-3×ESC-ABD-treated mice. Along the systemic effects (by improved glucose tolerance and body weight reduction), direct inhibition of the hepatocyte lipid uptake was suggested as the major mechanism of the anti-NAFLD effects. Overall, this study demonstrated the utility of the long-acting SUMO-3×ESC-ABD as a potent drug candidate for the treatment of NAFLD.

Chrysanthemum morifolium Ramat extract and probiotics combination ameliorates metabolic disorders through regulating gut microbiota and PPARα subcellular localization

BackgroundChrysanthemum morifolium Ramat, a traditional Chinese medicine, has the effects on liver clearing, vision improving, and anti-inflammation. C. morifolium and probiotics have been individually studied for their beneficial effects on metabolic diseases. However, the underlying molecular mechanisms were not completely elucidated. This study aims to elucidate the potential molecular mechanisms of C. morifolium and probiotics combination (CP) on alleviating nonalcoholic fatty liver disease (NAFLD) and the dysregulation of glucose metabolism in high-fat diet (HFD)-fed mice.MethodsThe therapeutic effect of CP on metabolism was evaluated by liver histology and serum biochemical analysis, as well as glucose tolerance test. The impact of CP on gut microbiota was analyzed by 16S rRNA sequencing and fecal microbiota transplantation. Hepatic transcriptomic analysis was performed with the key genes and proteins validated by RT-qPCR and western blotting. In addition, whole body Pparα knockout (Pparα−/−) mice were used to confirm the CP-mediated pathway.ResultsCP supplementation ameliorated metabolic disorders by reducing body weight and hepatic steatosis, and improving glucose intolerance and insulin resistance in HFD fed mice. CP intervention mitigated the HFD-induced gut microbiota dysbiosis, which contributed at least in part, to the beneficial effect of improving glucose metabolism. In addition, hepatic transcriptomic analysis showed that CP modulated the expression of genes associated with lipid metabolism. CP downregulated the mRNA level of lipid droplet-binding proteins, such as Cidea and Cidec in the liver, leading to more substrates for fatty acid oxidation (FAO). Meanwhile, the expression of CPT1α, the rate-limiting enzyme of FAO, was significantly increased upon CP treatment. Mechanistically, though CP didn’t affect the total PPARα level, it promoted the nuclear localization of PPARα, which contributed to the reduced expression of Cidea and Cidec, and increased expression of CPT1α, leading to activated FAO. Moreover, whole body PPARα deficiency abolished the anti-NAFLD effect of CP, suggesting the importance of PPARα in CP-mediated beneficial effect.ConclusionThis study revealed the hypoglycemic and hepatoprotective effect of CP by regulating gut microbiota composition and PPARα subcellular localization, highlighting its potential for therapeutic candidate for metabolic disorders.

From petals to healing: consolidated network pharmacology and molecular docking investigations of the mechanisms underpinning Rhododendron arboreum flower's anti-NAFLD effects.

Rhododendron arboreum: Sm., also known as Burans is traditionally used as an anti-inflammatory, anti-diabetic, hepatoprotective, adaptogenic, and anti-oxidative agent. It has been used since ancient times in Indian traditional medicine for various liver disorders. However, the exact mechanism behind its activity against NAFLD is not known. The aim of the present study is to investigate the molecular mechanism of Rhododendron arboreum flower (RAF) in the treatment of NAFLD using network pharmacology and molecular docking methods. Bioactives were also predicted for their drug-likeness score, probable side effects and ADMET profile. Protein-protein interaction (PPI) data was obtained using the STRING platform. For the visualisation of GO analysis, a bioinformatics server was employed. Through molecular docking, the binding affinity between potential targets and active compounds were assessed. A total of five active compounds of RAF and 30 target proteins were selected. The targets with higher degrees were identified through the PPI network. GO analysis indicated that the NAFLD treatment with RAF primarily entails a response to the fatty acid biosynthetic process, lipid metabolic process, regulation of cell death, regulation of stress response, and cellular response to a chemical stimulus. Molecular docking and molecular dynamic simulation exhibited that rutin has best binding affinity among active compounds and selected targets as indicated by the binding energy, RMSD, and RMSF data. The findings comprehensively elucidated toxicity data, potential targets of bioactives and molecular mechanisms of RAF against NAFLD, providing a promising novel strategy for future research on NAFLD treatment.

Shenling Baizhu powder alleviates non-alcoholic fatty liver disease by modulating autophagy and energy metabolism in high-fat diet-induced rats

BackgroundNon-alcoholic fatty liver disease (NAFLD) has emerged as a burgeoning health problem worldwide, but no specific drug has been approved for its treatment. Shenling Baizhu powder (SL) is extensively used to treat NAFLD in Chinese clinical practice. However, the therapeutic components and pharmacological mechanisms of SL against NAFLD have not been thoroughly investigated. PurposeThis study aimed to investigate the pharmacological impact and molecular mechanism of SL on NAFLD. MethodsFirst, we established an animal model of NAFLD by high-fat diet (HFD) feeding, and evaluated the therapeutic efficacy of SL on NAFLD by physiological, biochemical, pathological, and body composition analysis. Next, the effect of SL on autophagic flow in NAFLD rats was evaluated by ultrastructure, immunofluorescence staining, and western blotting. Moreover, an integrated strategy of targeted energy metabolomics and network pharmacology was performed to characterize autophagy-related genes and explore the synergistic effects of SL active compounds. UPLC-MS/MS, molecular docking combined with in vivo and in vitro experiments were conducted to verify the key compounds and genes. Finally, a network was established among SL-herb-compound-genes-energy metabolites-NAFLD, which explains the complicated regulating mechanism of SL on NAFLD. ResultsWe discovered that SL decreased hepatic lipid accumulation, hepatic steatosis, and insulin resistance, and improved systemic metabolic disorders and pathological abnormalities. Subsequently, an integrated strategy of targeted energy metabolomics and network pharmacology identified quercetin, ellagic acid, kaempferol, formononetin, stigmasterol, isorhamnetin and luteolin as key compounds; catalase (CAT), AKT serine/threonine kinase 1 (AKT), nitric oxide synthase 3 (eNOS), NAD(P)H quinone dehydrogenase 1 (NQO1), heme oxygenase 1 (HO-1) and hypoxia-inducible factor 1 subunit alpha (HIF-1α) were identified as key genes; while nicotinamide adenine dinucleotide phosphate (NADP) and succinate emerged as key energy metabolites. Mechanistically, we revealed that SL may exert its anti-NAFLD effect by inducing autophagy activation and forming a comprehensive regulatory network involving key compounds, key genes, and key energy metabolites, ultimately alleviating oxidative stress, endoplasmic reticulum stress, and mitochondrial dysfunction. ConclusionOur study demonstrated the therapeutic effect of SL in NAFLD models, and establishes a basis for the development of potential products from SL plant materials for the treatment of NAFLD.

Adrenarche-accompanied rise of adrenal sex steroid precursors prevents NAFLD in Young Female rats by converting into active androgens and inactivating hepatic Srebf1 signaling

BackgroundNon-alcoholic fatty liver disease (NAFLD) has rapidly become the most common cause of chronic liver disease in children and adolescents, but its etiology remains largely unknown. Adrenarche is a critical phase for hormonal changes, and any disturbance during this period has been linked to metabolic disorders, including obesity and dyslipidemia. However, whether there is a causal linkage between adrenarche disturbance and the increasing prevalence of NAFLD in children remains unclear.ResultsUsing the young female rat as a model, we found that the liver undergoes a transient slowdown period of growth along with the rise of adrenal-derived sex steroid precursors during adrenarche. Specifically blocking androgen actions across adrenarche phase using androgen receptor antagonist flutamide largely increased liver weight by 47.97% and caused marked fat deposition in liver, thus leading to severe NAFLD in young female rats. Conversely, further administrating nonaromatic dihydrotestosterone (DHT) into young female rats across adrenarche phase could effectively reduce liver fat deposition. But, administration of the aromatase inhibitor, formestane across adrenarche had minimal effects on hepatic de novo fatty acid synthesis and liver fat deposition, suggesting adrenal-derived sex steroid precursors exert their anti-NAFLD effects in young females by converting into active androgens rather than into active estrogens. Mechanistically, transcriptomic profiling and integrated data analysis revealed that active androgens converted from the adrenal sex steroid precursors prevent NAFLD in young females primarily by inactivating hepatic sterol regulatory element-binding transcription factor 1 (Srebf1) signaling.ConclusionsWe firstly evidenced that adrenarche-accompanied rise of sex steroid precursors plays a predominant role in preventing the incidence of NAFLD in young females by converting into active androgens and inactivating hepatic Srebf1 signaling. Our novel finding provides new insights into the etiology of NAFLD and is crucial in developing effective prevention and management strategies for NAFLD in children.

Diosgenin as a substitute for cholesterol alleviates NAFLD by affecting CYP7A1 and NPC1L1-related pathway

BackgroundNonalcoholic fatty liver disease (NAFLD) rapidly becomes the leading cause of end-stage liver disease or liver transplantation. Nowadays, there has no approved drug for NAFLD treatment. Diosgenin as the structural analogue of cholesterol attenuates hypercholesterolemia by inhibiting cholesterol metabolism, which is an important pathogenesis in NAFLD progression. However, there has been no few report concerning its effects on NAFLD so far. MethodsUsing a high-fat diet & 10% fructose-feeding mice, we evaluated the anti-NAFLD effects of diosgenin. Transcriptome sequencing, LC/MS analysis, molecular docking simulation, molecular dynamics simulations and Luci fluorescent reporter gene analysis were used to evaluate pathways related to cholesterol metabolism. ResultsDiosgenin treatment ameliorated hepatic dysfunction and inhibited NAFLD formation including lipid accumulation, inflammation aggregation and fibrosis formation through regulating cholesterol metabolism. For the first time, diosgenin was structurally similar to cholesterol, down-regulated expression of CYP7A1 and regulated cholesterol metabolism in the liver (p < 0.01) and further affecting bile acids like CDCA, CA and TCA in the liver and feces. Besides, diosgenin decreased expression of NPC1L1 and suppressed cholesterol transport (p < 0.05). Molecular docking and molecular dynamics further proved that diosgenin was more strongly bound to CYP7A1. Luci fluorescent reporter gene analysis revealed that diosgenin concentration-dependently inhibited the enzymes activity of CYP7A1. ConclusionOur findings demonstrated that diosgenin was identified as a specific regulator of cholesterol metabolism, which pave way for the design of novel clinical therapeutic strategies.

A novel bioinformatics strategy to uncover the active ingredients and molecular mechanisms of Bai Shao in the treatment of non-alcoholic fatty liver disease.

Introduction: As a new discipline, network pharmacology has been widely used to disclose the material basis and mechanism of Traditional Chinese Medicine in recent years. However, numerous researches indicated that the material basis of TCMs identified based on network pharmacology was the mixtures of beneficial and harmful substances rather than the real material basis. In this work, taking the anti-NAFLD (non-alcoholic fatty liver disease) effect of Bai Shao (BS) as a case, we attempted to propose a novel bioinformatics strategy to uncover the material basis and mechanism of TCMs in a precise manner. Methods: In our previous studies, we have done a lot work to explore TCM-induced hepatoprotection. Here, by integrating our previous studies, we developed a novel computational pharmacology method to identify hepatoprotective ingredients from TCMs. Then the developed method was used to discover the material basis and mechanism of Bai Shao against Non-alcoholic fatty liver disease by combining with the techniques of molecular network, microarray data analysis, molecular docking, and molecular dynamics simulation. Finally, literature verification method was utilized to validate the findings. Results: A total of 12 ingredients were found to be associated with the anti-NAFLD effect of BS, including monoterpene glucosides, flavonoids, triterpenes, and phenolic acids. Further analysis found that IL1-β, IL6, and JUN would be the key targets. Interestingly, molecular docking and molecular dynamics simulation analysis showed that there indeed existed strong and stable binding affinity between the active ingredients and the key targets. In addition, a total of 23 NAFLD-related KEGG pathways were enriched. The major biological processes involved by these pathways including inflammation, apoptosis, lipid metabolism, and glucose metabolism. Of note, there was a great deal of evidence available in the literature to support the findings mentioned above, indicating that our method was reliable. Discussion: In summary, the contributions of this work can be summarized as two aspects as follows. Firstly, we systematically elucidated the material basis and mechanism of BS against NAFLD from multiple perspectives. These findings further enhanced the theoretical foundation of BS on NAFLD. Secondly, a novel computational pharmacology research strategy was proposed, which would assist network pharmacology to uncover the scientific connotation TCMs in a more precise manner.

Biotransformed bear bile powder alleviates diet-induced nonalcoholic fatty liver disease in mice by regulating the gut microbiota and reversing lipid metabolism

BackgroundThe pathogenesis of metabolic syndrome was strongly associated with compromised metabolism homeostasis and gut microbiota imbalance. NAFLD is a progressive metabolic liver disease for which effective interventions are lacking. Bile acids exhibited appreciable metabolic regulatory effects and selective antimicrobial activity. Aim of the studyThis study was designed to investigate the effect of BBBP, which mainly contained bile acids, on NAFLD from the perspectives of gut microbiota and metabolomics. Materials and methodsThe present study was initiated on the anti-NAFLD effect of BBBP in HFD-fed mice. The efficacy of BBBP was evaluated by mice phenotypes, liver histopathological analysis and serum lipid and glucose levels. The activation of bile acid receptors such as Nr1h4, Nr1i2 and S1pr2 were detected by qRT-PCR analysis. Subsequently, untargeted metabolomics coupled with microbiomics was used to explore the mechanism of BBBP against NAFLD. Human L02 hepatocytes induced by OA and PA were used to investigate the effect of GABA on reducing lipid accumulation in vitro. ResultsBBBP significantly and dose-dependently alleviated the obese phenotype, lipid accumulation and liver injury in mice subjected to 18 weeks HFD diet. Untargeted metabolomics and microbiomics analysis revealed that BBBP could alleviate the disturbance of lipid and amino acid metabolism and the imbalance of gut microbiota. Furthermore, BBBP oral gavage activated liver bile acid receptors, as indicated by elevated mRNA levels of Nr1h4, Nr1i2 and S1pr2. Surprisingly, we determined that BBBP, which mainly contained bile acids that possessed antimicrobial activity, could promote the growth of Lactobacillus. Correlation analysis showed a remarkable correlation between Lactobacillus and endogenous metabolites such as valine, serine, glutamine, et al. Among them, GABA which could be produced by Lactobacillus significantly reduced the lipid accumulation in L02 cells. ConclusionsThe role of BBBP in regulating lipid metabolism might be achieved by activating bile acid receptors, or partially by promoting the levels of Lactobacillus and its metabolites such as GABA. Our study provided evidence that BBBP could be a novel therapeutic candidate for the treatment of NAFLD.

Astragalus polysaccharide attenuates nonalcoholic fatty liver disease through THDCA in high-fat diet-fed mice

Ethnopharmacological relevanceAstragalus polysaccharide (APS) extracted from Astragalus membranaceus (Fisch.) Bunge was proven to be effective in preventing high-fat diet (HFD) induced nonalcoholic fatty liver disease (NAFLD). However, the exact mechanisms were not completely elucidated. Aim of the studyThe aim was to reveal the mechanisms of APS on preventing NAFLD from the aspects of regulating bile acids (BAs) homeostasis. Materials and methodsSerum and liver BAs in HFD fed mice with or without APS intervention were quantified with an ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) system. The effect of APS on hepatic proteins involved in BAs synthesis were analyzed with Western blot. Finally, the effect of identified taurohyodeoxycholic acid (THDCA) that was significantly increased by APS on hepatic triglyceride (TG) accumulation was explored in vivo and in vitro. ResultsAPS regulated serum and liver BA profiles in HFD fed mice, especially increased serum THDCA. The levels of hepatic cholesterol 7a-hydroxylase (CYP7A1) and sterol 12a-hydroxylase (CYP8B1) which catalyzed the classical BAs synthesis pathway were significantly decreased by APS, while oxysterol 7a-hydroxylase (CYP7B1) which catalyzed the alternative BAs synthesis pathway was significantly increased by APS. THDCA reduced HFD-induced hepatic lipid accumulation and improved glucose homeostasis in mice, and decreased TG level in palmitic acid/oleic acid treated alpha mouse liver 12 (AML-12) cells. THDCA significantly downregulated the protein level of cluster of differentiation 36 (CD36) involved in fatty acid transport into the liver. Importantly, THDCA showed similar effect with APS in upregulating hepatic CYP7B1 and downregulating CYP7A1. ConclusionThis study revealed the protective effect of APS on NAFLD was associated with the regulation on BA profiles, and proved the potential anti-NAFLD effect of THDCA, highlighting the involvement of BA metabolism in efficacy of herb-derived polysaccharides on metabolism.

Comparative analysis of the synergetic effects of Diwuyanggan prescription on high fat diet-induced non-alcoholic fatty liver disease using untargeted metabolomics

Non-alcoholic fatty liver disease (NAFLD) is one of the most common chronic liver disorders worldwide and had no approved pharmacological treatments. Diwuyanggan prescription (DWYG) is a traditional Chinese medicine preparation composed of 5 kinds of herbs, which has been used for treating chronic liver diseases in clinic. Whereas, the synergistic mechanism of this prescription for anti-NAFLD remains unclear. In this study, we aimed to demonstrate the synergetic effect of DWYG by using the disassembled prescriptions and untargeted metabolomics research strategies. The therapeutic effects of the whole prescription of DWYG and the individual herb were divided into six groups according to the strategy of disassembled prescriptions, including DWYG, Artemisia capillaris Thunb. (AC), Curcuma longa L. (CL), Schisandra chinensis Baill. (SC), Rehmannia glutinosa Libosch. (RG) and Glycyrrhiza uralensis Fisch. (GU) groups. The high fat diets-induced NAFLD mice model was constructed to evaluate the efficacy effects of DWYG. An untargeted metabolomics based on the UPLC-QTOF-MS/MS approach was carried out to make clear the synergetic effect on the regulation of metabolites dissecting the united mechanisms. Experimental results on animals revealed that the anti-NAFLD effect of DWYG prescription was better than the individual herb group in reducing liver lipid deposition and restoring the abnormality of lipidemia. In addition, further metabolomics analysis indicated that 23 differential metabolites associated with the progression of NAFLD were identified and 19 of them could be improved by DWYG. Compared with five single herbs, DWYG showed the most extensive regulatory effects on metabolites and their related pathways, which were related to lipid and amino acid metabolisms. Besides, each individual herb in DWYG was found to show different degrees of regulatory effects on NAFLD and metabolic pathways. SC and CL possessed the highest relationship in the regulation of NAFLD. Altogether, these results provided an insight into the synergetic mechanisms of DWYG from the metabolic perspective, and also supported a scientific basis for the rationality of clinical use of this prescription.

Calenduloside E ameliorates non-alcoholic fatty liver disease via modulating a pyroptosis-dependent pathway.

Ethnopharmacological relevanceNon-alcoholic fatty liver disease (NAFLD) is a prevalent chronic liver condition that can have multiple underlying causes. There are no satisfactory chemical or biological drugs for the treatment of NAFLD. Longyasongmu, the bark and root of Aralia elata (Miq.) Seem, is used extensively in traditional Chinese medicine (TCM) and has been used in treating diverse liver diseases including NAFLD. Based on Aralia elata (Miq.) Seem as the main ingredient, Longya Gantai Capsules have been approved for use in China for the treatment of acute hepatitis and chronic hepatitis. Calenduloside E (CE), a natural pentacyclic triterpenoid saponin, is a significant component of saponin isolated from the bark and root of Aralia elata (Miq.) Seem. However, the role and mechanism of anti-NAFLD effects of CE is still unclear. Aim of the studyThe objective of this study was to examine the potential mechanisms underlying the protective effect of CE on NAFLD. Materials and methodsIn this study, an NAFLD model was established by Western diet in apoE−/− mice, followed by treatment with various doses of CE (5 mg/kg, 10 mg/kg). The anti-NAFLD effect of CE was assessed by the liver injury, lipid accumulation, inflammation, and pro-fibrotic phenotype. The mechanism of CE in ameliorating NAFLD was studied through transcriptome sequencing (RNA-seq). In vitro, the mouse hepatocytes (AML-12) were stimulated in lipid mixtures with CE and performed the exploration and validation of the relevant pathways using Western blot, immunofluorescence, etc. ResultsThe findings revealed a significant improvement in liver injury, lipid accumulation, inflammation, and pro-fibrotic phenotype upon CE administration. Furthermore, RNAseq analysis indicated that the primary pathway through which CE alleviates NAFLD involves pyroptosis-related inflammatory cascade pathways. Furthermore, it was observed that CE effectively suppressed inflammasome-mediated pyroptosis both in vivo and in vitro. Remarkably, the functional enrichment analysis of RNA-seq data revealed that the PI3K-Akt signaling pathway is the primarily Signaling transduction pathway modulated by CE treatment. Subsequent experimental outcomes provided further validation of CE's ability to hinder inflammasome-mediated pyroptosis through the inhibition of PI3K/AKT/NF-κB signaling pathway. ConclusionsThese findings present a novel pharmacological role of CE in exerting anti-NAFLD effects by inhibiting pyroptosis signaling pathways.

Berberine-silybin salt achieves improved anti-nonalcoholic fatty liver disease effect through regulating lipid metabolism

Ethnopharmacological relevanceBerberine (BBR) and silybin (SIY) are natural compounds obtained from Berberidaceae members and Silybum marianum (L.) Gaertn., respectively. These compounds have been demonstrated to regulate lipid metabolism and indue hepatoprotective effects, establishing their importance for the treatment of liver injury. Combination therapy has shown promise in treating ailments with complex pathophysiology, such as liver diseases. However, the inconsistent dissolution and poor absorption of BBR and SIY limit their efficacy. Aim of the studyThis study compared the salt formulation (BSS) and physical mixture (BSP) of BBR and SIY for their efficacy in treating nonalcoholic fatty liver disease (NAFLD). Materials and methodsThe formation of the BSS was confirmed using various techniques, including nuclear magnetic resonance spectroscopy, Fourier-transform infrared spectroscopy, differential scanning calorimetry, scanning electron microscopy, and powder X-ray diffractometry. In addition, dissolution, trans-epithelial permeability, and bioavailability experiments were conducted to evaluate the absorption and distribution of drugs. Pharmacodynamics and mechanisms were investigated through in vivo experiments. ResultsBSS form demonstrated synchronized dissolution of both components, unlike BSP. Additionally, the transepithelial permeability results revealed that BSS exhibited superior penetration and absorption of both BBR and SIY in comparison to BSP. Furthermore, BSS significantly increased the bioavailability of SIY in both plasma and the liver (2.2- and 4.5-fold, respectively) when compared with BSP. Moreover, BSS demonstrated a more potent inhibitory effect on lipid production in HepG2 cells than BSP. In mouse models (BALB/c) of NAFLD, BSS improved disease outcomes, as evidenced by decreased adipose levels, normalized blood lipid levels, and reduced liver parenchyma injury. Preliminary transcriptomics analysis suggested that BSS achieved its anti-NAFLD effect by regulating the expression of fatty acid transporter CD36, recombinant fatty acid binding protein 4, and stearyl coenzyme A dehydrogenase 1, which are associated with the synthesis and uptake of fatty acid-related proteins. ConclusionsThe study demonstrated that compared with physical mixing, salification improved the efficacy of BBR and SIY, as demonstrated in animal experiments. These findings provide valuable insights into the development of more effective treatments for NAFLD and provide new possibilities for combination therapies.

Hyodeoxycholic acid ameliorates nonalcoholic fatty liver disease by inhibiting RAN-mediated PPARα nucleus-cytoplasm shuttling

Nonalcoholic fatty liver disease (NAFLD) is usually characterized with disrupted bile acid (BA) homeostasis. However, the exact role of certain BA in NAFLD is poorly understood. Here we show levels of serum hyodeoxycholic acid (HDCA) decrease in both NAFLD patients and mice, as well as in liver and intestinal contents of NAFLD mice compared to their healthy counterparts. Serum HDCA is also inversely correlated with NAFLD severity. Dietary HDCA supplementation ameliorates diet-induced NAFLD in male wild type mice by activating fatty acid oxidation in hepatic peroxisome proliferator-activated receptor α (PPARα)-dependent way because the anti-NAFLD effect of HDCA is abolished in hepatocyte-specific Pparα knockout mice. Mechanistically, HDCA facilitates nuclear localization of PPARα by directly interacting with RAN protein. This interaction disrupts the formation of RAN/CRM1/PPARα nucleus-cytoplasm shuttling heterotrimer. Our results demonstrate the therapeutic potential of HDCA for NAFLD and provide new insights of BAs on regulating fatty acid metabolism.

Thyme (Thymus quinquecostatus Celak) Polyphenol-Rich Extract (TPE) Alleviates HFD-Induced Liver Injury in Mice by Inactivating the TLR4/NF-κB Signaling Pathway through the Gut-Liver Axis.

Non-alcoholic fatty liver disease (NAFLD) represents a significant and urgent global health concern. Thyme (Thymus quinquecostatus Celak) is a plant commonly used in cuisine and traditional medicine in Asian countries and possesses potential liver-protective properties. This study aimed to assess the hepatoprotective effects of thyme polyphenol-rich extract (TPE) on high-fat diet (HFD)-induced NAFLD and further explore possible mechanisms based on the gut-liver axis. HFD-induced liver injury in C57 mice is markedly ameliorated by TPE supplementation in a dose-dependent manner. TPE also regulates the expression of liver lipid metabolic genes (i.e., Hmgcr, Srebp-1, Fasn, and Cyp7a1), enhancing the production of SCFAs and regulating serum metabolites by modulating gut microbial dysbiosis. Furthermore, TPE enhances the intestinal barrier function and alleviates intestinal inflammation by upregulating tight junction protein expression (i.e., ZO-1 and occluding) and inactivating the intestinal TLR4/NF-κB pathway in HFD-fed mice. Consequently, gut-derived LPS translocation to the circulation was blocked, the liver TLR4/NF-κB signaling pathway was repressed, and subsequent pro-inflammatory cytokine production was restrained. Conclusively, TPE might exert anti-NAFLD effects through the gut-liver axis and has the potential to be used as a dietary supplement for the management of NAFLD.

The identification of material basis of Si Miao Formula effective for attenuating non-alcoholic fatty liver disease

Ethnopharmacological relevanceThe Si Miao Formula (SMF), a traditional Chinese medicine, originated from the "Cheng Fang Bian Du" during the Qing Dynasty and is commonly employed for the treatment of gout and hyperuricemia. We have demonstrated the anti-NAFLD effect of SMF by regulating hepatic lipid metabolism in high fat and high sucrose (HFHS) feeding mice in our previous report. However, the material basis of SMF for its anti-NAFLD effect remains unknown. Aim of the studyTo compare the effeciacy of different components of SMF and identify the material basis for its anti-NAFLD effect. Materials and methodsIn the present study, a “Leave-one out” strategy was adopted by removing one herb from SMF each time, and the anti-NAFLD effects of four decomposed recipes containing three herbs were evaluated in C57BL/6J mice fed with an HFHS diet for 16 weeks. The chemical components of SMF and the absorbed entities in serum were assayed using UHPLC-Q-Exactive-Orbitrap HRMS. Finally, a new chemical combination with four compounds (berberine, betaine, caffeic acid, p-coumaric acid, 2:2:1:1) were generated (SMF component composition, SMF_CC), and its anti-NAFLD effect was evaluated by comparing with the original SMF in the mouse model. ResultsVarified effects on NAFLD mice were observed among the decomposed recipes of SMF, while the original SMF showed advantages over its decomposed recipes. A total of 111 chemicals were identified from SMF, and 21 of them were detected in serum after oral administration of SMF. Comparing to SMF, SMF_CC showed comparable anti-NAFLD effect in HFHS-diet-fed mice, which was associated with the inhibition of hepatic fatty acid synthesis and transport, as well as inflammation. ConclusionOur current results suggested that the original SMF was better than its decomposed recipes in NAFLD management, and the derived SMF_CC was also effective in inhibiting NAFLD formation, highlighting its potential of being a novel natural agent for NAFLD therapy.

Exogenous galanin alleviates hepatic steatosis by promoting autophagy via the AMPK-mTOR pathway

Defective autophagy-induced intracellular lipid degradation is causally associated with non-alcoholic fatty liver disease (NAFLD) development. Therefore, agents that can restore autophagy may have potential clinical application prospects on this public health issue. Galanin (GAL) is a pleiotropic peptide that regulates autophagy and is a potential drug for the treatment of NAFLD. In this study, we used an MCD-induced NAFLD mouse model in vivo and an FFA-induced HepG2 hepatocyte model in vitro to evaluate the anti-NAFLD effect of GAL. Exogenous GAL supplementation significantly attenuated lipid droplet accumulation and suppressed hepatocyte TG levels in mice and cell models. Mechanistically, Galanin-mediated reduction of lipid accumulation was positively correlated with upregulated p-AMPK, as evidenced by upregulated protein expressions of fatty acid oxidation-related gene markers (PPAR-α and CPT1A), upregulated expressions of the autophagy-related marker (LC3B), and downregulated autophagic substrate p62 levels. In FFA-treated HepG2 cells, activation of fatty acid oxidation and autophagy-related proteins by galanin was reversed by autophagy inhibitors, chloroquine, and the AMPK inhibitor. Galanin ameliorates hepatic fat accumulation by inducing autophagy and fatty acid oxidation via the AMPK/mTOR pathway.