Astragaloside IV ameliorates diet-induced hepatic steatosis in obese mice by inhibiting intestinal FXR via intestinal flora remodeling.

BACKGROUND: Nonalcoholic fatty liver disease (NAFLD) is a major clinical and public health burden worldwide with no established pharmacological therapy. Changes in the intestinal flora and associated metabolite bile acids (BAs) have been described in NAFLD. Astragaloside IV (AS-IV) is a low drug permeability saponin with protective effects against multiple diseases. However, the specific mechanism underlying the involvement of AS-IV in the regulation of NAFLD is yet to be clarified. PURPOSE: This study aimed to investigate the effect of AS-IV on NAFLD and explore whether intestinal flora was involved. METHODS: The effect of AS-IV was evaluated on high-fat diet-fed mice. Real-time PCR, immunohistochemistry, immunofluorescence, and biochemical analyses were performed. 16S rRNA gene sequencing and UPLC-TQMS were used to determine the alterations in the intestinal flora and concentration of BAs. Fecal microbiota transplantation (FMT) and intestine-specific farnesoid X receptor (FXR) knockout were also performed. RESULTS: AS-IV treatment alleviated diet-induced metabolic impairments, particularly hepatic steatosis. These changes occurred in the setting of decreased intestinal bile salt hydrolase (BSH)-expressing flora. Further analysis showed that the reduced BSH activity increased intestinal tauro-ß-muricholic acid levels, an inhibitor of intestinal FXR. Inhibition of intestinal FXR signaling by AS-IV was accompanied by decreased expression of intestinal fibroblast growth factor 15 and subsequent hepatic FXR activation as well as increased glucagon-like peptide-1 and decreased ceramide production, all of which contribute to the inhibition of sterol regulatory element-binding protein-1c-mediated hepatic steatosis. Furthermore, intestine-specific Fxr knockout and FMT further demonstrated an FXR- and intestinal flora-dependent preventive effect of AS-IV on hepatic steatosis. CONCLUSION: These results show that the changes in intestinal flora and BAs serve an essential role in the remission of hepatic steatosis by AS-IV, thereby suggesting that AS-IV may be used as a prebiotic agent to provide viable treatment for NAFLD.

Phillygenin ameliorates nonalcoholic fatty liver disease via TFEB-mediated lysosome biogenesis and lipophagy.

BACKGROUND: Lipophagy is an autophagic process, which delivers the intracellular lipid droplets to the lysosomes for degradation. Recent studies revealed that the impairment of lysosomal biogenesis and autophagic flux led to dysregulation of lipophagy in hepatocytes, which exacerbated the development of nonalcoholic fatty liver disease (NAFLD). Therefore, agents restoring autophagic flux and lipophagy in hepatocytes may have therapeutic potential against this increasingly prevalent disease. Phillygenin (PHI), a lignin extracted from Forsythia suspense, exerts hepatoprotective and anti-inflammatory effects. However, the effect of PHI on NAFLD remains unknown. PURPOSE: This study aimed to investigate the protective effect of PHI on NAFLD and elucidate the underlying mechanism. METHODS: The effects of PHI were examined in palmitate (PA)-stimulated AML12 cells and primary hepatocytes, as well as in NAFLD mice induced by a high-fat diet (HFD). We also used transcription factor EB (TFEB) knockdown hepatocytes and hepatocyte-specific TFEB knockout (TFEBΔhep) mice for mechanistic studies. In vivo and in vitro studies were performed using western blots, immunofluorescence techniques, and transmission electron microscopy. RESULTS: Our results indicated that autophagic flux and lysosome biogenesis in PA-stimulated hepatocytes were impaired. PHI alleviated lipid deposition by increasing lysosomal biogenesis and autophagic flux. It also stimulated the release of endoplasmic reticulum Ca2+ to activate calcineurin, which regulated TFEB dephosphorylation and nuclear translocation, and promoted lysosomal biogenesis. In addition, PHI blocked the NLRP3 inflammasome pathway and improved hepatocyte inflammation in an autophagy-dependent manner. Consistent with the in vitro results, PHI improved hepatic steatosis and inflammation in HFD mice, but these beneficial effects were eliminated in hepatocyte-specific TFEB knockout mice. CONCLUSION: Despite PHI has been reported to have anti-hepatic fibrosis effects, whether it has a hepatoprotective effects against NAFLD and the underlying molecular mechanism remain unclear. Herein, we found that PHI restored lipophagy and suppressed lipid accumulation and inflammation by regulating the Ca2+-calcineurin-TFEB axis in hepatocytes. Thus, PHI represents a therapeutic candidate for the treatment of NAFLD.

Cardioprotection effect of Yiqi-Huoxue-Jiangzhuo formula in a chronic kidney disease mouse model associated with gut microbiota modulation and NLRP3 inflammasome inhibition.

BACKGROUND: The pathogenesis and treatment of cardiovascular disease mediated by chronic kidney disease (CKD) are key research questions. Specifically, the mechanisms underlying the cardiorenal protective effect of Yiqi-Huoxue-Jiangzhuo formula (YHJF), a traditional Chinese herbal medicine, have not yet been clarified. METHODS: A classical CKD mouse model was constructed by 5/6 nephrectomy (Nx) to study the effects of YHJF intervention on 5/6 Nx mice cardiorenal function, gut microbial composition, gut-derived metabolites, and NLRP3 inflammasome pathways. RESULTS: YHJF improved cardiac dysfunction and reversed left ventricular hypertrophy, myocardial hypertrophy, and interstitial fibrosis in 5/6 Nx mice. In addition, YHJF inhibited activation of the NLRP3 inflammasome and downregulated the expression of TNF-α and IL-1ß both in the heart and serum; reconstitution of the intestinal flora imbalance was also found in 5/6 Nx mice treated with YHJF. Spearman's correlation and redundancy analyses showed that changes in the intestinal flora of 5/6 Nx mice were related to clinical phenotype and serum inflammatory levels. CONCLUSIONS: Treatment with YHJF effectively protected the heart function of 5/6 Nx mice; this effect was attributed to inhibition of NLRP3 inflammasome activation and regulation of intestinal microbial composition and derived metabolites. YHJF has potential for improving intestinal flora imbalance and gut-derived toxin accumulation in patients with CKD, thereby preventing cardiovascular complications.

Qing-Re-Xiao-Zheng Formula Modulates Gut Microbiota and Inhibits Inflammation in Mice With Diabetic Kidney Disease.

Evidence indicates that the metabolic inflammation induced by gut microbiota dysbiosis contributes to diabetic kidney disease. Prebiotic supplementations to prevent gut microbiota dysbiosis, inhibit inflammatory responses, and protect the renal function in DKD. Qing-Re-Xiao-Zheng formula (QRXZF) is a Traditional Chinese Medicine (TCM) formula that has been used for DKD treatment in China. Recently, there are growing studies show that regulation of gut microbiota is a potential therapeutic strategy for DKD as it is able to reduce metabolic inflammation associated with DKD. However, it is unknown whether QRXZF is effective for DKD by regulating of gut microbiota. In this study, we investigated the reno-protective effect of QRXZF by exploring its potential mechanism between gut microbiota and downstream inflammatory pathways mediated by gut-derived lipopolysaccharide (LPS) in the kidney. High-fat diet (HFD) and streptozotocin injection-induced DKD mice model was established to assess the QRXZF effect in vivo. Mice treated with QRXZF for 8 weeks had significantly lower levels of urinary albumin, serum cholesterol and triglycerides. The renal injuries observed through histological analysis were attenuated as well. Also, mice in the QRXZF group had higher levels of Zonula occludens protein-1 (ZO-1) expression, lower levels of serum fluorescein-isothiocyanate (FITC)-dextran and less-damaged colonic mucosa as compared to the DKD group, implying the benefit role for the gut barrier integrity. QRXZF treatment also reversed gut dysbiosis and reduced levels of gut-derived LPS. Notably, the expression of toll-like receptor 4 (TLR4) and nuclear factor-κB (NF-κB), which are important inflammation pathways in DKD, were suppressed in the QRXZF groups. In conclusion, our results indicated that the reno-protective effects of QRXZF was probably associated with modulating gut microbiota and inhibiting inflammatory responses in the kidney.

A novel strategy based on targeted cellular metabolomics for quantitatively evaluating anti-aging effect and screening effective extracts of Erzhi Wan.

The complexity of ingredients in traditional Chinese medicine (TCM) makes it challenging to clarify its efficacy in an acceptable and scientific approach. The present study was aimed to use quantification results from targeted cellular metabolomics to evaluate anti-aging efficacy of a famous Chinese medicine formula, Erzhi Wan (EZW), and screen possible effective extracts, depending on the developed strategy integrating multivariate receiver operating characteristic (ROC) curve and analytic hierarchy process (AHP). In this study, senescent NRK cells induced by D-galactose were treated with drug-containing serum of EZW and four kinds of extracts (petroleum ether, ethyl acetate, butanol and water). Intermediates of two major metabolic pathways for energy synthesis, tricarboxylic acid (TCA) cycle and glycolysis, were accurately quantified by GC-MS/MS to identify discriminate metabolites for clarifying therapeutic mechanism of EZW based on multivariate statistical analysis. Senescent and non-senescent cells were successfully distinguished using these metabolites by ROC curve analysis. Next, these metabolites were used as evaluation indexes to quantitatively reflect different effect of EZW and its extracts, according to the role of them in distinguishing groups and in conjunction with AHP. In vitro detection of senescence-associated ß-galactosidase (SA-ß-gal) activity was used to verify the reliability of evaluation results. The reversal after treatment of drug-containing serum of EZW and extracts was observed, and the petroleum ether extract might be the potential active extract responsible for the major anti-aging effect of EZW, which was in agreement with in vitro experiments. Altogether, metabolomics was a powerful approach for evaluation efficacy and elucidation action mechanisms of TCM. The integrated evaluation strategy in this paper with properties of high practicality, feasibility and effectivity was expected to provide a new insight into comprehensive and quantitative efficacy evaluation.

GC-MS based metabolomic profiling of lung tissue couple with network pharmacology revealed the possible protection mechanism of Pudilan Xiaoyan Oral Liquid in LPS-induced lung injury of mice.

Pudilan Xiaoyan Oral Liquid (PDL) originated from "Pudilan" Classic Recipe of traditional Chinese medicine is one kind of anti-inflammatory Chinese patent medicine recorded in Chinese Pharmacopeia. PDL has been used clinically for treating inflammatory diseases of the respiratory tract. However, due to the complex composition of PDL, its potential anti-inflammation and the mechanism remain unknown. To identify the mechanism of the PDL in the treatment of lipopolysaccharide (LPS)-induced lung injury of mice. The mice models of lung injury were established and the changes of biochemical indices in serum and histopathology were detected to explore the effects of PDL. The approach of GC-MS metabolomics was used to find more significant metabolites, and the metabolic pathways were enriched through MetaboAnalyst. Then network analysis was applied to visualize the protein related to the important metabolites, merging into a protein-metabolite network via Cytoscape. The treatment of PDL could attenuate LPS-induced histopathological damage of lung tissues, followed by reducing pro-inflammation mediators including IL-10, TNF-a and NF-ĸB in serum. 11 potential metabolites were identified in lung tissue through metabolomics, which were significantly regulated to recover by PDL treatment. The correlated network was constructed by integrating potential metabolites and pathways. Aspartate and l-cysteine were selected as key metabolites and correlated proteins such as IL4I1 and ASPA were speculated as the potential target to treat LPS-induced lung injury using PDL. These results demonstrated that PDL might prevent the pathological process of lung injury through regulating the disturbed protein-metabolite network.

A review on traditional uses, phytochemistry and pharmacology of Eclipta prostrata (L.) L.

ETHNOPHARMACOLOGICAL RELEVANCE: Eclipta prostrata, a traditional herbal medicine, has long been used in Asia and South America for the therapy of hemorrhagic diseases (e.g. hemoptysis, hematemesis, hematuria, epistaxis and uterine bleeding), skin diseases, respiratory disorders, coronary heart disease, hair loss, vitiligo, snake bite and those caused by the deficiency of liver and kidney. AIM OF THE REVIEW: In this review, we highlight relatively comprehensive and up-to-date information of E. prostrata on traditional uses, phytochemistry, pharmacology and toxicity, along with featuring the gaps in current knowledge, aiming to provide references for future research and possible opportunities for well applications of this medicinal plant. MATERIALS AND METHODS: Information on E. prostrata was gathered from scientific databases (Google Scholar, Web of Science, Scifinder, Baidu Scholar, PubMed and CNKI). Information was also obtained from local books, Ph.D. theses and M.Sc. dissertations and Chinese Pharmacopoeia. The plant taxonomy was validated by the database "The Plant List". RESULTS: Various phytochemical classes has been identified and isolated from the plant covering triterpenes, flavonoids, thiopenes, coumestans, steroids and others. Among these, coumestans are reported as the most common ingredients. The isolated crude extracts and individual compounds have been reported to exhibit promising pharmacological properties, such as hepatoprotective, osteoprotective, cytotoxic, hypoglycaemic, anti-inflammatory, anti-microbial, hypolipidemic, promoting hair growth, rejuvenative and neuroprotective effects. CONCLUSIONS: Until now, significant progress has been witnessed in phytochemistry and pharmacology of E. prostrata. Thus, some traditional uses has been well supported and clarified by modern pharmacological studies. Moreover, E. prostrata also showed therapeutic potential in some refractory diseases such as cancer, dementia and diabetes. But, present findings are still insufficient that cannot satisfactorily explain some mechanisms of action. More well-designed studies in vitro especially in vivo are required to establish links between the traditional uses and bioactivities, discover new skeletons and activity molecules, as well as ensure safety before clinical use.

Broad range metabolomics coupled with network analysis for explaining possible mechanisms of Er-Zhi-Wan in treating liver-kidney Yin deficiency syndrome of Traditional Chinese medicine.

ETHNOPHARMACOLOGICAL RELEVANCE: Er-Zhi-Wan (EZW), a famous traditional Chinese formulation, is used to prevent, or to treat, various liver and kidney diseases for its actions of replenishing liver and kidney. However, the mechanisms of treating Liver-kidney Yin deficiency syndrome (LKYDS) of EZW have not been comprehensively investigated. AIM OF THE STUDY: In this study, a broad range metabolomics strategy coupled with network analysis was established to investigate possible mechanisms of EZW in treating LKYDS. MATERIALS AND METHOD: The rat models of LKYDS were established using the mixture of thyroxine and reserpine, and the changes of biochemical indices in serum and histopathology were detected to explore the effects of EZW. Next, a broad range metabolomics strategy based on RPLC-Q-TOF/MS and HILIC-Q-TOF/MS has been developed to find the possible significant metabolites in the serum and urine of LKYDS rats. Then, network analysis was applied to visualize the relationships between identified serum and urine metabolites and in detail to find hub metabolites, which might be responsible for the effect of EZW on rats of LKYDS. Furthermore, the shortest path of "disease gene-pathway protein-metabolite" was built to investigate the possible intervention path of EZW from the systematic perspective. RESULTS: Five hub metabolites, namely, arachidonic acid, L-arginine, testosterone, taurine and oxoglutaric acid, were screened out and could be adjusted to recover by EZW. After that, the shortest path starting from disease genes and ending in metabolites were identified and disclosed, and the genes of aging such as CAV1 and ACO1 were selected to explain the pathological mechanism of LKYDS. CONCLUSION: Broad range metabolomics coupled with network analysis could provide another perspective on systematically investigating the molecular mechanism of EZW in treating LKYDS at metabolomics level. In addition, EZW might prevent the pathological process of LKYDS through regulating the disturbed metabolic pathway and the aging genes such as CAV1 and ACO1, which may be potential targets for EZW in the treatment of LKYDS.