Effects of electroacupuncture of different intensities and durations on PERK/ATF4/CHOP signaling pathway in liver of non-alcoholic fatty liver disease rats. / ä¸åŒå¼ºåº¦åŠæ—¶ç¨‹ç”µé’ˆå¹²é¢„å¯¹éžé ’ç²¾æ€§è„‚è‚ªè‚ç— å¤§é¼ è‚è„PERK/ATF4/CHOP通路的影响.

OBJECTIVES: To analyze the effects of electroacupuncture (EA) at "Fenglong" (ST40) and "Zusanli" (ST36) of different intensities and durations on rats with non-alcoholic fatty liver disease (NAFLD) based on the protein kinase R-like endoplasmic reticulum kinase (PERK)-activating transcription factor 4 (ATF4)-C/EBP homologous protein (CHOP) signaling pathway, so as to explore its mechanism underlying improvement of NAFLD. METHODS: SD rats were randomly divided into normal diet group, high-fat model group, sham EA group, strong stimulation EA (SEA) group, and weak stimulation EA (WEA) group, with 15 rats in each group. Each group was further divided into 2, 3, and 4-week subgroups. NAFLD rat model was established by feeding a high-fat diet. After successful modeling, rats in the SEA and WEA groups received EA at bilateral ST40 and ST36 with dense and sparse waves (4 Hz/20 Hz) at current intensities of 4 mA (SEA group) and 2 mA (WEA group), lasting for 20 minutes, once a day, 5 days a week with 2 days of rest. The sham EA group only had the EA apparatus connected without electricity. Different duration subgroups were intervened for 2, 3, and 4 weeks. After the intervention, the contents of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in rats were detected by an automatic biochemical analyzer；liver morphological changes were observed by Oil Red O staining；real-time fluorescence quantitative PCR and Western blot were used to detect the expression of PERK, ATF4, and CHOP mRNAs and proteins in the rat liver tissue. RESULTS: In the high-fat model group, there was a significant accumulation of red lipid droplets in the liver cells, which was reduced significantly in the SEA group at the 4th week. Compared with the normal diet group with the same treatment duration, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs and proteins in the liver tissue were elevated (P<0.01) in the high-fat model group . Compared with the high-fat model group with the same treatment duration, the contents of serum ALT, AST, and the expression of PERK, ATF4, CHOP mRNAs and proteins in the liver tissue were decreased (P<0.01, P<0.05) in the SEA and WEA groups. Compared with the sham EA group with the same treatment duration, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs were decreased (P<0.01, P<0.05) in the SEA and WEA groups, the expression of PERK, ATF4, and CHOP proteins in the liver tissue was decreased (P<0.01) in the SEA group at the 2nd, 3rd, and 4th week, the expression of PERK and CHOP proteins at the 2nd, 3rd, 4th week and ATF4 protein at 2nd week in the liver tissue were decreased (P<0.01, P<0.05) in the WEA group. Compared with the SEA group with the same treatment duration, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs and proteins in the liver tissue were elevated (P<0.05, P<0.01) in the WEA group. Compared with the 2-week time point within the groups, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs and PERK proteins in the liver tissue were decreased (P<0.01, P<0.05) in the SEA and WEA groups at 3rd and 4th week, the expression of ATF4 proteins in the liver tissue was decreased (P<0.01) in the SEA group at 3rd and 4th week, and the expression of CHOP proteins in the liver tissue was decreased (P<0.01) in the SEA group at 4th week and in the WEA group at 3rd and 4th week. Compared with the 3-week time point within the groups, the contents of serum ALT, AST, and the expression of PERK, ATF4, and CHOP mRNAs were significantly decreased (P<0.05, P<0.01) in the SEA and WEA groups at 4th week, the expression of PERK and CHOP proteins in the liver tissue was decreased (P<0.01) in the SEA and WEA groups at 4th week, and the expression of ATF4 protein in the liver tissue was decreased (P<0.05) in the SEA group at 4th week. CONCLUSIONS: EA at ST40 and ST36 can significantly improve liver function in NAFLD rats, and its mechanism of action may involve inhibiting PERK expression thereby targeting the downstream ATF4/CHOP signaling pathway to suppress endoplasmic reticulum stress, exerting a liver protective effect；the optimal effect was observed with EA intensity of 4 mA for 4 weeks.

[Effect of Erchen Decoction on liver mitochondrial function by inhibiting mTORC1/SREBP1/CAV1 pathway in mice with high-fat diet].

This study aims to investigate the effect of Erchen Decoction(ECD) on liver mitochondrial function in mice with a high-fat diet and its possible mechanism. A total of sixty C57BL/6J mice were randomly divided into a normal group, high-fat group, ECD group, mTORC1 activator(MHY) group, ECD+MHY group, and polyene phosphatidyl choline(PPC) group, with 10 rats in each group. The normal group was given a normal diet, and the other groups were fed a high-fat diet for 20 weeks. At the 17th week, the ECD group and ECD+MHY group were given ECD(8.7 g·kg~(-1)) daily, and the PPC group was given PPC(0.18 g·kg~(-1)) daily, while the remaining groups were given normal saline(0.01 mL·g~(-1)) daily for four weeks. In the 19th week, the MHY group and ECD+MHY group were injected intraperitoneally with MHY(5 mg·kg~(-1)) every other day for two weeks. During the experiment, the general conditions of the mice were observed. The contents of triglyceride(TG) and total cholesterol(TC) in serum were measured. Morphological changes in liver tissue were examined through HE and oil red O staining. The content of adenosine triphosphate(ATP) was determined using chemiluminescence, and mitochondrial membrane potential was assessed using a fluorescence probe(JC-1). Western blot was performed to detect the expression of rapamycin target protein complex 1(mTOR1), ribosomal protein S6 kinase B1(S6K), sterol regulatory element binding protein 1(SREBP1), and caveolin 1(CAV1). RESULTS:: revealed that compared with the normal group, the mice in the high-fat group exhibited significant increases in body weight and abdominal circumference(P<0.01). Additionally, there were significant increases in TG and TC levels(P<0.01). HE and oil red O staining showed that the boundaries of hepatic lobules were unclear; hepatocytes were enlarged, round, and irregularly arranged, with obvious lipid droplet deposition and inflammatory cell infiltration. The liver ATP content and mitochondrial membrane potential decreased significantly(P<0.01). The expression of p-mTOR, p-S6K, and n-SREBP1 increased significantly(P<0.01), while the expression of CAV1 decreased significantly(P<0.01). Compared with the high-fat group, the body weight and TG content of mice in the ECD group and PPC group decreased significantly(P<0.05). Improvements were observed in hepatocyte morphology, lipid deposition, and inflammatory cell infiltration. Furthermore, there were significant increases in ATP content and mitochondrial membrane potential(P<0.05 or P<0.01). The expression of p-mTOR, p-S6K, and n-SREBP1 decreased significantly in the ECD group(P<0.01), while CAV1 expression increased significantly(P<0.01). However, the indices mentioned above did not show improvement in the MHY group. When the ECD+MHY group was compared with the MHY group, there were significant reductions in body weight and TG contents(P<0.05). The morphological changes of hepatocytes, lipid deposition, and inflammatory cell infiltration were recovered. Moreover, there were significant increases in liver ATP content and mitochondrial membrane potential(P<0.05 or P<0.05). The expression of p-mTOR, p-S6K, and n-SREBP1 decreased significantly(P<0.01), while CAV1 expression increased significantly(P<0.01). In conclusion, ECD can improve mitochondrial function by regulating the mTORC1/SREBP1/CAV1 pathway. This mechanism may be involved in the resolution of phlegm syndrome and the regulation of lipid metabolism.

Alpha lipoic acid administration improved both peripheral sensitivity to insulin and liver clearance of insulin reducing potential risk of diabetes and nonalcoholic fatty liver disease in overweight/obese PCOS patients.

OBJECTIVE: To evaluate the effects of alpha lipoic acid (ALA) on hormonal and metabolic parameters in a group of overweight/obese Polycystic Ovary Syndrome (PCOS) patients. METHODS: This was a retrospective study in which thirty-two overweight/obese patients with PCOS (n = 32) not requiring hormonal treatment were selected from the database of the ambulatory clinic of the Gynecological Endocrinology Center at the University of Modena and Reggio Emilia, Italy. The hormonal profile, routine exams and insulin and C-peptide response to oral glucose tolerance test (OGTT) were evaluated before and after 12 weeks of complementary treatment with ALA (400 mg/day). Hepatic Insulin Extraction (HIE) index was also calculated. RESULTS: ALA administration significantly improved insulin sensitivity and decreased ALT and AST plasma levels in all subjects, though no changes were observed on reproductive hormones. When PCOS patients were subdivided according to the presence or absence of familial diabetes background, the higher effects of ALA were observed in the former group that showed AST and ALT reduction and greater HIE index decrease. CONCLUSION: ALA administration improved insulin sensitivity in overweight/obese PCOS patients, especially in those with familial predisposition to diabetes. ALA administration improved both peripheral sensitivity to insulin and liver clearance of insulin. Such effects potentially decrease the risk of nonalcoholic fat liver disease and diabetes in PCOS patients.

[Effect of Zuogui Jiangtang Qinggan Formula on lipid metabolism in db/db mouse model of type 2 diabetes mellitus complicated with non-alcoholic fatty liver disease].

This study aims to explore the effect of Zuogui Jiangtang Qinggan Formula(ZGJTQGF) on the lipid metabolism in the db/db mouse model of type 2 diabetes mellitus(T2DM) complicated with non-alcoholic fatty liver disease(NAFLD) via the insulin receptor(INSR)/adenosine 5'-monophosphate(AMP)-activated protein kinase(AMPK)/sterol-regulatory element-binding protein 2(SREBP-2) signaling pathway. Twenty-four db/db mice were randomized into positive drug(metformin, 0.067 g·kg~(-1)) and low-(7.5 g·kg~(-1)) and high-dose(15 g·kg~(-1)) ZGJTQGF groups. Six C57 mice were used as the blank group and administrated with an equal volume of distilled water. The mice in other groups except the blank group were administrated with corresponding drugs by gavage for 6 consecutive weeks. At the end of drug administration, fasting blood glucose(FBG) and blood lipid levels were measured, and oral glucose tolerance test was performed. Compared with the blank group, the mice treated with ZGJTQGF showed decreased body mass and liver weight coefficient, lowered levels of FBG, total cholesterol(TC), triglyceride(TG), and low-density lipoprotein(LDL), and weakened liver function. The pathological changes and lipid accumulation in the liver tissue were examined. Western blot was employed to measure the protein levels of INSR, AMPK, p-AMPK, and SREBP-2. Compared with the blank group, the model group showed down-regulated protein levels of INSR and p-AMPK/AMPK and up-regulated protein level of SREBP-2. Compared with the model group, high-dose ZGJTQGF up-regulated the protein levels of INSR and p-AMPK/AMPK and down-regulated the protein level of SREBP-2. Low-dose ZGJTQGF slightly up-regulated the protein levels of INSR and p-AMPK/AMPK and down-regulated the protein level of SREBP-2, without significant differences. The results suggested that ZGJTQGF may alleviate insulin resistance and improve lipid metabolism in db/db mice by activating the INSR/AMPK/SREBP-2 signaling pathway.

[Tanshinone â ¡_A inhibits ferroptosis via Nrf2 signaling pathway to protect liver in rats of non-alcoholic fatty liver disease].

This study investigated the protective effect of tanshinone Ⅱ_A(TSⅡ_A) on the liver in the rat model of non-alcoholic fatty liver disease(NAFLD) and the mechanism of TSⅡ_A in regulating ferroptosis via the nuclear factor E2-related factor 2(Nrf2) signaling pathway. The rat model of NAFLD was established with a high-fat diet for 12 weeks. The successfully modeled rats were assigned into model group, low-and high-dose TSⅡ_A groups, and inhibitor group, and normal control group was set. Enzyme-linked immunosorbent assay was employed to determine the content of superoxide dismutase(SOD) and malondialdehyde(MDA) in the serum of rats in each group. A biochemical analyzer was used to measure the content of aspartate aminotransferase(AST), alaninl aminotransferase(ALT), total cholesterol(TC), and triglycerides(TG). Hematoxylin-eosin(HE) staining was used to detect pathological damage in liver tissue. Terminal-deoxynucleoitidyl transferase-mediated nick end labeling(TUNEL) was employed to examine the apoptosis of the liver tissue. Oil red O staining, MitoSOX staining, and Prussian blue staining were conducted to reveal lipid deposition, the content of reactive oxygen species(ROS), and iron deposition in liver tissue. Western blot was employed to determine the expression of Nrf2, heme oxygenase-1(HO-1), glutathione peroxidase 4(GPX4), ferroptosis suppressor protein 1(FSP1), B cell lymphoma-2(Bcl-2), and Bcl-2 associated X protein(Bax) in the liver tissue. The result showed that TSⅡ_A significantly reduced the content of MDA, AST, ALT, TC, and TG in the serum, increased the activity of SOD, decreased the apoptosis rate, lipid deposition, ROS, and iron deposition in the liver tissue, up-regulated the expression of Nrf2, HO-1, FSP1, GPX, and Bcl-2, and inhibited the expression of Bax in the liver tissue of NAFLD rats. However, ML385 partially reversed the protective effect of TSⅡ_A on the liver tissue. In conclusion, TSⅡ_A could inhibit ferroptosis in the hepatocytes and decrease the ROS and lipid accumulation in the liver tissue of NAFLD rats by activating the Nrf2 signaling pathway.

The lack of PPARα exacerbated the progression of non-alcoholic steatohepatitis in mice with spleen deficiency syndrome by triggering an inflammatory response.

Background: In addition to abnormal liver inflammation, the main symptoms of non-alcoholic steatohepatitis (NASH) are often accompanied by gastrointestinal digestive dysfunction, consistent with the concept of spleen deficiency (SD) in traditional Chinese medicine. As an important metabolic sensor, whether peroxisome proliferator-activated receptor alpha (PPARα) participates in regulating the occurrence and development of NASH with SD (NASH-SD) remains to be explored. Methods: Clinical liver samples were collected for RNA-seq analysis. C57BL/6J mice induced by folium sennae (SE) were used as an SD model. qPCR analysis was conducted to evaluate the inflammation and metabolic levels of mice. PPARα knockout mice (PPARαko) were subjected to SE and methionine-choline-deficient (MCD) diet to establish the NASH-SD model. The phenotype of NASH and the inflammatory indicators were measured using histopathologic analysis and qPCR as well. Results: The abnormal expression of PPARα signaling, coupled with metabolism and inflammation, was found in the results of RNA-seq analysis from clinical samples. SD mice showed a more severe inflammatory response in the liver evidenced by the increases in macrophage biomarkers, inflammatory factors, and fibrotic indicators in the liver. qPCR results also showed differences in PPARα between SD mice and control mice. In PPARαko mice, further evidence was found that the lack of PPARα exacerbated the inflammatory response phenotype as well as the lipid metabolism disorder in NASH-SD mice. Conclusion: The abnormal NR signaling accelerated the vicious cycle between lipotoxicity and inflammatory response in NAFLD with SD. Our results provide new evidence for nuclear receptors as potential therapeutic targets for NAFLD with spleen deficiency.

Lian-Mei-Yin formula alleviates diet-induced hepatic steatosis by suppressing Yap1/FOXM1 pathway-dependent lipid synthesis.

Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease, with a global prevalence of 25%. Patients with NAFLD are more likely to suffer from advanced liver disease, cardiovascular disease, or type II diabetes. However, unfortunately, there is still a shortage of FDA-approved therapeutic agents for NAFLD. Lian-Mei-Yin (LMY) is a traditional Chinese medicine formula used for decades to treat liver disorders. It has recently been applied to type II diabetes which is closely related to insulin resistance. Given that NAFLD is another disease involved in insulin resistance, we hypothesize that LMY might be a promising formula for NAFLD therapy. Herein, we verify that the LMY formula effectively reduces hepatic steatosis in diet-induced zebrafish and NAFLD model mice in a time- and dose-dependent manner. Mechanistically, LMY suppresses Yap1-mediated Foxm1 activation, which is crucial for the occurrence and development of NAFLD. Consequently, lipogenesis is ameliorated by LMY administration. In summary, the LMY formula alleviates diet-induced NAFLD in zebrafish and mice by inhibiting Yap1/Foxm1 signaling-mediated NAFLD pathology.

Natural Products in the Modulation of Farnesoid X Receptor Against Nonalcoholic Fatty Liver Disease.

Nonalcoholic fatty liver disease (NAFLD) is a global health concern with a high prevalence and increasing economic burden, but official medicine remains unavailable. Farnesoid X receptor (FXR), a nuclear receptor member, is one of the most promising drug targets for NAFLD therapy that plays a crucial role in modulating bile acid, glucose, and lipid homeostasis, as well as inhibits hepatic inflammation and fibrosis. However, the rejection of the FXR agonist, obecholic acid, by the Food and Drug Administration for treating hepatic fibrosis raises a question about the functions of FXR in NAFLD progression and the therapeutic strategy to be used. Natural products, such as FXR modulators, have become the focus of attention for NAFLD therapy with fewer adverse reactions. The anti-NAFLD mechanisms seem to act as FXR agonists and antagonists or are involved in the FXR signaling pathway activation, indicating a promising target of FXR therapeutic prospects using natural products. This review discusses the effective mechanisms of FXR in NAFLD alleviation, and summarizes currently available natural products such as silymarin, glycyrrhizin, cycloastragenol, berberine, and gypenosides, for targeting FXR, which can facilitate development of naturally targeted drug by medicinal specialists for effective treatment of NAFLD.

SIRT1: Harnessing multiple pathways to hinder NAFLD.

Non-alcoholic fatty liver disease (NAFLD) encompasses hepatic steatosis, non-alcoholic steatohepatitis (NASH), fibrosis, cirrhosis, and hepatocellular carcinoma. It is the primary cause of chronic liver disorders, with a high prevalence but no approved treatment. Therefore, it is indispensable to find a trustworthy therapy for NAFLD. Recently, mounting evidence illustrates that Sirtuin 1 (SIRT1) is strongly associated with NAFLD. SIRT1 activation or overexpression attenuate NAFLD, while SIRT1 deficiency aggravates NAFLD. Besides, an array of therapeutic agents, including natural compounds, synthetic compounds, traditional Chinese medicine formula, and stem cell transplantation, alleviates NALFD via SIRT1 activation or upregulation. Mechanically, SIRT1 alleviates NAFLD by reestablishing autophagy, enhancing mitochondrial function, suppressing oxidative stress, and coordinating lipid metabolism, as well as reducing hepatocyte apoptosis and inflammation. In this review, we introduced the structure and function of SIRT1 briefly, and summarized the effect of SIRT1 on NAFLD and its mechanism, along with the application of SIRT1 agonists in treating NAFLD.

Yanggan Jiangmei Formula alleviates hepatic inflammation and lipid accumulation in non-alcoholic steatohepatitis by inhibiting the NF-κB/NLRP3 signaling pathway.

The role of NF-κB and the NLRP3 inflammasome in the chronic inflammatory microenvironment of non-alcoholic steatohepatitis (NASH) has been posited as crucial. The Yanggan Jiangmei Formula (YGJMF) has shown promise in ameliorating hepatic steatosis in NASH patients, yet its pharmacological mechanisms remain largely unexplored. This study was conducted to investigate the efficacy of YGJMF in NASH and to elucidate its pharmacological underpinnings. To simulate NASH both in vivo and in vitro, high-fat-diet (HFD) rats and HepG2 cells stimulated with free fatty acids (FFAs) were utilized. The severity of liver injury and lipid deposition was assessed using serum indicators, histopathological staining, micro-magnetic resonance imaging (MRI), and the liver-to-muscle signal intensity ratio (SIRL/M). Furthermore, a combination of enzyme-linked immunosorbent assay (ELISA), immunohistochemistry (IHC), immunofluorescence, real-time quantitative polymerase chain reaction (RT-qPCR), and Western blotting analyses was employed to investigate the NF-κB/NLRP3 signaling pathway and associated cytokine levels. The results from liver pathology, MRI assessments, and biochemical tests in rat models demonstrated YGJMF's significant effectiveness in reducing liver damage and lipid accumulation. Additionally, YGJMF markedly reduced hepatocyte inflammation by downregulating inflammatory cytokines in both liver tissue and serum. Furthermore, YGJMF was found to disrupt NF-κB activation, consequently inhibiting the assembly of the NLRP3 inflammasome in both the in vitro and in vivo models. The preliminary findings of this study suggest that YGJMF may alleviate hepatic steatosis and inhibit the NF-κB/NLRP3 signaling pathway, thereby exerting anti-inflammatory effects in NASH.

GBA3 promotes fatty acid oxidation and alleviates non-alcoholic fatty liver by increasing CPT2 transcription.

BACKGROUND: Excessive lipids accumulation and hepatocytes death are prominent characteristics of non-alcoholic fatty liver disease (NAFLD). Nonetheless, the precise pathophysiological mechanisms are not fully elucidated. METHODS: HepG2 cells stimulated with palmitic acids and rats fed with high-fat diet were used as models for NAFLD. The impact of Glucosylceramidase Beta 3 (GBA3) on fatty acid oxidation (FAO) was assessed using Seahorse metabolic analyzer. Lipid content was measured both in vitro and in vivo. To evaluate NAFLD progression, histological analysis was performed along with measurements of inflammatory factors and liver enzyme levels. Western blot and immunohistochemistry were employed to examine the activity levels of necroptosis. Flow cytometry and reactive oxygen species (ROS) staining were utilized to assess levels of oxidative stress. RESULTS: GBA3 promoted FAO and enhanced the mitochondrial membrane potential without affecting glycolysis. These reduced the lipid accumulation. Rats supplemented with GBA3 exhibited lower levels of inflammatory factors and liver enzymes, resulting in a slower progression of NAFLD. GBA3 overexpression reduced ROS and the ratio of cell apoptosis. Phosphorylation level was reduced in the essential mediator, MLKL, implicated in necroptosis. Mechanistically, as a transcriptional coactivator, GBA3 promoted the expression of Carnitine Palmitoyltransferase 2 (CPT2), which resulted in enhanced FAO. CONCLUSIONS: Increased FAO resulting from GBA3 reduced oxidative stress and the production of ROS, thereby inhibiting necroptosis and delaying the progression of NAFLD. Our research offers novel insights into the potential therapeutic applications of GBA3 and FAO in the management and treatment of NAFLD.

Extract of Silphium perfoliatum L. improve lipid accumulation in NAFLD mice by regulating AMPK/FXR signaling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Globally, the incidence rate and number of patients with nonalcoholic fatty liver disease are increasing, which has become one of the greatest threats to human health. However, there is still no effective therapy and medicine so far. Silphium perfoliatum L. is a perennial herb native to North America, which is used to improve physical fitness and treat liver and spleen related diseases in the traditional medicinal herbs of Indian tribes. This herb is rich in chlorogenic acids, which have the functions of reducing blood lipids, losing weight and protecting liver. However, the effect of these compounds on nonalcoholic fatty liver disease remains unclear. AIM OF THE STUDY: Clarify the therapeutic effects and mechanism of the extract (CY-10) rich in chlorogenic acid and its analogues from Silphium perfoliatum L. on non-alcoholic fatty liver disease, and to determine the active compounds. MATERIALS AND METHODS: A free fatty acid-induced steatosis model of HepG2 cells was established to evaluate the in vitro activity of CY-10 in promoting lipid metabolism. Further, a high-fat diet-induced NAFLD model in C57BL/6 mice was established to detect the effects of CY-10 on various physiological and biochemical indexes in mice, and to elucidate the in vivo effects of the extract on regulating lipid metabolism, anti-inflammation and hepatoprotection, and nontarget lipid metabolomics was performed to analyze differential metabolites of fatty acids in the liver. Subsequently, western blotting and immunohistochemistry were used to analyze the target of the extract and elucidate its mechanism of action. Finally, the active compounds in CY-10 were elucidated through in vitro activity screening. RESULTS: The results indicated that CY-10 significantly attenuated lipid droplet deposition in HepG2 cells. The results of in vivo experiments showed that CY-10 significantly reduce HFD-induced mouse body weight and organ index, improve biochemical indexes, oxidation levels and inflammatory responses in the liver and serum, thereby protecting the liver tissue. It can promote the metabolism of unsaturated fatty acids in the liver and reduce the generation of saturated fatty acids. Furthermore, it is clarified that CY-10 can promote lipid metabolism balance by regulating AMPK/FXR/SREPB-1c/PPAR-γ signal pathway. Ultimately, the main active compound was proved to be cryptochlorogenic acid, which has a strong promoting effect on the metabolism of fatty acids in cells. Impressively, the activities of CY-10 and cryptochlorogenic acid were stronger than simvastatin in vitro and in vivo. CONCLUSION: For the first time, it is clarified that the extract rich in chlorogenic acids and its analogues in Silphium perfoliatum L. have good therapeutic effects on non-alcoholic fatty liver disease. It is confirmed that cryptochlorogenic acid is the main active compound and has good potential for medicine.

Si-Wu-Tang attenuates hepatocyte PANoptosis and M1 polarization of macrophages in non-alcoholic fatty liver disease by influencing the intercellular transfer of mtDNA.

ETHNOPHARMACOLOGICAL RELEVANCE: Non-alcoholic fatty liver disease (NAFLD) represents a burgeoning challenge for public health with potential progression to malignant liver diseases. PANoptosis, an avant-garde conceptualization of cell deaths, is closely associated with mitochondrial damage and linked to multiple liver disorders. Si-Wu-Tang (SWT), a traditional Chinese herbal prescription renowned for regulating blood-related disorders and ameliorating gynecological and hepatic diseases, has been demonstrated to alleviate liver fibrosis by regulating bile acid metabolism and immune responses. AIM OF THE STUDY: However, the mechanisms by which mtDNA is released from PANoptotic hepatocytes, triggering macrophage activation and hepatitis and whether this process can be reversed by SWT remain unclear. MATERIALS AND METHODS: Here, sophisticated RNA-sequencing complemented by molecular approaches were applied to explore the underlying mechanism of SWT against NAFLD in methionine/choline-deficient diet (MCD)-induced mice and relative in vitro models. RESULTS: We revealed that SWT profoundly repaired mitochondrial dysfunction, blocked mitochondrial permeability transition and mtDNA released to the cytoplasm, subsequently reversing hepatocyte PANoptosis and macrophage polarization both in MCD-stimulated mice and in vitro. Mechanically, loaded lipids dramatically promoted the opening of mPTP and oligomerization of VDAC2 to orchestrate mtDNA release, which was combined with ZBP1 to promote hepatocyte PANoptosis and also taken by macrophages to trigger M1 polarization via the FSTL1 and PKM2 combination. SWT effectively blocked NOXA signaling and reversed all these detrimental outcomes. CONCLUSION: Our findings show that SWT protects against hepatitis-mediated hepatocyte PANoptosis and macrophage M1 polarization by influencing intrahepatic synthesis, release and intercellular transfer of mtDNA, suggesting a potential therapeutic strategy for ameliorating NAFLD.

Docosahexaenoic Acid Modulates Nonalcoholic Fatty Liver Disease by Suppressing Endocannabinoid System.

SCOPE: Endocannabinoid signaling regulates energy homeostasis, and is tightly associated with nonalcoholic fatty liver disease (NAFLD). The study previously finds that supplementation of docosahexaenoic acid (DHA) has superior function to ameliorate NAFLD compared with eicosapentaenoic acid (EPA), however, the underlying mechanism remains elusive. The present study aims to investigate whether DHA intervention alleviates NAFLD via endocannabinoid system. METHODS AND RESULTS: In a case-control study, the serum endocannabinoid ligands in 60 NAFLD and 60 healthy subjects are measured. Meanwhile, NAFLD model is established in mice fed a high-fat and -cholesterol diet (HFD) for 9 weeks. DHA or EPA is administrated for additional 9 weeks. Serum primary endocannabinoid ligands, namely anandamide (AEA) and 2-arachidoniylglycerol (2-AG), are significantly higher in individuals with NAFLD compared with healthy controls. NAFLD model shows that serum 2-AG concentrations and adipocyte cannabinoid receptor 1 expression levels are significantly lower in DHA group compared with HFD group. Lipidomic and targeted ceramide analyses further confirm that endocannabinoid signaling inhibition has exerted deletion of hepatic C16:0-ceramide contents, resulting in down-regulation of de novo fatty acid synthesis and up-regulation of fatty acid ß-oxidation related protein expression levels. CONCLUSIONS: This work elucidates that DHA has improved NAFLD by suppressing endocannabinoid system.

Smilax China L. polysaccharide prevents HFD induced-NAFLD by regulating hepatic fat metabolism and gut microbiota.

BACKGROUND: The increasing incidence of nonalcoholic fatty liver disease (NAFLD) has urged the development of new therapeutics. NAFLD is intimately linked to gut microbiota due to the hepatic portal system, and utilizing natural polysaccharides as prebiotics has become a prospective strategy for preventing NAFLD. Smilax china L. polysaccharide (SCP) possesses excellent hepatoprotective and anti-inflammatory activity. However, its protective effects on NAFLD remains unclear. PURPOSE: The goal of this study was to explore the protective effects of SCP on high-fat diet (HFD)-induced NAFLD mice by regulating hepatic fat metabolism and gut microbiota. METHODS: Extraction and isolation from Smilax china L. rhizome to obtain SCP. C57BL/6 J mice were distributed to six groups: Control (normal chow diet), HFD-fed mice were assigned to HFD, simvastatin (SVT), and low-, medium-, high-doses of SCP for 12 weeks. The body, liver, and different adipose tissues weights were detected, and lipids in serum and liver were assessed. RT-PCR and Western blot were used to detect the hepatic fat metabolism-related genes and proteins. Gut microbiota of cecum contents was profiled through 16S rRNA gene sequencing. RESULTS: SCP effectively reversed HFD-induced increase weights of body, liver, and different adipose tissues. Lipid levels of serum and liver were also significantly reduced after SCP intervention. According to the results of RT-PCR and western blot analysis, SCP treatment up-regulated the genes and proteins related to lipolysis were up-regulated, while lipogenesis-related genes and proteins were down-regulated. Furthermore, the HFD-induced dysbiosis of intestinal microbiota was similarly repaired by SCP intervention, including enriching beneficial bacteria and depleting harmful bacteria. CONCLUSION: SCP could effectively prevent HFD-induced NAFLD, might be considered as a prebiotic agent due to its excellent effects on altering hepatic fat metabolism and maintaining gut microbiota homeostasis.

Hedan tablet ameliorated non-alcoholic steatohepatitis by moderating NF-κB and lipid metabolism-related pathways via regulating hepatic metabolites.

Non-alcoholic steatohepatitis (NASH) is a severe form of fatty liver disease. If not treated, it can lead to liver damage, cirrhosis and even liver cancer. However, advances in treatment have remained relatively slow, and there is thus an urgent need to develop appropriate treatments. Hedan tablet (HDP) is used to treat metabolic syndrome. However, scientific understanding of the therapeutic effect of HDP on NASH remains limited. We used HDP to treat a methionine/choline-deficient diet-induced model of NASH in rats to elucidate the therapeutic effects of HDP on liver injury. In addition, we used untargeted metabolomics to investigate the effects of HDP on metabolites in liver of NASH rats, and further validated its effects on inflammation and lipid metabolism following screening for potential target pathways. HDP had considerable therapeutic, anti-oxidant, and anti-inflammatory effects on NASH. HDP could also alter the hepatic metabolites changed by NASH. Moreover, HDP considerable moderated NF-κB and lipid metabolism-related pathways. The present study found that HDP had remarkable therapeutic effects in NASH rats. The therapeutic efficacy of HDP in NASH mainly associated with regulation of NF-κB and lipid metabolism-related pathways via arachidonic acid metabolism, glycine-serine-threonine metabolism, as well as steroid hormone biosynthesis.

Vitamin D Mitigates Hepatic Fat Accumulation and Inflammation and Increases SIRT1/AMPK Expression in AML-12 Hepatocytes.

Emerging evidence has demonstrated a strong correlation between vitamin D status and fatty liver disease. Aberrant hepatic fat infiltration contributes to oxidant overproduction, promoting metabolic dysfunction, and inflammatory responses. Vitamin D supplementation might be a good strategy for reducing hepatic lipid accumulation and inflammation in non-alcoholic fatty liver disease and its associated diseases. This study aimed to investigate the role of the most biologically active form of vitamin D, 1,25-dihydroxyvitamin D (1,25(OH)2D), in hepatic fat accumulation and inflammation in palmitic acid (PA)-treated AML-12 hepatocytes. The results indicated that treatment with 1,25(OH)2D significantly decreased triglyceride contents, lipid peroxidation, and cellular damage. In addition, mRNA levels of apoptosis-associated speck-like CARD-domain protein (ASC), thioredoxin-interacting protein (TXNIP), NOD-like receptor family pyrin domain-containing 3 (NLRP3), and interleukin-1ß (IL-1ß) involved in the NLRP3 inflammasome accompanied by caspase-1 activity and IL-1ß expression were significantly suppressed by 1,25(OH)2D in PA-treated hepatocytes. Moreover, upon PA exposure, 1,25(OH)2D-incubated AML-12 hepatocytes showed higher sirtulin 1 (SIRT1) expression and adenosine monophosphate-activated protein kinase (AMPK) phosphorylation. A SIRT1 inhibitor alleviated the beneficial effects of 1,25(OH)2D on PA-induced hepatic fat deposition, IL-1ß expression, and caspase-1 activity. These results suggest that the favorable effects of 1,25(OH)2D on hepatic fat accumulation and inflammation may be, at least in part, associated with the SIRT1.

Tangshen formula targets the gut microbiota to treat non-alcoholic fatty liver disease in HFD mice: A 16S rRNA and non-targeted metabolomics analyses.

BACKGROUND: Tangshen formula (TSF) has an ameliorative effect on hepatic lipid metabolism in non-alcoholic fatty liver disease (NAFLD), but the role played by the gut microbiota in this process is unknown. METHOD: We conducted three batches of experiments to explore the role played by the gut microbiota: TSF administration, antibiotic treatment, and fecal microbial transplantation. NAFLD mice were induced with a high-fat diet to investigate the ameliorative effects of TSF on NAFLD features and intestinal barrier function. 16S rRNA sequencing and serum untargeted metabolomics were performed to further investigate the modulatory effects of TSF on the gut microbiota and metabolic dysregulation in the body. RESULTS: TSF ameliorated insulin resistance, hypercholesterolemia, lipid metabolism disorders, inflammation, and impairment of intestinal barrier function. 16S rRNA sequencing analysis revealed that TSF regulated the composition of the gut microbiota and increased the abundance of beneficial bacteria. Antibiotic treatment and fecal microbiota transplantation confirmed the importance of the gut microbiota in the treatment of NAFLD with TSF. Subsequently, untargeted metabolomics identified 172 differential metabolites due to the treatment of TSF. Functional predictions suggest that metabolisms of choline, glycerophospholipid, linoleic acid, alpha-linolenic acid, and arachidonic acid are the key metabolic pathways by which TSF ameliorates NAFLD and this may be influenced by the gut microbiota. CONCLUSION: TSF treats the NAFLD phenotype by remodeling the gut microbiota and improving metabolic profile, suggesting that TSF is a functional gut microbial and metabolic modulator for the treatment of NAFLD.

Integrated network pharmacology, metabolomics, and transcriptomics of Huanglian-Hongqu herb pair in non-alcoholic fatty liver disease.

ETHNOPHARMACOLOGICAL RELEVANCE: The Huanglian-Hongqu herb pair (HH) is a synergistic drug combination used to treat non-alcoholic fatty liver disease (NAFLD). However, the molecular mechanism underlying the therapeuticeffects of HH requires further elucidation. AIM OF THE STUDY: The present study explored the potential mechanism of HH in treating NAFLD. MATERIALS AND METHODS: UPLC-Q-TOF-MS was employed to identify the drug constituents in HH. A NAFLD rat model was induced by a high-fat diet (HFD) and treated with different doses of HH. The functional mechanism of HH in NAFLD rats was predicted using network pharmacology, metabolomics and transcriptomics. Immunohistochemistry, real-time PCR, and Western blot were performed to validate the key mechanisms. RESULTS: Pharmacodynamic assessment demonstrated that HH exhibited improvements in lipid deposition and reduced hepatic oxidative stress in NAFLD rats. Hepatic wide-target metabolomics revealed that HH primarily modulated amino acids and their metabolites, fatty acids, organic acids and their derivatives, bile acids, and other liver metabolites. The enriched pathways included metabolic pathways, primary bile acid biosynthesis, and bile secretion. Network pharmacology analysis indicated that HH regulated the key pathways in NAFLD, notably PPAR, AMPK, NF-κB and other signaling pathways. Furthermore, hepatic transcriptomics, based on Illumina RNA-Seq sequencing analyses, suggested that HH improved NAFLD through metabolic pathways, the PPAR signaling pathway, primary bile acid biosynthesis, and fatty acid metabolism. Further mechanistic studies indicated that HH could regulate the genes and proteins associated with the PPAR signaling pathway. CONCLUSION: Our findings demonstrated that the potential therapeutic benefits of HH in ameliorating NAFLD by targeting the PPAR signaling pathway, thereby facilitating a more extensive use of HH in NAFLD.

Effects of Ecklonia stolonifera Extract on Metabolic Dysregulation in High-Fat Diet-Induced Obese Mice.

This study aimed to test the hypothesis that long-term and low-dose supplementation with an ethanol extract of Ecklonia stolonifera may confer protection against high-fat diet (HFD)-induced obesity in mice. Male C57BL/6J mice were divided into two groups, one of which was fed an HFD (40 kcal% fat) and the other an HFD+E. stolonifera (0.006%, w/w, ∼5 mg/kg body weight/day) for 16 weeks. E. stolonifera supplementation significantly reduced body weight from week 3 and until the end of the experiment. E. stolonifera-supplemented mice also exhibited lower fat mass (epididymal, perirenal, and mesenteric fat) and smaller adipocyte size than HFD control mice. The two groups displayed similar food intakes, but E. stolonifera markedly decreased lipogenesis and increased lipolysis and fatty acid oxidation in adipose tissue. Moreover, E. stolonifera significantly decreased plasma and hepatic lipid levels, hepatic lipid droplet accumulation, plasma aminotransferase levels, and liver weight by decreasing lipogenesis and increasing fatty acid oxidation. As E. stolonifera-supplemented mice showed improvements in hyperglycemia, insulin resistance, and inflammation, compared to control mice, it is possible that the beneficial effects of E. stolonifera on obesity might be associated with decreased inflammation and insulin resistance. Collectively, these results indicate that E. stolonifera could be used as a novel means of preventing and treating obesity and obesity-related metabolic disorders.