Berberine alleviates fructose-induced hepatic injury via ADK/AMPK/Nrf2 pathway: A novel insight.

Berberine (BBR) is a major active component of traditional Chinese medicine Rhizoma Coptidis and Cortex Phellodendri, which have been frequently used to treat liver diseases. Oxidative stress and inflammation are two pivotal hepatic pathological hallmarks. This study aimed to explore the potential effect and underlying mechanism of BBR on fructose-induced rat liver injury model, and hepatocyte damage in HepG2 and BRL-3A cells. Our results indicated that BBR effectively reversed fructose-induced body weight gain, glucose intolerance, and insulin resistance, observably attenuated abnormal histopathological alterations and ameliorated serum activities of ALT and AST. In vivo and in vitro, BBR significantly alleviated the secretion of pro-inflammatory cytokines IL-6 and TNF-α, and elevated levels of anti-inflammatory cytokine IL-10. BBR also attenuated oxidative stress by markedly decreasing intracellular contents of ROS and MDA, and increasing SOD enzymatic activity and GSH level. Furthermore, BBR substantially upregulated the protein expression of Nrf2, HO-1 and p-AMPK, and the fluorescence level of p-AMPK. In addition, BBR significantly increased the level of AMP, the ratio of AMP/ATP, and promoted the expression of ADK. Nevertheless, siADK abolished the benefits exerted by BBR on HepG2 and BRL-3A cells. Conclusively, the hepatoprotective effect of BBR was believed to be intimately associated with anti-inflammatory and antioxidant action mediated, at least partially, via ADK/AMPK/Nrf2 signaling. This work provided further support for the traditional application of Rhizoma Coptidis and Cortex Phellodendri in liver protection and might shed novel dimension to the clinical application of BBR, providing a promising lead compound for drug design.

Sulfonation metabolism in the gut microbiota is the main metabolic pathway of cholesterol in hypercholesterolemic mice.

The interactions between dietary cholesterol and intestinal microbiota strongly affect host health. Sulfonation is a major conjugating pathway responsible for regulating the chemical and functional homeostasis of endogenous and exogenous molecules. However, the role of cholesterol sulfonation metabolism in the host remains unclear. This work was designed to profile cholesterol-specific host-microbe interaction and conversion focusing on cholesterol sulfonation metabolism. Results indicated that the serum and fecal cholesterol sulfate (CHS) levels were significantly higher than those of total bile acid (TBA) levels in hypercholesterolemic mice. Deletion of the gut microbiota by antibiotics could dramatically increase total cholesterol (TC) levels but it decreased CHS levels in a pseudo-germ-free (PGF) mouse host. 16S rRNA gene sequencing assay and correlation analysis between the abundance of various intestinal bacteria (phylum and class) and the CHS/TC ratio showed that the intestinal genera Bacteroides contributed essentially to cholesterol sulfonation metabolism. These results were further confirmed in an in situ and ex vivo mouse intestinal model, which indicated that the sulfonation metabolism rate of cholesterol could reach 42% under high cholesterol conditions. These findings provided new evidence that the sulfonation metabolic pathway dominated cholesterol metabolism in hypercholesterolemic mice and microbial conversion of cholesterol-to-CHS was of vital importance for cholesterol-lowering by Bacteroides. This suggested that the gut microbiota could regulate cholesterol metabolism and that it was feasible to reduce cholesterol levels by dietary interventions involving the gut microbiota.

Patchouli alcohol alleviates metabolic dysfunction-associated steatohepatitis via inhibiting mitochondria-associated endoplasmic reticulum membrane disruption-induced hepatic steatosis and inflammation in rats.

Metabolic dysfunction-associated steatohepatitis (MASH) is a severe metabolic dysfunction-associated steatotic liver disease (MASLD) characterized by abnormal hepatic steatosis and inflammation. Previous studies have shown that Patchouli alcohol (PA), the primary component of Pogostemonis Herba, can alleviate digestive system diseases. However, its protection against MASH remains unclear. This study explored the protective effects and underlying mechanism of PA against high-fat diet-induced MASH in rats. Results showed that PA considerably reduced body weight, epididymal fat, and liver index and attenuated liver histological injury in MASH rats. PA alleviated hepatic injury by inhibiting steatosis and inflammation. These effects are associated with the improvement of SREBP-1c- and PPARα-mediated lipid metabolism and inhibition of the STING-signaling pathway-mediated inflammatory response. Moreover, PA-inhibited hepatic endoplasmic reticulum (ER) stress and mitochondrial dysfunction, reducing SREBP-1c and STING expressions and enhance PPARα expression. PA treatment had the strongest effect on the regulation of mitogen fusion protein 2 (Mfn2) in inhibiting mitochondrial dysfunction. Mfn2 is an important structural protein for binding ERs and mitochondria to form mitochondria-associated ER membranes (MAMs). MASH-mediated disruption of MAMs was inhibited after PA treatment-induced Mfn2 activation. Therefore, the pharmacological effect of PA on MASH is mainly attributed to the inhibition of MAM disruption-induced hepatic steatosis and inflammation. The findings of this study may have implications for MASH treatment that do not neglect the role of Mfn2-mediated MAMs.

Kidney targeting and modulating macrophage polarization through AMPK signaling: Therapeutic mechanism of berberine in uric acid nephropathy.

Uric acid nephropathy (UAN), caused by a common metabolic disorder resulting from hyperuricemia (HUA), has an increasing incidence. Previous studies have shown that berberine (BBR) has clear urate-lowering and anti-inflammatory effects in UAN mice, but its mechanism needs to be further clarified. Therefore, Potassium Oxonate (PO) combined with hypoxanthine (HX) induced UAN mice model and MSU induced THP-1 cells polarization model were adopted to investigate the mechanism of BBR on UAN in terms of tissue distribution and molecular pharmacology. Study unveiled that BBR was first found to bind to red blood cells (RBCs), which were recognized and phagocytosed by monocytes, then recruited by the injured kidney. Subsequently, BBR was enriched and functional in damaged kidney. The results of in vivo experiments revealed that, BBR reduced UA, BUN, CRE levels as well as the release of TNF-α, IL-1ß, IL-18 and IL-6, and alleviated renal injury in UAN mice, as consistent with previous studies. Additionally, BBR decreased MCP-1 expression, while diminishing macrophage infiltration and decreasing M1 proportion as determined by RT-qPCR. In vitro experiments, demonstrated that MSU promoted inflammatory polarization of THP-1 cells, while BBR reduced synthesis of inflammatory factors and inhibited MSU-induced inflammatory polarization. These effects of BBR were dependent on AMPK activation along with indirect inhibition of NF-κB signaling pathway mediated. However, the anti-inflammatory and macrophage polarization regulation effects of BBR were completely reversed upon administration of Compound C, an AMPK inhibitor. Therefore, BBR ameliorated kidney injury via regulating macrophage polarization through AMPK, which has therapeutic potential for UAN patients.

9-Hydroxy-8-oxypalmatine, a novel liver-mediated oxymetabolite of palmatine, alleviates hyperuricemia and kidney inflammation in hyperuricemic mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Palmatine is a main bioactive alkaloid of Cortex Phellodendri, which has been commonly prescribed for the treatment of hyperuricemia (HUA) in China. The metabolites of palmatine were crucial to its prominent biological activity. 9-Hydroxy-8-oxypalmatine (9-OPAL) is a novel liver-mediated secondary oxymetabolite of palmatine. AIM OF THE STUDY: The current study was to assess the efficacy of 9-OPAL, a novel liver-mediated secondary oxymetabolite of palmatine derived from Cortex Phellodendri, in experimental HUA mouse model and further explore its underlying mechanism. MATERIALS AND METHODS: An in vitro metabolic experiment with oxypalmatine was carried out using liver samples. We separated and identified a novel liver metabolite, and investigated its anti-HUA effect in mice. HUA mice were induced by potassium oxonate and hypoxanthine daily for one week. After 1 h of modeling, mice were orally administered with different doses of 9-OPAL (5, 10 and 20 mg/kg). The pathological changes of the kidneys were evaluated using hematoxylin-eosin staining (H&E). The acute toxicity of 9-OPAL was assessed. The effects of 9-OPAL on serum levels of uric acid (UA), adenosine deaminase (ADA), xanthine oxidase (XOD), creatinine (CRE), blood urea nitrogen (BUN) and inflammatory cytokines were measured by enzyme-linked immunosorbent assay (ELISA) or biochemical method. Furthermore, Western blot, quantitative real-time PCR (qRT-PCR) and molecular docking were used to investigate the effect of 9-OPAL on the expression of renal urate transporters and NLRP3 signaling pathway in HUA mice. RESULTS: 9-OPAL had been discovered to be a novel liver-mediated oxymetabolite of palmatine for the first time. Treatment with 9-OPAL significantly reduced the UA, CRE as well as BUN levels, and also effectively attenuated abnormal renal histopathological deterioration with favorable safety profile. Besides, 9-OPAL significantly decreased the serum and hepatic activities of XOD and ADA, dramatically inhibited the up-regulation of UA transporter protein 1 (URAT1) and glucose transporter protein 9 (GLUT9), and reversed the down-regulation of organic anion transporter protein 1 (OAT1). Additionally, 9-OPAL effectively mitigated the renal inflammatory markers (TNF-α, IL-1ß, IL-6 and IL-18), and downregulated the transcriptional and translational expressions of renal Nod-like receptor family pyrin domain containing 3 (NLRP3), caspase-1, apoptosis-associated speck-like (ASC) and IL-1ß in HUA mice. Molecular docking results revealed 9-OPAL bound firmly with XOD, OAT1, GLUT9, URAT1, NLRP3, caspase-1, ASC and IL-1ß. CONCLUSIONS: 9-OPAL was found to be a novel liver-mediated secondary metabolite of palmatine with favorable safety profile. 9-OPAL had eminent anti-hyperuricemic and renal-protective effects, and the mechanisms might be intimately associated with repressing XOD activities, modulating renal urate transporter expression and suppressing the NLRP3 inflammasome activation. Our investigation might also provide further experimental evidence for the traditional application of Cortex Phellodendri in the treatment of HUA.

Supersaturated Drug Delivery System of Oxyberberine Based on Cyclodextrin Nanoaggregates: Preparation, Characterization, and in vivo Application.

Propose: Oxyberberine (OBB), one of the main metabolites of berberine derived from intestinal and erythrocyte metabolism, exhibits appreciable anti-hyperuricemic activity. However, the low water solubility and poor plasma concentration-effect relationship of OBB hamper its development and utilization. Therefore, an OBB-hydroxypropyl-ß-cyclodextrin (HP-ß-CD) supersaturated drug delivery system (SDDS) was prepared and characterized in this work. Methods: OBB-HP-ß-CD SDDS was prepared using the ultrasonic-solvent evaporation method and characterized. Additionally, the in vitro and in vivo release experiments were conducted to assess the release kinetics of OBB-HP-ß-CD SDDS. Subsequently, the therapeutic efficacy of OBB-HP-ß-CD SDDS on hyperuricemia (HUA) was investigated by means of histopathological examination and evaluation of relevant biomarkers. Results: The results of FT-IR, DSC, PXRD, NMR and molecular modeling showed that the crystallized form of OBB was transformed into an amorphous OBB-HP-ß-CD complex. Dynamic light scattering indicated that this system was relatively stable and maintained by formation of nanoaggregates with an average diameter of 23 nm. The dissolution rate of OBB-HP-ß-CD SDDS was about 5 times higher than that of OBB raw material. Furthermore, the AUC0-t of OBB-HP-ß-CD SDDS (10.882 µg/mL*h) was significantly higher than that of the raw OBB counterpart (0.701 µg/mL*h). The oral relative bioavailability of OBB-HP-ß-CD SDDS was also enhanced by 16 times compared to that of the raw material. Finally, in vivo pharmacodynamic assay showed the anti-hyperuricemic potency of OBB-HP-ß-CD SDDS was approximately 5-10 times higher than that of OBB raw material. Conclusion: Based on our findings above, OBB-HP-ß-CD SDDS proved to be an excellent drug delivery system for increasing the solubility, dissolution, bioavailability, and anti-hyperuricemic potency of OBB.

Enhancing hepatoprotective action: oxyberberine amorphous solid dispersion system targeting TLR4.

Oxyberberine (OBB) is a significant natural compound, with excellent hepatoprotective properties. However, the poor water solubility of OBB hinders its release and absorption thus resulting in low bioavailability. To overcome these drawbacks of OBB, amorphous spray-dried powders (ASDs) of OBB were formulated. The dissolution, characterizations, and pharmacokinetics of OBB-ASDs formulation were investigated, and its hepatoprotective action was disquisitive in the D-GalN/LPS-induced acute liver injury (ALI) mouse model. The characterizations of OBB-ASDs indicated that the crystalline form of OBB active pharmaceutical ingredients (API) was changed into an amorphous form in OBB-ASDs. More importantly, OBB-ASDs showed a higher bioavailability than OBB API. In addition, OBB-ASDs treatment restored abnormal histopathological changes, improved liver functions, and relieved hepatic inflammatory mediators and oxidative stress in ALI mice. The spray drying techniques produced an amorphous form of OBB, which could significantly enhance the bioavailability and exhibit excellent hepatoprotective effects, indicating that the OBB-ASDs can exhibit further potential in hepatoprotective drug delivery systems. Our results provide guidance for improving the bioavailability and pharmacological activities of other compounds, especially insoluble natural compounds. Meanwhile, the successful development of OBB-ASDs could shed new light on the research process of poorly soluble medicine.

Discovery of 2,8-dihydroxyadenine in HUA patients with uroliths and biomarkers for its associated nephropathy.

Currently, it is acknowledged that gout is caused by uric acid (UA). However, some studies have revealed no correlation between gout and UA levels, and growing evidence suggests that 2,8-dihydroxyadenine (2,8-DHA), whose structural formula is similar to UA but is less soluble, may induce gout. Hence, we hypothesized that uroliths from hyperuricemia (HUA) patients, which is closely associated with gout, may contain 2,8-DHA. In this study, 2,8-DHA in uroliths and serum of HUA patients were determined using HPLC. Moreover, bioinformatics was used to investigate the pathogenic mechanisms of 2,8-DHA nephropathy. Subsequently, a mouse model of 2,8-DHA nephropathy established by the gavage administration of adenine, as well as a model of injured HK-2 cells induced by 2,8-DHA were used to explore the pathogenesis of 2,8-DHA nephropathy. Interestingly, 2,8-DHA could readily deposit in the cortex of the renal tubules, and was found in the majority of these HUA patients. Additionally, the differentially expressed genes between 2,8-DHA nephropathy mice and control mice were found to be involved in inflammatory reactions. Importantly, CCL2 and IL-1ß genes had the maximum degree, closeness, and betweenness centrality scores. The expressions of CCL2 and IL-1ß genes were significantly increased in the serum of 24 HUA patients with uroliths, indicating that they may be significant factors for 2,8-DHA nephropathy. Further analysis illustrated that oxidative damage and inflammation were the crucial processes of 2,8-DHA renal injury, and CCL2 and IL-1ß genes were verified to be essential biomarkers for 2,8-DHA nephropathy. These findings revealed further insights into 2,8-DHA nephropathy, and provided new ideas for its diagnosis and treatment.

Tetrahydroberberine alleviates high-fat diet-induced hyperlipidemia in mice via augmenting lipoprotein assembly-induced clearance of low-density lipoprotein and intermediate-density lipoprotein.

The promotion of excess low-density lipoprotein (LDL) clearance stands as an effective clinical approach for treating hyperlipidemia. Tetrahydroberberine, a metabolite of berberine, exhibits superior bioavailability compared to berberine and demonstrates a pronounced hypolipidemic effect. Despite these characteristics, the impact of tetrahydroberberine on improving excessive LDL clearance in hyperlipidemia has remained unexplored. Thus, this study investigates the potential effects of tetrahydroberberine on high-fat diet-induced hyperlipidemia in mice. The findings reveal that tetrahydroberberine exerts a more potent lipid-lowering effect than berberine, particularly concerning LDL-cholesterol in hyperlipidemic mice. Notably, tetrahydroberberine significantly reduces serum levels of upstream lipoproteins, including intermediate-density lipoprotein (IDL) and very low-density lipoprotein, by promoting their conversion to LDL. This reduction is further facilitated by the upregulation of hepatic LDL receptor expression induced by tetrahydroberberine. Intriguingly, tetrahydroberberine enhances the apolipoprotein E (ApoE)/apolipoprotein B100 (ApoB100) ratio, influencing lipoprotein assembly in the serum. This effect is achieved through the activation of the efflux of ApoE-containing cholesterol in the liver. The ApoE/ApoB100 ratio exhibits a robust negative correlation with serum levels of LDL and IDL, indicating its potential as a diagnostic indicator for hyperlipidemia. Moreover, tetrahydroberberine enhances hepatic lipid clearance without inducing lipid accumulation in the liver and alleviates existing liver lipid content. Importantly, no apparent hepatorenal toxicity is observed following tetrahydroberberine treatment for hyperlipidemia. In summary, tetrahydroberberine demonstrates a positive impact against hyperlipidemia by modulating lipoprotein assembly-induced clearance of LDL and IDL. The ApoE/ApoB100 ratio emerges as a promising diagnostic indicator for hyperlipidemia, showcasing the potential clinical significance of tetrahydroberberine in lipid management.

Self-Assembled nanoparticles Combining Berberine and Sodium Taurocholate for Enhanced Anti-Hyperuricemia Effect.

Propose: Berberine (BBR) is extensively studied as an outstanding anti-hyperuricemia drug. However, the clinical application of BBR was limited due to its poor absorption and low bioavailability. Therefore, there is an urgent necessity to find a novel drug formulation to address the issues of BBR in clinical application. Methods: Herein, we conducted the solubility, characterization experiments to verify whether BBR and sodium taurocholate (STC) self-assembled nanoparticles (STC@BBR-SANPs) could form. Furthermore, we proceeded the release experiment in vitro and in vivo to investigate the drug release effect. Finally, we explored the therapeutic effect of STC@BBR-SANPs on hyperuricemia (HUA) through morphological observation of organs and measurement of related indicators. Results: The solubility, particle size, scanning electron microscopy (SEM), and stability studies showed that the stable STC@BBR-SANPs could be formed in the BBR-STC system at ratio of 1:4. Meanwhile, the tissue distribution experiments revealed that the STC@BBR-SANPs could accelerate the absorption and distribution of BBR. In addition, the pharmacology study demonstrated that both BBR and STC@BBR-SANPs exhibited favorable anti-HUA effects and nephroprotective effects, while STC@BBR-SANPs showed better therapeutic action than that of BBR. Conclusion: This work indicated that STC@BBR-SANPs can be self-assembly formed, and exerts excellent uric acid-lowering effect. STC@BBR-SANPs can help to solve the problems of poor solubility and low absorption rate of BBR in clinical use, and provide a new perspective for the future development of BBR.

Curcumin Alleviates High-fat Diet-induced Nonalcoholic Steatohepatitis via Improving Hepatic Endothelial Function with Microbial Biotransformation in Rats.

Hepatic endothelial function is central to the development of nonalcoholic steatohepatitis (NASH). Curcumin (Cur) is reportedly hepatoprotective, however, it remains unknown whether Cur improves hepatic endothelial function in NASH. Additionally, the poor bioavailability of Cur renders it difficult to elucidate its hepatoprotective effect, hence, its biotransformation should be considered. Herein, we investigated the effects and mechanisms of Cur and its bioconversion on hepatic endothelial function against high-fat diet-induced NASH in rats. The results revealed that Cur improved hepatic lipid accumulation, inflammation, and endothelial dysfunction by inhibiting NF-κB and PI3K/Akt/HIF-1α pathways, however, these effects were weakened via antibiotic addition, which was closely related to reduced tetrahydrocurcumin (THC) produce in the liver and intestinal content. Moreover, THC exerted a better effect than Cur on restoring liver sinusoidal endothelial cells function to attenuate steatosis and injury in L02 cells. Thus, these findings indicate that the effect of Cur on NASH is closely related to hepatic endothelial function improvement with intestinal microbial biotransformation.

Intestines-erythrocytes-mediated bio-disposition deciphers the hypolipidemic effect of berberine from Rhizoma Coptidis: A neglected insight.

ETHNOPHARMACOLOGICAL RELEVANCE: Rhizoma Coptidis (RC), the dried rhizome of Coptis Chinensis Franch., can dispel dampness and heat within the body and has been traditionally used for the treatment of cardiovascular disease (CVD)-associated problems including hyperlipidemia in China. Berberine (BBR) is the main active component of RC, which has been shown to possess significant therapeutic potential. However, only 0.14% of BBR is metabolized in the liver, and the extremely low bioavailability (<1%) and blood concentration of BBR in experimental and clinical settings is insufficient to achieve the effects as observed under in vitro conditions, which imposes challenges to explain its excellent pharmacological actions. Intense efforts are currently being devoted to defining its specific pharmacological molecular targets, while the exploration from the perspective of its pharmacokinetic disposition has rarely been reported to date, which could hardly make a comprehensive understanding of its hypolipidemic enigma. AIM OF THE STUDY: This study made a pioneering endeavor to unveil the hypolipidemic mechanism of BBR from RC focusing on its unique intestines-erythrocytes-mediated bio-disposition. MATERIALS AND METHODS: The fate of BBR in intestines and erythrocytes was probed by a rapid and sensitive LC/MS-IT-TOF method. To analyze the disposition of BBR, a reliable HPLC method was subsequently developed and validated for simultaneous determination of BBR and its key active metabolite oxyberberine (OBB) in whole blood, tissues, and excreta. Meanwhile, the enterohepatic circulation (BDC) of BBR and OBB was verified by bile duct catheterization rats. Finally, lipid overloading models of L02 and HepG2 cells were employed to probe the lipid-lowering activity of BBR and OBB at in vivo concentration. RESULTS: The results showed that BBR underwent biotransformation in both intestines and erythrocytes, and converted into the major metabolite oxyberberine (OBB). The AUC0-t ratio of total BBR to OBB was approximately 2:1 after oral administration. Besides, the AUC0-t ratio of bound BBR to its unbound counterpart was 4.6:1, and this ratio of OBB was 2.5:1, indicative of abundant binding-type form in the blood. Liver dominated over other organs in tissue distribution. BBR was excreted in bile, while the excretion of OBB in feces was significantly higher than that in bile. Furthermore, the bimodal phenomenon of both BBR and OBB disappeared in BDC rats and the AUC0-t was significantly lower than that in the sham-operated control rats. Interestingly, OBB significantly decreased triglycerides and cholesterol levels in lipid overloading models of L02 and HepG2 cells at in vivo-like concentration, which was superior to the prodrug BBR. CONCLUSIONS: Cumulatively, BBR underwent unique extrahepatic metabolism and disposition into OBB by virtue of intestines and erythrocytes. BBR and OBB were mainly presented and transported in the protein-bound form within the circulating erythrocytes, potentially resulting in hepatocyte targeting accompanied by obvious enterohepatic circulation. The unique extrahepatic disposition of BBR via intestines and erythrocytes conceivably contributed enormously to its hypolipidemic effect. OBB was the important material basis for the hypolipidemic effect of BBR and RC.

Berberine attenuates uric acid-induced cell injury by inhibiting NLRP3 signaling pathway in HK-2 cells.

Hyperuricemia (HUA) is a common chronic metabolic disease that can cause renal failure and even death in severe cases. Berberine (BBR) is an isoquinoline alkaloid derived from Phellodendri Cortex with strong antioxidant, anti-inflammatory, and anti-apoptotic properties. The purpose of this study was to investigate the protective effects of berberine (BBR) against uric acid (UA)-induced HK-2 cells and unravel their regulatory potential mechanisms. The CCK8 assay was carried out to detect cell viability. The expression levels of inflammatory factors interleukin-1ß (IL-1ß) and interleukin-18 (IL-18) and Lactate dehydrogenase (LDH) were measured using Enzyme-linked immunosorbent assays (ELISA). The expression of the apoptosis-related protein (cleaved-Caspase3, cleaved-Caspase9, BAX, BCL-2) was detected by western blot. The effects of BBR on the activities of the NOD-like receptor family pyrin domain containing 3 (NLRP3) and the expression of the downstream genes were determined by RT-PCR and western blot in HK-2 cells. From the data, BBR significantly reversed the up-regulation of inflammatory factors (IL-1ß, IL-18) and LDH. Furthermore, BBR down-regulated protein expression of pro-apoptotic proteins BAX, cleaved caspase3 (cl-Caspase3), cleaved caspase9 (cl-Caspase9), and enhanced the expression of antiapoptotic protein BCL-2. Simultaneously, BBR inhibited the activated NLPR3 and reduced the mRNA levels of NLRP3, Caspase1, IL-18, and IL-1ß. Also, BBR attenuated the expression of NLRP3 pathway-related proteins (NLRP3, ASC, Caspase1, cleaved-Caspase1, IL-18, IL-1ß, and GSDMD). Furthermore, specific NLRP3-siRNA efficiently blocked UA-induced the level of inflammatory factors (IL-1ß, IL-18) and LDH and further inhibited activated NLRP3 pathway. Collectively, our results suggested that BBR can alleviate cell injury induced by UA. The underlying unctionary mechanism may be through the NLRP3 signaling pathway.

Brucea javanica oil alleviates intestinal mucosal injury induced by chemotherapeutic agent 5-fluorouracil in mice.

Background: Brucea javanica (L.) Merr, has a long history to be an anti-dysentery medicine for thousand of years, which is commonly called "Ya-Dan-Zi" in Chinese. The common liquid preparation of its seed, B. javanica oil (BJO) exerts anti-inflammatory action in gastrointestinal diseases and is popularly used as an antitumor adjuvant in Asia. However, there is no report that BJO has the potential to treat 5-Fluorouracil (5-FU)-induced chemotherapeutic intestinal mucosal injury (CIM). Aim of the study: To test the hypothesis that BJO has potential intestinal protection on intestinal mucosal injury caused by 5-FU in mice and to explore the mechanisms. Materials and methods: Kunming mice (half male and female), were randomly divided into six groups: normal group, 5-FU group (5-FU, 60 mg/kg), LO group (loperamide, 4.0 mg/kg), BJO group (0.125, 0.25, 0.50 g/kg). CIM was induced by intraperitoneal injection of 5-FU at a dose of 60 mg/kg/day for 5 days (from day 1 to day 5). BJO and LO were given orally 30 min prior to 5-FU administration for 7 days (from day 1 to day 7). The ameliorative effects of BJO were assessed by body weight, diarrhea assessment, and H&E staining of the intestine. Furthermore, the changes in oxidative stress level, inflammatory level, intestinal epithelial cell apoptosis, and proliferation, as well as the amount of intestinal tight junction proteins were evaluated. Finally, the involvements of the Nrf2/HO-1 pathway were tested by western blot. Results: BJO effectively alleviated 5-FU-induced CIM, as represented by the improvement of body weight, diarrhea syndrome, and histopathological changes in the ileum. BJO not only attenuated oxidative stress by upregulating SOD and downregulating MDA in the serum, but also reduced the intestinal level of COX-2 and inflammatory cytokines, and repressed CXCL1/2 and NLRP3 inflammasome activation. Moreover, BJO ameliorated 5-FU-induced epithelial apoptosis as evidenced by the downregulation of Bax and caspase-3 and the upregulation of Bcl-2, but enhanced mucosal epithelial cell proliferation as implied by the increase of crypt-localized proliferating cell nuclear antigen (PCNA) level. Furthermore, BJO contributed to the mucosal barrier by raising the level of tight junction proteins (ZO-1, occludin, and claudin-1). Mechanistically, these anti-intestinal mucositis pharmacological effects of BJO were relevant for the activation of Nrf2/HO-1 in the intestinal tissues. Conclusion: The present study provides new insights into the protective effects of BJO against CIM and suggests that BJO deserves to be applied as a potential therapeutic agent for the prevention of CIM.

Stereoisomers of octahydrocurcumin, the hydrogenated metabolites of curcumin, display stereoselective activity on the CYP2E1 enzyme in L-02 cells.

As the final hydrogenated metabolite of curcumin, octahydrocurcumin (OHC) exhibits increased powerful bioactivities. The chiral and symmetric chemical structure indicated that there were two OHC stereoisomers, (3R,5S)-octahydrocurcumin (Meso-OHC) and (3S,5S)-octahydrocurcumin ((3S,5S)-OHC), which may induce different effects on metabolic enzymes and bioactivities. Thus, we detected OHC stereoisomers from rat metabolites (blood, liver, urine and feces) after oral administration of curcumin. In addition, OHC stereoisomers were prepared and then their different influences on cytochrome P450 enzymes (CYPs) and UDP-glucuronyltransferases (UGTs) in L-02 cells were tested to explore the potential interaction and different bioactivities. Our results proved that curcumin could be metabolised into OHC stereoisomers first. In addition, Meso-OHC and (3S,5S)-OHC exhibited slight induction or inhibition effects on CYP1A2, CYP2A6, CYP2C8, CYP2C9, CYP3A4 and UGTs. Furthermore, Meso-OHC exhibited more intensive inhibition toward CYP2E1 expression than (3S,5S)-OHC, ascribed to the different mode of binding to the enzyme protein (P < 0.05), which finally induced more effective liver protection effects in acetaminophen-induced L-02 cell injury.

High fructose-induced skeletal muscle insulin resistance could be alleviated by berberine via AMPD1 and ADSL.

AMP-activated protein kinase (AMPK) is a master regulator of energy homeostasis that is activated in response to an elevated intracellular AMP/ATP ratio. Although many studies have shown berberine is an AMPK activator widely used in metabolic syndrome, how to properly control AMPK activity remains obscure. Our present study aimed to examine the protective effect of berberine against fructose-induced insulin resistance in rats and L6 cells, as well as its potential activation mechanism on AMPK. The results showed that berberine effectively reversed body weight gain, Lee's index, dyslipidemia and insulin intolerance. Moreover, berberine alleviated inflammatory response, antioxidant capacity and promoted glucose uptake in vivo and in vitro. The beneficial effect was associated with upregulation of both Nrf2 and AKT/GLUT4 pathways, which were regulated by AMPK. Notably, berberine could increase the level of AMP and the ratio of AMP/ATP, then further activate AMPK. Mechanistic experiments revealed that berberine suppressed the expression of adenosine monophosphate deaminase 1 (AMPD1) and promoted the expression of adenylosuccinate synthetase (ADSL). Taken together, berberine exerted excellent therapeutic effect on insulin resistance. And its mode of action may be related to the AMP-AMPK pathway by regulating AMPD1 and ADSL.

Huangqin Decoction ameliorates ulcerative colitis by regulating fatty acid metabolism to mediate macrophage polarization via activating FFAR4-AMPK-PPARα pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Huangqin Decoction (HQD), a traditional Chinese medicine (TCM) formula chronicled in Shang Han Lun, is safe and effective for treatment of ulcerative colitis (UC). AIM OF THE STUDY: To investigate the effect of HQD against dextran sulfate sodium (DSS)-induced UC mice by regulating gut microbiota and metabolites, and further explore the mechanism of fatty acid metabolism on macrophage polarization. MATERIALS AND METHODS: Based on 3% dextran sulfate sodium (DSS)-induced UC mice model, clinical symptoms observation (body weight, DAI, and colon length) and histological inspection were used to evaluate the efficacy of HQD and fecal microbiota transplantation (FMT) from HQD-treated mice. The gut microbiota and metabolites were detected by 16S rRNA sequencing and metabolomics analysis. The parameters of fatty acid metabolism, macrophage polarization, and FFAR1/FFAR4-AMPK-PPARα pathway were analyzed by immunofluorescence analysis, western blotting, and real-time PCR. Then, the effects of FFAR1 and FFAR4 on macrophage polarization were examined by agonists based on LPS-induced RAW264.7 cell model. RESULTS: The results showed that FMT, like HQD, ameliorated UC by improving weight loss, restoring colon length, and reducing DAI scores and histopathological scores. Besides, HQD and FMT both enhanced the richness of gut microbiota, and modulated intestinal bacteria and metabolites to achieve a new balance. Untargeted metabolomics analysis revealed that fatty acids, especially long-chain fatty acids (LCFAs), dominated in HQD against DSS-induced UC by regulating the gut microenvironment. Further, FMT and HQD recovered the expression of fatty acid metabolism-related enzymes, and simultaneously activated FFAR1/FFAR4-AMPK-PPARα pathway but suppressed NF-κB pathway. Combined with cell experiment, HQD and FMT promoted macrophage polarization from M1 toward M2, which were well associated with anti-inflammatory cytokines and combined with the activated FFAR4. CONCLUSIONS: The mechanism of HQD against UC was related to regulating fatty acid metabolism to mediate M2 macrophage polarization by activating the FFAR4-AMPK-PPARα pathway.

Patchouli alcohol attenuates the cognitive deficits in a transgenic mouse model of Alzheimer's disease via modulating neuropathology and gut microbiota through suppressing C/EBPß/AEP pathway.

BACKGROUND: Alzheimer's disease (AD) is a chronic neurodegenerative disease characterized by progressive cognitive dysfunctions and behavioral impairments. Patchouli alcohol (PA), isolated from Pogostemonis Herba, exhibits multiple pharmacological properties, including neuroprotective effects. This study aimed to investigate the therapeutic effects of PA against AD using the TgCRND8 transgenic AD mouse model, and to explore the underlying mechanisms targeting CCAAT/enhancer-binding protein ß/asparagine endopeptidase (C/EBPß/AEP) signaling pathway. METHODS: After genotyping to confirm the transgenicity, drug treatments were administered intragastrically once daily to 3-month-old TgCRND8 mice for 4 consecutive months. Several behavioral tests were applied to assess different aspects of neurological functions. Then the brain and colon tissues were harvested for in-depth mechanistic studies. To further verify whether PA exerts anti-AD effects via modulating C/EBPß/AEP signaling pathway in TgCRND8 mice, adeno-associated virus (AAV) vectors encoding CEBP/ß were bilaterally injected into the hippocampal CA1 region in TgCRND8 mice to overexpress C/EBPß. Additionally, the fecal microbiota transplantation (FMT) experiment was performed to verify the potential role of gut microbiota on the anti-AD effects of PA. RESULTS: Our results showed that PA treatment significantly improved activities of daily living (ADL), ameliorated the anxiety-related behavioral deficits and cognitive impairments in TgCRND8 mice. PA modulated the amyloid precursor protein (APP) processing. PA also markedly reduced the levels of beta-amyloid (Aß) 40 and Aß42, suppressed Aß plaque burdens, inhibited tau protein hyperphosphorylation at several sites and relieved neuroinflammation in the brains of TgCRND8 mice. Moreover, PA restored gut dysbiosis and inhibited the activation of the C/EBPß/AEP signaling pathway in the brain and colon tissues of TgCRND8 mice. Interestingly, PA strikingly alleviated the AD-like pathologies induced by the overexpression of C/EBPß in TgCRND8 mice. Additionally, the FMT of fecal microbiota from the PA-treated TgCRND8 mice significantly alleviated the cognitive impairments and AD-like pathologies in the germ-free TgCRND8 mice. CONCLUSION: All these findings amply demonstrated that PA could ameliorate the cognitive deficits in TgCRND8 mice via suppressing Aß plaques deposition, hyperphosphorylation of tau protein, neuroinflammation and gut dysbiosis through inhibiting the activation of C/EBPß/AEP pathway, suggesting that PA is a promising naturally occurring chemical worthy of further development into the pharmaceutical treatment of AD.

Oxyberberrubine, a novel liver microsomes-mediated secondary metabolite of berberine, alleviates hyperuricemic nephropathy in mice.

BACKGROUND: Berberrubine (BRB), one of the major metabolites of berberine (BBR), exerts an anti-hyperuricemic effect even superior to BBR. Liver is an important location for drug transformation. Nevertheless, there are few studies on the bioactivities and metabolites of BRB. PURPOSE: We investigated whether oxyberberrubine (OBR), a liver metabolite of BRB, exerted urate-lowering and reno-protective effects in hyperuricemic mice. METHODS: Liver microsomes were used to incubate BRB for studying its biotransformation. We isolated and identified its new metabolite OBR, and investigated its anti-hyperuricemic and reno-protective effects. In this work, the hyperuricemic mice model was established by receiving potassium oxonate (PO) and hypoxanthine (HX) for 7 consecutive days. 1 h after modeling, different dosages of OBR (5, 10 and 20 mg/kg), BRB (20 mg/kg) or febuxostat (Fex, 5 mg/kg) were given mice by gavage. RESULTS: Results showed that OBR possessed potent anti-hyperuricemic and reno-protective effects in hyperuricemic mice. Serum uric acid (UA) level was lowered, and the activities of xanthine oxidase (XOD) as well as adenosine deaminase (ADA) in the liver were suppressed after treatment with OBR. Hepatic expressions of XOD were remarkably decreased at mRNA and protein levels by OBR treatment. In addition, OBR prominently alleviated renal injury, embodied in markedly reduced serum creatinine and blood urea nitrogen (BUN) levels, decreased inflammatory mediators (TNF-α, IL-1ß, IL-6 and IL-18) levels, mRNA expression of CYP27B1 and repairment of renal tissues damage. Besides, OBR down-regulated renal expression of urate transporter 1 (URAT1), glucose transporter 9 (GLUT9), NOD-like receptor 3 (NLRP3), apoptosis-associated speck-like protein containing CARD (ASC), and caspase-1 at mRNA and protein levels. CONCLUSIONS: In short, our study indicated that OBR possessed superior anti-hyperuricemic and reno-protective effects, at least in part, through the inhibition of XOD, URAT1, GLUT9 and NLRP3 inflammasome signaling pathway in the kidney.

Hypouricemic and nephroprotective effects of palmatine from Cortex Phellodendri Amurensis: A uric acid modulator targeting Keap1-Nrf2/NLRP3 axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Palmatine (Pal) is a major bioactive alkaloid originated from ancient Chinese herbal medicine Cortex Phellodendri Amurensis (CPA), which has long been applied to treat hyperuricemia (HUA)-related diseases. Pal possesses potent anti-inflammatory and anti-oxidant effects against metabolic diseases. However, its potential beneficial effect against PO (potassium oxonate)/HX (hypoxanthine)-induced HUA remains elusive. AIM OF THE STUDY: This study aimed to investigate the potential pharmacological effect and mechanism of Pal on PO/HX-induced HUA in mice. MATERIAL AND METHODS: A mouse model of HUA was established by co-administration of PO/HX once daily for 7 consecutive days. The HUA mice were orally given three doses (25, 50 and 100 mg/kg) of Pal daily for a week. Febuxostat (Feb, 5 mg/kg) was given as a positive control. At the scheduled termination of the experiment, the whole blood, liver and kidney were collected for subsequent analyses. The concentrations of uric acid (UA), creatinine (CRE) and blood urea nitrogen (BUN), and activities of adenosine deaminase (ADA) and xanthine oxidase (XOD) were evaluated. Histopathological alterations of the kidney were detected by H&E staining. The inflammatory and oxidative stress status was detected by assay kits. Additionally, key proteins involved in the urate transporter, Keap1-Nrf2 and TXNIP/NLRP3 signaling pathways were evaluated by immunohistochemistry and Western blotting. Finally, molecular docking was employed to probe the binding characteristics of Pal and target proteins Keap1, NLRP3, URAT1 and HO-1. RESULTS: Administration of Pal substantially decreased the elevated kidney weight, lowered UA, CRE and BUN levels, and attenuated abnormal histopathological alterations. Meanwhile, treatment with Pal also dramatically lowered hepatic XOD and ADA activities. Besides, Pal treatment effectively mitigated the renal inflammatory and oxidative stress markers. Further mechanistic investigation indicated Pal distinctly downregulated the protein levels of GLUT9 and URAT1, while up-regulated the expression levels of OAT1 and ABCG2. Pal also restored Nrf2 activation, promoted subsequent expression of anti-oxidative enzymes, and downregulated the expressions of TXNIP, NLRP3, apoptosis-associated speck-like (ASC), caspase-1, IL-1ß and IL-18. Molecular docking analysis also indicated Pal firmly bound with Keap1, NLRP3, URAT1 and HO-1. CONCLUSIONS: These findings indicated that Pal exhibited favorable anti-HUA effect via modulating the expressions of transporter-related proteins and suppressing XOD activity. Furthermore, Pal also alleviated HUA-induced kidney injury, which was at least partially related to restoring Keap1-Nrf2 pathway and inhibiting TXNIP/NLRP3 inflammasome. Our investigation was envisaged to provide experimental support for the traditional application of CPA and CPA-containing classical herbal formulas in the management of HUA-related diseases and might provide novel dimension to the clinical application of Pal.