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.

Isorhynchophylline exerts antidepressant-like effects in mice via modulating neuroinflammation and neurotrophins: involvement of the PI3K/Akt/GSK-3ß signaling pathway.

Isorhynchophylline (IRN), an oxindole alkaloid isolated from Uncaria rhynchophylla, elicited distinct antidepressant-like activity in mice. The present study aimed to investigate the antidepressant-like effects of IRN in chronic unpredictable mild stress (CUMS)-induced depressive-like behaviors in mice and to illustrate its possible mechanisms of action. The mice were subjected to CUMS for 6 wk and administered with IRN (20 or 40 mg/kg) daily by oral gavage for 3 wk. The PI3K/protein kinase B (Akt) inhibitor and glycogen synthase kinase-3ß (GSK-3ß) inhibitors were used to determine the involvement of the PI3K/Akt/GSK-3ß pathway in the antidepressant-like effects of IRN in the mice. The results showed that CUMS caused depression-like behaviors in the mice, such as behavioral despair by the forced swim test (FST) and anhedonia by the sucrose preference test. In addition, CUMS could significantly reduce the levels of nerve growth factor and brain-derived neurotrophic factor but markedly increase the release of TNF-α and IL-6 in the hippocampus and cerebral cortex of the mice. Western blotting analysis showed that CUMS markedly suppressed the levels of phosphorylated GSK-3ß (Ser9) and phosphorylated Akt (Ser473) but significantly enhanced the translocation of NF-κB p65 from cytosol to nuclei in the hippocampus and cerebral cortex of the mice. CUMS could also significantly increase the NF-κB binding activity in the hippocampus and cerebral cortex of the mice, whereas IRN treatment could significantly reverse the behavioral and biochemical changes induced by CUMS in the mice. Moreover, the antidepressant-like effect of IRN was completely abolished by the PI3K/Akt inhibitor. Combination treatment with IRN and GSK-3ß inhibitors in the mice exerted a synergistic anti-immobility action in the FST. The results of mechanistic investigations indicated that the antidepressant-like action of IRN was mediated, at least in part, by enhancing neurotrophins and attenuating neuroinflammation via modulating the PI3K/Akt/GSK-3ß pathway.-Xian, Y.-F., Ip, S.-P., Li, H.-Q., Qu, C., Su, Z.-R., Chen, J.-N., Lin, Z.-X. Isorhynchophylline exerts antidepressant-like effects in mice via modulating neuroinflammation and neurotrophins: involvement of the PI3K/Akt/GSK-3ß signaling pathway.

Patchouli Oil Attenuates High Fat Diet-induced Non-alcoholic Hepatic Steatosis.

Non-alcoholic fatty liver disease (NAFLD) is one of the most common chronic liver diseases worldwide. Nevertheless, no first-line therapy exists. Hepatic steatosis is the earliest stage of NAFLD, which is characterized by an accumulation of hepatic lipids. Patchouli oil (PO), which is isolated from the well-known Chinese herb named Pogostemon cablin (Blanco) Benth. (Lamiaceae), inhibits hepatic lipid accumulation effectively. However, its potential ability for the treatment of NAFLD had not been reported before. Thus, the objective of this study was to investigate the effectiveness of PO against hepatic steatosis and its underlying mechanisms. We used a high fat diet (HFD)-induced hepatic steatosis model of rats to estimate the effect of PO against NAFLD. Hematoxylin-eosin and oil red O staining were used to analyze the hepatic histopathological changes. ELISA, RT-qPCR, and Western blotting analysis were applied to evaluate the parameters for hepatic steatosis. Our results showed that PO significantly attenuated the lipid profiles and the serum enzymes, evidenced by quantitative and histopathological analyses. It also markedly down-regulated the expression of sterol regulatory element-binding protein 1 (SREPB-1c) with its downstream factors in de novo lipogenesis. And, likewise, in lipid export by very low-density lipoproteins (VLDL), related molecules were dramatically improved. Furthermore, PO observably normalized the aberrant peroxisome proliferator-activated receptor α (PPAR-α) signal in fatty acids oxidation. In conclusion, PO exerted a preventing effect against HFD-induced steatosis and might be due to decrease de novo lipogenesis, promote export of lipids, as well as owing to improve fatty acids oxidation.

Palmatine ameliorated murine colitis by suppressing tryptophan metabolism and regulating gut microbiota.

Inflammatory bowel disease (IBD), majorly include Crohn's disease (CD) and ulcerative colitis (UC), is chronic and relapsing inflammatory disorders of the gastrointestinal tract, which treatment options remain limited. Here we examined the therapeutic effects of an isoquinoline alkaloid, Palmatine (Pal), on mice experimental colitis induced by dextran sulfate sodium (DSS) and explored underlying mechanisms. Colitis was induced in BALB/c mice by administering 3% DSS in drinking water for 7 days. Pal (50 and 100 mg kg-1) and the positive drug Sulfasalazine (SASP, 200 mg kg-1) were orally administered for 7 days. Disease activity index (DAI) was evaluated on day 8, and colonic tissues were collected for biochemistry analysis. The fecal microbiota was characterized by high-throughput Illumina MiSeq sequencing. And plasma metabolic changes were detected by UPLC-MS. Our results showed that Pal treatment significantly reduced DAI scores and ameliorated colonic injury in mice with DSS-induced colitis. Mucosal integrity was improved and cell apoptosis was inhibited. Moreover, gut microbiota analysis showed that mice received Pal-treatment have higher relative abundance of Bacteroidetes and Firmicutes, but reduced amount of Proteobacteria. Moreover, Pal not only suppressed tryptophan catabolism in plasma, but also decreased the protein expression of indoleamine 2,3-dioxygenase 1 (IDO-1, the rate-limiting enzyme of tryptophan catabolism) in colon tissue. This is consolidated by molecular docking, which suggested that Pal is a potent IDO-1 inhibitor. Taken together, our findings demonstrate that Pal ameliorated DSS-induced colitis by mitigating colonic injury, preventing gut microbiota dysbiosis, and regulating tryptophan catabolism, which indicated that Pal has great therapeutic potential for colitis.

Patchouli alcohol ameliorates dextran sodium sulfate-induced experimental colitis and suppresses tryptophan catabolism.

Despite the increased morbidity of ulcerative colitis (UC) in recent years, available treatments remain unsatisfactory. Pogostemon cablin has been widely applied to treat a variety of gastrointestinal disorders in clinic for centuries, in which patchouli alcohol (PA, C15H26O) has been identified as the major active component. This study attempted to determine the bioactivity of PA on dextran sulfate sodium (DSS)-induced mice colitis and clarify the mechanism of action. Acute colitis was induced in mice by 3% DSS for 7 days. The mice were then given PA (10, 20 and 40mg/kg) or sulfasalazine (SASP, 200mg/kg) as positive control via oral administration for 7 days. At the end of study, animals were sacrificed and samples were collected for pathological and other analysis. In addition, a metabolite profiling and a targeted metabolite analysis, based on the Ultra-Performance Liquid Chromatography coupled with mass spectrometry (UPLC-MS) approach, were performed to characterize the metabolic changes in plasma. The results revealed that PA significantly reduced the disease activity index (DAI) and ameliorated the colonic injury of DSS mice. The levels of colonic MPO and cytokines involving TNF-α, IFN-γ, IL-1ß, IL-6, IL-4 and IL-10 also declined. Furthermore, PA improved the intestinal epithelial barrier by enhancing the level of colonic expression of the tight junction (TJ) proteins, for instance ZO-1, ZO-2, claudin-1 and occludin, and by elevating the levels of mucin-1 and mucin-2 mRNA. The study also demonstrated that PA inhibited the DSS-induced cell death signaling by modulating the apoptosis related Bax and Bcl-2 proteins and down-regulating the necroptosis related RIP3 and MLKL proteins. By comparison, up-regulation of IDO-1 and TPH-1 protein expression in DSS group was suppressed by PA, which was in line with the declined levels of kynurenine (Kyn) and 5-hydroxytryptophan (5-HTP) in plasma. The therapeutic effect of PA was evidently reduced when Kyn was given to mice. In summary, the study successfully demonstrated that PA ameliorated DSS-induced mice acute colitis by suppressing inflammation, maintaining the integrity of intestinal epithelial barrier, inhibiting cell death signaling, and suppressing tryptophan catabolism. The results provided valuable information and guidance for using PA in treatment of UC.

Patchoulene Epoxide Isolated from Patchouli Oil Suppresses Acute Inflammation through Inhibition of NF-κB and Downregulation of COX-2/iNOS.

According to the GC-MS analysis, compositional variation was observed between samples of patchouli oil, of which an unknown compound identified as patchoulene epoxide (PAO) was found only in the long-stored oil, whose biological activity still remains unknown. Therefore, the present study aimed to evaluate the potential anti-inflammatory activity with three in vivo inflammatory models: xylene-induced ear edema, acetic acid-induced vascular permeability, and carrageenan-induced paw edema. Further investigation into its underlying mechanism on carrageenan-induced paw edema was conducted. Results demonstrated that PAO significantly inhibited the ear edema induced by xylene, lowered vascular permeability induced by acetic acid and decreased the paw edema induced by carrageenan. Moreover, PAO markedly decreased levels of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), interleukin-6 (IL-6), prostaglandin E2 (PGE2), and nitric oxide (NO), but increased levels of interleukin-4 (IL-4) and interleukin-10 (IL-10). PAO was also shown to significantly downregulate the protein and mRNA expressions of cyclooxygenase-2 (COX-2) and inducible nitric-oxide synthase (iNOS). Western blot analysis revealed that PAO remarkably inhibited p50 and p65 translocation from the cytosol to the nucleus by suppressing IKKß and IκBα phosphorylation. In conclusion, PAO exhibited potent anti-inflammatory activity probably by suppressing the activation of iNOS, COX-2 and NF-κB signaling pathways.

The immune-regulating effect of Xiao'er Qixingcha in constipated mice induced by high-heat and high-protein diet.

BACKGROUND: Xiao'er Qixingcha (EXQ) has been extensively applied to relieve dyspepsia and constipation in children for hundreds of years in China. However, the therapeutic mechanism underlying its efficacy remained to be defined. The present study aimed to clarify the possible laxative and immune-regulating effects of EXQ on two models of experimental constipation in mice, which mimicked the pediatric constipation caused by high-heat and high-protein diet (HHPD). METHODS: The two models of constipated mice were induced by HHPD or HHPD + atropine respectively. To investigate the laxative and immune-regulating activities of EXQ, animals were treated with three doses of EXQ (0.75, 1.5 and 3 g/kg) for 7 consecutive days. The fecal output parameters (number and weight), weight of intestinal content and, the thymus and spleen indexes were measured. The levels of sIgA, IL-10, TNF-α and LPS in colon and serum were determined by ELISA. Furthermore, the pathological changes of colon tissue were examined after routine H&E staining. RESULTS: Both HHPD and HHPD + atropine treatments obviously inhibited the fecal output and reduced the colonic sIgA, prominently increased the levels of IL-10 and TNF-α in colonic tissue and elevated the contents of LPS in serum and colonic tissues. In contrast, oral administration of EXQ significantly improved the feces characters and dose-dependently decreased the intestinal changes in both models. In HHPD model test, EXQ efficaciously boosted the sIgA level in a dose-dependent manner, significantly elicited decreases in TNF-α and IL-10 levels, and evidently decreased the spleen and thymus indexes. In HHPD + atropine model test, EXQ treatment reversed the pathological changes by not only dramatically decreasing the spleen index and the levels of LPS and IL-10, but also markedly elevating the thymus index. Furthermore, microscopic observation revealed that EXQ treatment maintained the integrity of colonic mucosa, and protected the colonic tissues from inflammation in the both models. CONCLUSIONS: EXQ exhibited prominent laxative activity and effectively protected the colonic mucosal barrier in two models of constipated mice, of which the mechanism might be closely associated with its propulsive and immune-regulating properties. The current results not only validated the rationale for the clinical application of EXQ in pediatric constipation related symptoms, but also threw new light on the immune-inflammatory responses accompanied with chronic constipation pathology.

Synthesis of pogostone by one-step.

Pogostone, isolated from Pogostemon cablin, has many biological activities such as potential antibacterial, anticandida, and antifungal. Traditional extraction leads to low output of PO about 17.6 mg/kg from Herba Pogostemonis. The previous literature had reported a synthetic study and the yield had reached 4.48% with strictly controlled reaction conditions. The two methods above cannot meet the large demand of PO; we report a new synthesis method. 4-hydroxy-6-methyl-2-pyrone (1) was added in toluene, with the existence of acylation catalyst 4-dimethylaminopyridine (DMAP), 4-methylvaleric acid (2), and condensing agent dicyclohexylcarbodiimide (DCC), PO was synthesized after the combination of 3-carbon of (1) with 1-OH of (2) in the acylation reaction. The purity had reached 98%, determined by HPLC. The structure was confirmed by spectroscopic methods including infrared, electron ionization mass spectrometry, and nuclear magnetic resonance spectroscopy. PO was totally synthesized in one step including cyclization, with total yield of 27.2%.

Andrographolide sodium bisulphite-induced inactivation of urease: inhibitory potency, kinetics and mechanism.

BACKGROUND: The inhibitory effect of andrographolide sodium bisulphite (ASB) on jack bean urease (JBU) and Helicobacter pylori urease (HPU) was performed to elucidate the inhibitory potency, kinetics and mechanism of inhibition in 20 mM phosphate buffer, pH 7.0, 2 mM EDTA, 25 °C. METHODS: The ammonia formations, indicator of urease activity, were examined using modified spectrophotometric Berthelot (phenol-hypochlorite) method. The inhibitory effect of ASB was characterized with IC50 values. Lineweaver-Burk and Dixon plots for JBU inhibition of ASB was constructed from the kinetic data. SH-blocking reagents and competitive active site Ni2+ binding inhibitors were employed for mechanism study. Molecular docking technique was used to provide some information on binding conformations as well as confirm the inhibition mode. RESULTS: The IC50 of ASB against JBU and HPU was 3.28±0.13 mM and 3.17±0.34 mM, respectively. The inhibition proved to be competitive and concentration- dependent in a slow-binding progress. The rapid formation of initial ASB-JBU complex with an inhibition constant of Ki=2.86×10(-3) mM was followed by a slow isomerization into the final complex with an overall inhibition constant of Ki*=1.33×10(-4) mM. The protective experiment proved that the urease active site is involved in the binding of ASB. Thiol reagents (L-cysteine and dithiothreithol) strongly protect the enzyme from the loss of enzymatic activity, while boric acid and fluoride show weaker protection, indicating that the active-site sulfhydryl group of JBU was potentially involved in the blocking process. Moreover, inhibition of ASB proved to be reversible since ASB-inactivated JBU could be reactivated by dithiothreitol application. Molecular docking assay suggested that ASB made contacts with the important sulfhydryl group Cys-592 residue and restricted the mobility of the active-site flap. CONCLUSIONS: ASB was a competitive inhibitor targeting thiol groups of urease in a slow-binding manner both reversibly and concentration-dependently, serving as a promising urease inhibitor for the treatment of urease-related diseases.

Selective antibacterial activity of patchouli alcohol against Helicobacter pylori based on inhibition of urease.

The aim of this study is to evaluate the antibacterial activity and urease inhibitory effects of patchouli alcohol (PA), the bioactive ingredient isolated from Pogostemonis Herba, which has been widely used for the treatment of gastrointestinal disorders. The activities of PA against selected bacteria and fungi were determined by agar dilution method. It was demonstrated that PA exhibited selective antibacterial activity against Helicobacter pylori, without influencing the major normal gastrointestinal bacteria. Noticeably, the antibacterial activity of PA was superior to that of amoxicillin, with minimal inhibition concentration value of 78 µg/mL. On the other hand, PA inhibited ureases from H.pylori and jack bean in concentration-dependent fashion with IC50 values of 2.67 ± 0.79 mM and 2.99 ± 0.41 mM, respectively. Lineweaver-Burk plots indicated that the type of inhibition was non-competitive against H.pylori urease whereas uncompetitive against jack bean urease. Reactivation of PA-inactivated urease assay showed DL-dithiothreitol, the thiol reagent, synergistically inactivated urease with PA instead of enzymatic activity recovery. In conclusion, the selective H.pylori antibacterial activity along with urease inhibitory potential of PA could make it a possible drug candidate for the treatment of H.pylori infection.

Characterization of solid dispersions of Patchouli alcohol with different polymers: effects on the inhibition of reprecipitation and the improvement of dissolution rate.

Solid dispersion technique is known to be an effective approach for the polymer to keep drugs stable in the solid state, thereby improving the dissolution rate and oral bioavailability through inhibiting reprecipitation in supersaturated solution. In this study, to evaluate the inhibitory effect of polyethylene glycol-6000 (PEG), Polyvinylpyrrolidone K30 (PVP) and Aminoalkyl methacrylate copolymer (Eudragit), the reprecipitation profiles were observed from supersaturated solutions of Patchouli alcohol (PA) in the presence and absence of the polymers. Furthermore, the dissolution profiles of PA solid dispersions formulated with PEG, PVP or Eudragit were compared for investigating the effect on improving dissolution of each polymer. Solid dispersions formulated with Eudragit were found to result in solution with the highest extent of supersaturation. By contrast, PEG and PVP were less effective. At equivalent supersaturation, all three polymers are capable of mitigating reprecipitation relative to that of PA alone. In addition, in the PA solid dispersion with Eudragit (E-SD (1/3)), the highest concentration of supersaturation of PA was maintained for prolonged time. These results unambiguously indicate that it is imperative to select the appropriate polymer and drug/polymer ratio in addition to considering the stability of the supersaturated solution, which was generated following dissolution of amorphous solid dispersion.

The protective effects of the supercritical-carbon dioxide fluid extract of Chrysanthemum indicum against lipopolysaccharide-induced acute lung injury in mice via modulating Toll-like receptor 4 signaling pathway.

The supercritical-carbon dioxide fluid extract of Chrysanthemum indicum Linné. (CFE) has been demonstrated to be effective in suppressing inflammation. The aim of this study is to investigate the preventive action and underlying mechanisms of CFE on acute lung injury (ALI) induced by lipopolysaccharide (LPS) in mice. ALI was induced by intratracheal instillation of LPS into lung, and dexamethasone was used as a positive control. Results revealed that pretreatment with CFE abated LPS-induced lung histopathologic changes, reduced the wet/dry ratio and proinflammatory cytokines productions (TNF-α, IL-1ß, and IL-6), inhibited inflammatory cells migrations and protein leakages, suppressed the levels of MPO and MDA, and upregulated the abilities of antioxidative enzymes (SOD, CAT, and GPx). Furthermore, the pretreatment with CFE downregulated the activations of NF-κB and the expressions of TLR4/MyD88. These results suggested that CFE exerted potential protective effects against LPS-induced ALI in mice and was a potential therapeutic drug for ALI. Its mechanisms were at least partially associated with the modulations of TLR4 signaling pathways.

Protective effects of hydroxysafflor yellow A on ß-amyloid-induced neurotoxicity in PC12 cells.

The accumulation of extracellular amyloid-ß peptide (Aß) has been considered as one of the important causes of Alzheimer's disease (AD), the most prevalent form of dementia. Hydroxysafflor yellow A (HSYA), a major active chemical component isolated from Carthamus tinctorius L., has been shown to possess neuroprotective actions in various ischemic models in vivo. The present study aimed to investigate the potential protective effect of HSYA against Aß-induced neurotoxicity in cultured rat pheochromocytoma (PC12) cells. The PC12 cells were pretreated with different concentrations (20, 40 and 80 µM) of HSYA for 2 h and then further treated with Aß (20 µM) for 24 h. The results showed that Aß could significantly decrease cell viability, glutathione level, mitochondrial membrane potential and the ratio of Bcl-2/Bax protein expression, while elevate the release of lactate dehydrogenase, the formation of DNA fragmentation, the levels of malondialdehyde and intracellular reactive oxygen species in PC12 cells. However, pretreatment with HSYA could effectively reverse these changes induced by Aß in PC12 cells. Our experimental results demonstrate that HSYA may be a potential neuroprotective agent warranting further development for treatment of AD.

Gastroprotective and anti-Helicobacter pylori potential of herbal formula HZJW: safety and efficacy assessment.

BACKGROUND: A traditional Chinese Medicine (TCM) formula, HZJW, has been applied in clinics in China for gastrointestinal disorders. However, the therapeutic mechanism underlying its efficacy and safety remained to be defined. The present investigation was undertaken to evaluate the formula HZJW for its gastroprotective potential, possible effect on Helicobacter pylori along with safety to justify its anti-ulcer action and safe clinical application. METHODS: The gastroduodenal cytoprotective potential was evaluated in rodent experimental models (HCl/Ethanol and NSAID-induced ulcer protocols). The anti-H. pylori property was assessed by agar dilution assay in vitro and analysis in vivo including rapid urease test, immunogold test and histopathology. For toxicity assessment, acute toxicity study was performed according to fixed dose procedure with a single oral administration of HZJW to mice. In the oral chronic toxicity, rats (80 males, 80 females) were administrated HZJW orally in 0, 1000, 2500, or 5000 mg/kg/day doses for 26 weeks (n = 40/group of each sex). Clinical signs, mortality, body weights, feed consumption, ophthalmology, hematology, serum biochemistry, gross findings, organ weights and histopathology were examined at the end of the 13- and 26-week dosing period, as well as after the 4-week recovery period. RESULTS: In the HCl/Ethanol-induced ulcer model, it was observed that oral administration with HZJW (260, 520 and 1040 mg/kg) and ranitidine (250 mg/kg) significantly reduced the ulcerative lesion index (116.70 ± 36.4, 102.20 ± 18.20, 84.10 ± 12.1 and 73.70 ± 16.70) in a dose-dependent manner, respectively, with respect to control group (134.10 ± 31.69). Significant inhibition was also observed in ulcerative index from aspirin-induced ulcer model, with decreases of 35.40 ± 5.93, 31.30 ± 8.08, 26.80 ± 8.27and 20.40 ± 6.93 for the groups treated with HZJW and ranitidine, in parallel to controls (41.60 ± 10.80). On the other hand, treatment with HZJW efficaciously eradicated H. pylori in infected mice in rapid urease test (RUT) and immunogold antibody assay, as further confirmed by reduction of H. pylori presence in histopathological analysis. In the in vitro assay, MICs for HZJW and amoxicillin (positive control) were 125 and 0.12 µg/mL respectively. The LD50 of HZJW was over 18.0 g/kg for mice. No drug-induced abnormalities were found as clinical signs, body weight, food consumption, hematology, blood biochemistry, ophthalmology and histopathology results across three doses. No target organ was identified. The No Observed Adverse Effect Level (NOAEL) of HZJW was determined to be 5,000 mg/kg/day for both sexes, a dose that was equivalent to 50 times of human dose. CONCLUSIONS: These results suggested the efficacy and safety of HZJW in healing peptic ulcer and combating H. pylori, which corroborated their conventional indications and contributed to their antiulcer pharmacological validation, lending more credence to its clinical application for the traditional treatment of stomach complaints symptomatic of peptic ulcer disease (PUD). HZJW might have the potential for further development as a safe and effective alternative/complementary to conventional medication in treating gastrointestinal (GI) disorders.

Anti-inflammatory property of the ethanol extract of the root and rhizome of Pogostemon cablin (Blanco) Benth.

The aim of this study was to investigate the anti-inflammatory property of the ethanol extract of the root and rhizome of Pogostemon cablin (ERP). The anti-inflammatory effect was evaluated using four animal models including xylene-induced mouse ear edema, acetic acid-induced mouse vascular permeability, carrageenan-induced mouse pleurisy, and carrageenan-induced mouse hind paw edema. Results indicated that oral administration of ERP (120, 240, and 480 mg/kg) significantly attenuated xylene-induced ear edema, decreased acetic acid-induced capillary permeability, inhibited carrageenan-induced neutrophils recruitment, and reduced carrageenan-induced paw edema, in a dose-dependent manner. Histopathologically, ERP (480 mg/kg) abated inflammatory response of the edema paw. Preliminary mechanism studies demonstrated that ERP decreased the level of MPO and MDA, increased the activities of anti-oxidant enzymes (SOD, GPx, and GRd), attenuated the productions of TNF-α, IL-1ß, IL-6, PGE2 and NO, and suppressed the activities of COX-2 and iNOS. This work demonstrates that ERP has considerable anti-inflammatory potential, which provided experimental evidences for the traditional application of the root and rhizome of Pogostemon cablin in inflammatory diseases.

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.

Protective effect of isorhynchophylline against ß-amyloid-induced neurotoxicity in PC12 cells.

Beta-amyloid peptide (Aß), a major protein component of senile plaques, has been considered as a critical cause in the pathogenesis of Alzheimer's disease (AD). Modulation of the Aß-induced neurotoxicity has emerged as a possible therapeutic approach to ameliorate the onset and progression of AD. The present study aimed to evaluate the protective effect of isorhynchophylline, an oxindole alkaloid isolated from a Chinese herb Uncaria rhynchophylla, on Aß-induced neurotoxicity in cultured rat pheochromocytoma (PC12) cells. The results showed that pretreatment with isorhynchophylline significantly elevated cell viability, decreased the levels of intracellular reactive oxygen species and malondialdehyde, increased the level of glutathione, and stabilized mitochondrial membrane potential in Aß(25-35)-treated PC12 cells. In addition, isorhynchophylline significantly suppressed the formation of DNA fragmentation and the activity of caspase-3 and moderated the ratio of Bcl-2/Bax. These results indicate that isorhynchophylline exerts a neuroprotective effect against Aß(25-35)-induced neurotoxicity in PC12 cells, at least in part, via inhibiting oxidative stress and suppressing the mitochondrial pathway of cellular apoptosis.

Protective effects of pinostrobin on ß-amyloid-induced neurotoxicity in PC12 cells.

Beta-Amyloid peptide (Aß), a major protein component of brain senile plaques in Alzheimer's disease (AD), has been considered as a critical cause in the pathogenesis of AD. Pinostrobin, a potent flavonoid inducer, is the major and most active ingredient of Folium cajani. The present study aimed to investigate whether pinostrobin could provide protective effect against Aß(25-35)-induced neurotoxicity in cultured rat pheochromocytoma (PC12) cells. The PC12 cells were pretreated with different concentrations of pinostrobin for 2 h, followed by the challenge with 20 µM Aß(25-35) for 24 h. The results showed that pretreatment with pinostrobin significantly elevated cell viability, decreased the lactate dehydrogenase activity, the levels of intracellular reactive oxygen species and calcium, and mitochondrial membrane potential in Aß(25-35)-treated PC12 cells. In addition, pinostrobin significantly suppressed the formation of DNA fragmentation and increased the ratio of Bcl-2/Bax. These results indicate that pinostrobin was able to exert a neuroprotective effect against Aß(25-35)-induced neurotoxicity in PC12 cells, at least in part, via inhibiting oxidative damage and calcium overload, as well as suppressing the mitochondrial pathway of cellular apoptosis.

Fingerprinting and simultaneous determination of alkaloids in Picrasma quassioides from different locations by high performance liquid chromatography with photodiode array detection.

A simple and sensitive method was developed and validated for profiling and simultaneous quantitation of seven alkaloids (6-hydroxy-ß-carboline-1-carboxylic acid, ß-carboline-1-carboxylic acid, ß-carboline-1-propanoic acid, 3-methylcanthin-5,6-dione, 5-hydroxy-4-methoxycanthin-6-one, 1-methoxycarbony-ß-carboline, and 4,5-dimethoxycanthin-6-one) in Picrasma quassioide grown in different locations by high-performance liquid chromatography with photodiode array detection. The analysis was conducted on a Phenomenex Gemini C(18) column at 35°C with mobile phase of 25 mM aqueous ammonium acetate (pH 4.0, adjusted by glacial acetate acid) and acetonitrile. A common fingerprint chromatograph under 254 nm consisting of 27 peaks was constructed for the evaluation of the similarities among 31 P. quassioide samples. Samples from Guangdong and Guangxi Provinces were found to be within group linkage and showed significant difference from that of Jiangxi Province origin by using principal component analysis and hierarchical clustering analysis. In addition, the seven alkaloids were identified by electrospray ionization mass spectrometry and comparing with reference standards and literature data. All of them were determined simultaneously using the established HPLC method. This rapid and effective analytical method could be employed for quality assessment of P. quassioide, as well as pharmaceutical products containing this herbal material.

Patchouli alcohol ameliorates the learning and memory impairments in an animal model of Alzheimer's disease via modulating SIRT1.

BACKGROUND: Alzheimer's disease (AD) is one of the most prevalent neurodegenerative diseases. Patchouli alcohol (PA), a major active ingredient isolated from Pogostemonis Herba, exhibits extensive bioactivity in the central nervous system (CNS) and exerts neuroprotective effects. PURPOSE: This study aimed to investigate the anti-AD effects of PA in an animal model of AD and to elucidate the underlying molecular mechanisms. METHODS: The gas chromatography (GC) was used to determine the ability of PA to pass the blood-brain barrier (BBB) in rats after oral administration. The sporadic AD rat model was established by intracerebroventricularly (ICV) injection with streptozotocin (STZ). PA (25 and 50 mg/kg) was given to rat orally once daily for 42 consecutive days. Morris water maze (MWM) test was performed to determine the learning and memory functions of the STZ-induced AD rats. EX527, a silent information regulator 1 (SIRT1) selective inhibitor, was used to investigate the involvement of SIRT1 in the anti-AD effects of PA in rats. RESULTS: PA could penetrate the BBB. MWM test results showed that PA could significantly ameliorate the learning and memory deficits induced by STZ in rats. Meanwhile, PA enhanced the expression of SIRT1, and markedly alleviated the tau pathology by inhibiting the hyperacetylation (at the site of Lys174) and hyperphosphorylation (at the sites of Thr181, Thr205, Ser396 and Ser404) of tau protein. PA also efficiently suppressed the activation of microglia and astrocytes, and the beta-amyloid (Aß) expression and the deacetylation of nuclear factor-kappa B (NF-κB) at Lys 310 (K310) in the STZ-treated AD rats. EX527, a SIRT1 selective inhibitor, could partially abolish the cognitive deficits improving effect of PA and inhibit the down-regulation of acetylated tau and acetylated NF-κB p65, suggesting that PA exhibited neuroprotective effects against AD via upregulating SIRT1. CONCLUSION: This study reported for the first time that PA could penetrate the BBB to exert its protective effects on the brain after a single-dose oral administration. The current experimental findings also amply demonstrated that PA could improve the cognitive and memory impairments in the STZ-induced AD rat model. The underlying mechanisms involve the alleviations of neuroinflammation, tau pathology and Aß deposition via modulating of SIRT1 and NF-κB pathways. All these findings strongly suggest that PA is a promising naturally occurring compound worthy of further development into an anti-AD pharmaceutical.