Role and mechanism of MiR-542-3p in regulating TLR4 in nonylphenol-induced neuronal cell pyroptosis.

BACKGROUND: This study aimed to investigate the spatial learning/memory and motor abilities of rats and the alteration of miR-542-3p and pyroptosis in the midbrain nigrostriatal area in vivo after nonylphenol (NP) gavage and to explore the mechanism of miR-542-3p regulation of Toll-like receptor 4 (TLR4) in NP-induced pyroptosis in BV2 microglia in vitro. METHODS: In vivo: Thirty-six specific-pathogen-free-grade Sprague-Dawley rats were divided into three equal groups: blank control group (treated with pure corn oil), NP group (treated with NP, 80 mg/kg body weight per day for 90 days), and positive control group [treated with lipopolysaccharide (LPS), 2 mg/kg body weight for 7 days]. In vitro: The first part of the experiment was divided into blank group (control, saline), LPS group [1 µg/ml + 1 mM adenosine triphosphate (ATP)], and NP group (40 µmol/L). The second part was divided into mimics NC (negative control) group, miR-542-3p mimics group, mimics NC + NP group, and miR-542-3p mimics + NP group. RESULTS: In vivo: Behaviorally, the spatial learning/memory and motor abilities of rats after NP exposure declined, as detected via Y-maze, open field, and rotarod tests. Some microglia in the substantia nigra of the NP-treated rats were activated. The downregulation of miR-542-3p was observed in rat brain tissue after NP exposure. The mRNA/protein expression of pyroptosis-related indicators (TLR4), NOD-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein (ASC), gasdermin-D (GSDMD), cysteinyl aspartate-specific proteinase-1 (caspase-1), and interleukin-1ß (IL-1ß) in the substantia nigra of the midbrain increased after NP exposure. In vitro: ASC fluorescence intensity increased in BV2 cells after NP exposure. The mRNA and/or protein expression of pyroptosis-related indicators (TLR4, NLRP3, GSDMD, caspase-1, and IL-1ß) in BV2 cells was upregulated after NP exposure. The transfection of miR-542-3p mimics inhibited NP-induced ASC expression in BV2 cells. The overexpression of miR-542-3p, followed by NP exposure, significantly reduced TLR4, NLRP3, ASC, caspase-1, and IL-1ß gene and/or protein expression. CONCLUSIONS: This study suggested that NP exposure caused a decline in spatial learning memory and whole-body motor ability in rats. Our study was novel in reporting that the upregulation of miR-542-3p targeting and regulating TLR4 could inhibit NLRP3 inflammatory activation and alleviate NP-induced microglia pyroptosis.

Herbal formula Jiawei Xiaochengqi decoction prevents postoperative abdominal adhesion in a rat model through inhibition of CXCL2-CXCR2 pathway.

BACKGROUND: Postoperative abdominal adhesion (PAA) is the most common complication after abdominal surgeries, which can lead to intestinal obstruction, chronic abdominal pain or female infertility. Jiawei Xiaochengqi decoction (JWXCQ) is a hospital preparation widely used for PAA treatment in Nanfang Hospital of Southern Medical University for more than twenty years. PURPOSE: This study aimed to investigate the therapeutic effects and potential mechanism of JWXCQ against PAA and provide beneficial information for its clinical application. METHODS: The main active components of JWXCQ were identified using ultra high performance liquid chromatography (UHPLC) combined with standard substance comparison. The efficacy and underlying mechanism of JWXCQ were evaluated through in vivo experiments with a postsurgical-induced peritoneal adhesion rat model, and in vitro studies with LPS-stimulated Raw 264.7 macrophages and primary fibroblasts. H&E and Masson staining were performed to assess histopathological changes. The levels of cytokines/proteins-associated with inflammation and degradation of extracellular matrix as well as CXCL2-CXCR2 pathway-related proteins were determined by ELISA, qRT-PCR, western blot assays or immunohistochemistry, respectively. Furthermore, siCXCR2 transfection was used to validate the mechanism of action of JWXCQ. RESULTS: JWXCQ treatment significantly reduced the formation of PAA, inhibited the inflammation and collagen deposition, and facilitated the secretion of MMP9, decreased the levels of IL-1ß, IL-6, TIMP1, COL-1, and suppressed the CXCL2-CXCR2 pathway in PAA rats. Furthermore, JWXCQ inhibited its downstream pathways, the JAK2-STAT3 and PI3K-AKT signaling, as indicated by the suppression of the phosphorylation levels of STAT3 and AKT. In vitro cell experiments revealed that JWXCQ reduced IL-1ß and IL-6 secretion in Raw 264.7 macrophages and COL-1 in primary fibroblasts. The CXCL2-CXCR2, JAK2-STAT3 and PI3K-AKT pathways were also inhibited after JWXCQ treatment, which were consistent with the in vivo results. More importantly, silence of CXCR2 eliminated the regulatory effects of JWXCQ. CONCLUSION: JWXCQ could effectively prevent the PAA formation by alleviating inflammation and collagen deposition, which was associated with the inhibition of CXCL2-CXCR2 pathway. This study investigated the relevant pharmacological mechanisms of JWXCQ, providing further evidence for the application of JWXCQ in clinical PAA treatment.

Oridonin restores hepatic lipid homeostasis in an LXRα-ATGL/EPT1 axis-dependent manner.

Hepatosteatosis is characterized by abnormal accumulation of triglycerides (TG), leading to prolonged and chronic inflammatory infiltration. To date, there is still a lack of effective and economical therapies for hepatosteatosis. Oridonin (ORI) is a major bioactive component extracted from the traditional Chinese medicinal herb Rabdosia rubescens. In this paper, we showed that ORI exerted significant protective effects against hepatic steatosis, inflammation and fibrosis, which was dependent on LXRα signaling. It is reported that LXRα regulated lipid homeostasis between triglyceride (TG) and phosphatidylethanolamine (PE) by promoting ATGL and EPT1 expression. Therefore, we implemented the lipidomic strategy and luciferase reporter assay to verify that ORI contributed to the homeostasis of lipids via the regulation of the ATGL gene associated with TG hydrolysis and the EPT1 gene related to PE synthesis in a LXRα-dependent manner, and the results showed the TG reduction and PE elevation. In detail, hepatic TG overload and lipotoxicity were reversed after ORI treatment by modulating the ATGL and EPT1 genes, respectively. Taken together, the data provide mechanistic insights to explain the bioactivity of ORI in attenuating TG accumulation and cytotoxicity and introduce exciting opportunities for developing novel natural activators of the LXRα-ATGL/EPT1 axis for pharmacologically treating hepatosteatosis and metabolic disorders.

Correlation of Glucose Metabolism with Cancer and Intervention with Traditional Chinese Medicine.

Cancer is a complex disease with several distinct characteristics, referred to as "cancer markers" one of which is metabolic reprogramming, which is a common feature that drives cancer progression. Over the last ten years, researchers have focused on the reprogramming of glucose metabolism in cancer. In cancer, the oxidative phosphorylation metabolic pathway is converted into the glycolytic pathway in order to meet the growth requirements of cancer cells, thereby creating a microenvironment that promotes cancer progression. The precise mechanism of glucose metabolism in cancer cells is still unknown, but it is thought to involve the aberrant levels of metabolic enzymes, the influence of the tumor microenvironment (TME), and the activation of tumor-promoting signaling pathways. It is suggested that glucose metabolism is strongly linked to cancer progression because it provides energy to cancer cells and interferes with antitumor drug pharmacodynamics. Therefore, it is critical to unravel the mechanism of glucose metabolism in tumors in order to gain a better understanding of tumorigenesis and to lay the groundwork for future research into the identification of novel diagnostic markers and therapeutic targets for cancer treatment. Traditional Chinese Medicine (TCM) has the characteristics of multiple targets, multiple components, and less toxic side effects and has unique advantages in tumor treatment. In recent years, researchers have found that a variety of Chinese medicine monomers and compound recipes play an antitumor role by interfering with the reprogramming of tumor metabolism. The underlying mechanisms of metabolism reprogramming of tumor cells and the role of TCM in regulating glucose metabolism are reviewed in this study, so as to provide a new idea for antitumor research in Chinese medicine.

Magnetic carbon nanotubes-molecularly imprinted polymer coupled with HPLC for selective enrichment and determination of ferulic acid in traditional Chinese medicine and biological samples.

A molecularly imprinted polymer (MIP) with magnetic carbon nanotubes (MCNTs) as carrier was synthesized and used for the enrichment and determination of ferulic acid (FA) by high-performance liquid chromatography (HPLC). The morphology and structure of the FA magnetic carbon nanotubes-molecularly imprinted polymers (MCNTs@FA-MIPs) were characterized by Fourier transform infrared spectroscopy, scanning electron microscopy, transmission electron microscopy, and X-ray diffraction. The results demonstrated that the prepared MCNTs@FA-MIPs had excellent magnetic properties and uniform appearance. The adsorption properties of the novel MIP were studied by kinetic, and isothermal adsorption experiments. The results showed that the MCNTs@FA-MIPs exhibited relatively good uptake kinetics (equilibrium time, 2 h), high adsorption capacity (50 mg⋅g-1), fast separation, and good selectivity for the template molecule with a separation factor α of 1.73. The MCNTs@FA-MIPs combined with HPLC were successfully applied to the separation, enrichment, and determination of FA in a Ligusticum chuanxiong extract and in plasma of rats which had been administered with Taitai beauty essence. The recoveries for FA were 95.53-100.03 % with relative standard deviations (RSDs) less than 5.5%. The results confirmed that the proposed MCNTs@FA-MIPs offered efficient extraction of FA from traditional Chinese medicinal preparations and blood samples and with high specificity.

Imperatae rhizoma-Hedyotis diffusa Willd. herbal pair alleviates nephrotic syndrome by integrating anti-inflammatory and hypolipidaemic effects.

BACKGROUND: Nephrotic syndrome (NS) is a common nephropathy with a complex and diverse aetiology. Both Imperatae rhizoma and Hedyotis diffusa Willd. are herbs that are widely used as medicine and functional food. In traditional Chinese medicine theory, they are used as an herbal pair (HP) to treat inflammation-related diseases in the clinic, especially disorders of the kidney. PURPOSE: This study aimed to investigate the anti-inflammatory and hypolipidaemic effects of HP in an NS rat model and provide scientific data for its clinical application. METHODS: An NS model was established by two-dose injection of Sprague-Dawley rats with adriamycin. Seven groups, including the sham, model, HP treatment (0.25, 0.5 and 1.0 g/kg/d), prednisone (positive control, 5 mg/kg/d), and atorvastatin (positive control, 4 mg/kg/d) groups, were tested. The biochemical indexes of renal function and inflammatory cytokines were determined by ELISA kits and/or qPCR assays, and the crucial protein involved in the signalling pathway were subsequently tested by qPCR and/or Western blotting. Based on specific compounds identified by LC-Q-TOF-MS, network pharmacological study was carried out. RESULTS: The levels of BUN, Scr, Upro, UA, Alb, TC, TG, and LDL-C were significantly elevated in model rats. HP treatment for four weeks improved the renal function and the dyslipidaemia by decreasing the levels of all parameters, except BUN and Scr. HP treatment (0.5 and 1.0 g/kg/d) upregulated the expression of PPARγ, CYP7b1, and LDLR in the liver, while it down-regulated PCSK9, showing a regulatory effect on lipid metabolism disorder. The levels of TNF-α and IL-1ß in the plasma and the mRNA expression of TNF-α, IL-1ß, MCP-1, and TGF-ß1 in the kidney were decreased in HP groups, revealing its anti-inflammatory effect in NS rats. The HP exerted an alleviation effect on the inflammatory response through the NF-κB pathway by inhibiting the mRNA and protein expression of p50 and p65. There were 34 compounds identified or tentatively characterized in HP. In the network pharmacological study, PPARG(PPARγ), PCSK9, RELA(p65), and NF-κB1(p50) were the top 20 targets for HP, supporting the animal experimental results. CONCLUSION: HP exhibited protective effects on NS rats. These effects might be closely related to the inhibition of NF-κB and PCSK9-LDLR and activation of the PPARγ-CYP7B1 signalling pathways.

Isolation and Purification of Five Phloroglucinol Derivatives From Agrimonia Pilosa Ledeb by Counter-Current Chromatography Using Non-Aqueous Solvent Systems.

Five non-polar phloroglucinol derivatives, viz. pseudo-aspidin, α-kosin and agripinol A-C were isolated and purified from Agrimonia pilosa Ledeb by semi-preparative counter-current chromatography. The separation was performed by a two-step elution with non-aqueous solvent systems. In the first step, an elution mode of a two-phase solvent system consisting of n-hexane-acetonitrile-dichloromethane-methanol (6:6:0.5:0.5, v/v/v/v) was used. We obtained sample Ι containing three components (47.0 mg) and sample ΙΙ containing two components (24.8 mg) from crude extract (371.0 mg). In the second step, sample Ι was successfully separated by closed-loop recycling mode with a solvent system consisting of n-hexane-acetonitrile-dichloromethane (10:7:3, v/v/v), yielding 17.8 mg of pseudo-aspidin, 18.5 mg of α-kosin and 6.4 mg of agripinol A. The other two compounds-8.7 mg of agripinol B and 13.6 mg of agripinol C-were obtained from sample ΙΙ in the same manner. All the isolated compounds had a high purity exceeding 95%.

[Thermosensitive gel of polysaccharide from Ganoderma applanatum combined with paclitaxel for mice with 4T1 breast cancer].

Polysaccharide from Ganoderma applanatum has the activities of anti-tumor and enhancing immune function. There were no reports on antitumor effect of its intratumoral injection. In this study, the polysaccharide was extracted from G. applanatum by water extraction and alcohol precipitation, and purified by ceramic membrane after removing protein by Sevage method. The total polysaccharide content from G. applanatum(PGA)was about 63%. The combination of PGA and paclitaxel showed synergistic effect on cytotoxicity of 4 T1 cells at lower concentrations in vitro. In addition, the growth curve of 4 T1 cells showed that PGA could retard the growth of 4 T1 cells gradually. The PGA thermosensitive gel(PGA-TG)was prepared by using poloxamer 188 and 407. The gel temperature was 36 ℃, and the PGA-TG could effectively slow down the release rate of PGA in vitro. 4 T1 breast cancer-bearing mice were used as a model to evaluate the therapeutic effect of intratumoral injection of PGA combined with tail vein injection of nanoparticle albumin-bound paclitaxel(nab-PTX). In high and low dose PGA groups, each mice was given with 2.25, 1.125 mg PGA respectively, twice in total, and the dosage of paclitaxel was 15 mg·kg~(-1), once every 3 days, for a total of five times. The tumor inhibition rate was 29.65% in the high dose PGA-TG group, 58.58% in the nab-PTX group, 63.37% in low dose PGA-TG combined with nab-PTX group, and 68.10% in high dose PGA-TG combined with nab-PTX group respectively. The inhibitory effect in high dose PGA-TG group combined with nab-PTX on tumors was significantly higher than that in nab-PTX group(P<0.05). The results showed that paclitaxel therapy combined with intratumoral injection of PGA-TG could improve the therapeutic effect for 4 T1 mice and reduce the side effects of chemotherapy.

Metabolism and Toxicity of Emodin: Genome-Wide Association Studies Reveal Hepatocyte Nuclear Factor 4α Regulates UGT2B7 and Emodin Glucuronidation.

Emodin is the main toxic component in Chinese medicinal herbs such as rhubarb. Our previous studies demonstrated that genetic polymorphisms of UDP-glucuronosyltransferase 2B7 (UGT2B7) had an effect on the glucuronidation and detoxification of emodin. This study aimed to reveal the transcriptional regulation mechanism of UGT2B7 on emodin glucuronidation and its effect on toxicity. Emodin glucuronic activity and genome and transcriptome data were obtained from 36 clinical human kidney tissues. The genome-wide association studies (GWAS) identified that four single nucleotide polymorphisms (SNPs) (rs6093966, rs2868094, rs2071197, and rs6073433), which were located on the hepatocyte nuclear factor 4α (HNF4A) gene, were significantly associated with the emodin glucuronidation (p < 0.05). Notably, rs2071197 was significantly associated with the gene expression of HNF4A and UGT2B7 and the glucuronidation of emodin. The gene expression of HNF4A showed a high correlation with UGT2B7 (R2 = 0.721, p = 5.83 × 10-11). The luciferase activity was increased 7.68-fold in 293T cells and 2.03-fold in HepG2 cells, confirming a significant transcriptional activation of UGT2B7 promoter by HNF4A. The knockdown of HNF4A in HepG2 cells (36.6%) led to a significant decrease of UGT2B7 (19.8%) and higher cytotoxicity (p < 0.05). The overexpression of HNF4A in HepG2 cells (31.2%) led to a significant increase of UGT2B7 (24.4%) and improved cell viability (p < 0.05). Besides, HNF4A and UGT2B7 were both decreased in HepG2 cells and rats after treatment with emodin. In conclusion, emodin used long term or in high doses could inhibit the expression of HNF4A, thereby reducing the expression of UGT2B7 and causing hepatotoxicity.

Development of an immobilized liposome chromatography method for screening and characterizing α-glucosidase-binding compounds.

Immobilized liposome chromatography (ILC) is a powerful tool in screening and analyzing membrane-permeable components in natural medicinal herbs. In this study, the stationary phase of a new receptor liposome biomembrane chromatography (RLBC) was employed to screen the active ingredients in seven natural medicinal herbs. As a model system, α-glucosidase was immobilized in porous silica gel by incorporating α-glucosidase into liposome vesicles. Combined with HPLC, this stationary phase can be used to evaluate the interaction of liposome-protein compounds with compounds and drugs. The surface characteristics of the RLBC phase was characterized by Scanning electron microscope (SEM), Fourier transform infrared spectroscopy (FTIR) and Energy dispersive spectrometer (EDS). RLBC was successfully used to screen and analyze permeable compounds in natural medicinal herbs, and screen the extracts from Schisandra chinensis. This method was used to establish that Schisandrin in Schisandra chinensis is an active component. Furthermore, the hypoglycemic effect of Schisandrin was verified in vivo in rats. This study further modeled the relationship between the activity of inhibitor and retention behavior with RLBC with a mathematical equation. RLBC stationary phase combined with HPLC can be used for rapid screening of drug candidates.

Isoalantolactone inhibits RANKL-induced osteoclast formation via multiple signaling pathways.

The metabolicosteopathy known as postmenopausal osteoporosisiscaused by disruption of the balance between bone resorption and osteogenesis, processes that are mediated by osteoclasts and osteoblasts, respectively. The current therapeutic approaches to treating osteoporosis have several limitations. In this study, we demonstrated that the natural chemical compound isoalantolactone (IAL) could inhibit osteoclastogenesis, without affecting osteogenesis. This is the first study reporting a role of IAL in suppressing the receptor activator of nuclear factor-kappa B ligand (RANKL)-induced osteoclast formation in a dose-dependent manner, and downregulating the expression of osteoclast-related marker genes. Furthermore, IAL abrogated the phosphorylation of c-Jun N-terminal kinase (JNK)/p38, NF-κB, and phosphatidylinositol 3-kinase (PI3K)-AKT, and also diminished the expression of osteoclastogenesis-related proteins. In conclusion, our results indicated that IAL has promise for the treatment of osteoporosis and other metabolicbone diseases.

[Evaluation of key production processes of Schizonepetae Herba formula granules based on characteristic chromatogram and quantitative transfer relationship].

The aim of this paper was to evaluate the key production processes of Schizonepetae Herba formula granules based on the new model of combining characteristic chromatogram with quantitative transfer relationship. The rationality of production process design was evaluated by studying the intermediates in different processes of formula granules, analyzing the loss of index component pulegone in each step, and establishing the characteristic chromatogram. The content of pulegone in 10 batches of standard decoction ranged between 0.067% and 0.124%(70%-130% of the average value), and the transfer rate of pulegone was 44.58%-93.97%. After the improvement of the production process, the content of pulegone in Schizonepetae Herba formula granules was 0.093%, and the transfer rate of pulegone was 68.38%, which was consistent with the parameters range of standard decoction. This study emphasized the integrality of the research process of traditional Chinese medicine(TCM) formula granules, and provided a new idea for the quality control of TCM with content determination as the main evaluation index for a long time.

Glucuronidation and its effect on the bioactivity of amentoflavone, a biflavonoid from Ginkgo biloba leaves.

OBJECTIVES: Ginkgo biloba leaves contain amentoflavone (AMF), a dietary flavonoid that possesses antioxidant and anticancer activity. Flavonoids are extensively subjected to glucuronidation. This study aimed to determine the metabolic profile of AMF and the effect of glucuronidation on AMF bioactivity. METHODS: A pharmacokinetic study was conducted to determine the plasma concentrations of AMF and its metabolites. The metabolic profile of AMF was elucidated using different species of microsomes. The antioxidant activity of AMF metabolites was determined using DPPH/ABTS radical and nitric oxide assays. The anticancer activity of AMF metabolites was evaluated in U87MG/U251 cells. KEY FINDINGS: Pharmacokinetic studies indicated that the oral bioavailability of AMF was 0.06 ± 0.04%, and the area under the curve of the glucuronidated AMF metabolites (410.938 ± 62.219 ng/ml h) was significantly higher than that of AMF (194.509 ± 16.915 ng/ml h). UGT1A1 and UGT1A3 greatly metabolized AMF. No significant difference was observed in the antioxidant activity between AMF and its metabolites. The anticancer activity of AMF metabolites significantly decreased. CONCLUSIONS: A low AMF bioavailability was due to extensive glucuronidation, which was mediated by UGT1A1 and UGT1A3. Glucuronidated AMF metabolites had the same antioxidant but had a lower anticancer activity than that of AMF.

Sodium aescinate significantly suppress postoperative peritoneal adhesion by inhibiting the RhoA/ROCK signaling pathway.

BACKGROUND: Although mechanical barriers and modern surgical techniques have been developed to prevent postoperative adhesion formation, high incidence of adhesions still represents an important challenge in abdominal surgery. So far, there has been no available therapeutic drug in clinical practice. PURPOSE: In this study, we explored the efficacy of sodium aescinate (AESS) treatment against postoperative peritoneal adhesions, the potential molecular mechanism was also investigated. STUDY DESIGN AND METHODS: Sixty male Sprague-Dawley rats were randomly divided into 6 groups for the study: the blank, vehicle, positive control and three AESS administration groups (0.5, 1 and 2 mg/kg/d, intravenous administration for 7 days). Adhesions were induced by discretely ligating peritoneal sidewall. An IL-1ß-induced HMrSV5 cell model was also performed to explore possible functional mechanism. RESULTS: The results indicated that the incidence and severity of peritoneal adhesions were significantly lower in the AESS-treated groups than that in the vehicle and positive control group. AESS-treated groups showed that the secretion, activity, and expression of tPA in rat peritoneum were notably increased. The FIB levels in rat plasma were decreased. The immunohistochemical staining analysis demonstrated that collagen I and α-SMA deposition were significantly attenuated in AESS-treated peritoneal tissues. Besides, we found that AESS treatment reduced the protein levels of p-MYPT1. To further explore the mechanisms of AESS, both activator and inhibitors of RhoA/ROCK pathway were employed in this study. It was found that AESS-induced up-regulation of tPA was reversed by activator of ROCK, but the effects of ROCK inhibitors were consistent with AESS. CONCLUSION: Taken together, the findings of in vivo and in vitro experiments proved that AESS could significantly suppress postoperative peritoneal adhesion formation through inhibiting the RhoA/ROCK signaling pathway. Our researches provide important pharmacological basis for AESS development as a potential therapeutic agent on peritoneal adhesions.

Extracellular IL-37 promotes osteogenic differentiation of human bone marrow mesenchymal stem cells via activation of the PI3K/AKT signaling pathway.

Interleukin (IL)-37, a pivotal anti-inflammatory cytokine and a fundamental inhibitor of innate immunity, has recently been shown to be abnormally expressed in several autoimmune-related orthopedic diseases, including rheumatoid arthritis, ankylosing spondylitis, and osteoporosis. However, the role of IL-37 during osteogenic differentiation of mesenchymal stem cells (MSCs) remains largely unknown. In this study, extracellular IL-37 significantly increased osteoblast-specific gene expression, the number of mineral deposits, and alkaline phosphatase activity of MSCs. Moreover, a signaling pathway was activated in the presence of IL-37. The enhanced osteogenic differentiation of MSCs due to supplementation of IL-37 was partially rescued by the presence of a PI3K/AKT signaling inhibitor. Using a rat calvarial bone defect model, IL-37 significantly improved bone healing. Collectively, these findings indicate that extracellular IL-37 enhanced osteogenesis of MSCs, at least in part by activation of the PI3K/AKT signaling pathway.

Transplantation of fecal microbiota rich in short chain fatty acids and butyric acid treat cerebral ischemic stroke by regulating gut microbiota.

The gut microbiota and its short chain fatty acid (SCFA) metabolites have been established to play an important protective role against neurodegenerative diseases. Our previous study demonstrated that cerebral ischemic stroke triggers dysfunctional gut microbiota and increased intestinal permeability. In this study, we aimed to clarify the mechanism by which gut microbiota and SCFAs can treat cerebral ischemic stroke in rat middle cerebral artery occlusion models and use the information to develop new therapies. Our results show that oral administration of non-absorbable antibiotics reduced neurological impairment and the cerebral infarct volume, relieved cerebral edemas, and decreased blood lipid levels by altering the gut microbiota. We also found that ischemic stroke decreased intestinal levels of SCFAs. And that transplanting fecal microbiota rich in these metabolites was an effective means of treating the condition. Compared with other SCFAs, butyric acid showed the highest negative correlation with ischemic stroke. Supplementation with butyric acid treated models of ischemic stroke effectively by remodeling the gut microbiota, enriching the beneficial Lactobacillus, and repairing the leaky gut. In conclusion, interfering with the gut microbiota by transplanting fecal bacteria rich in SCFAs and supplementing with butyric acid were found to be effective treatments for cerebral ischemic stroke.

CYP3A4/5 mediates the metabolic detoxification of humantenmine, a highly toxic alkaloid from Gelsemium elegans Benth.

Gelsemium elegans Benth., a well-known toxic herbal plant, is widely used to treat rheumatic arthritis, inflammation and other diseases. Gelsemium contains humantenmine (HMT), which is an important bioactive and toxic alkaloid. Cytochrome P450 enzymes (CYPs) play important roles in the elimination and detoxification of exogenous substances. This study aimed to investigate the roles of CYPs in the metabolism and detoxification of HMT. First, metabolic studies were performed in vitro by using human liver microsomes, selective chemical inhibitors and recombinant human CYPs. Results indicated that four metabolites, including hydroxylation and oxidation metabolites, were found in human liver microsomes and identified based on their high-resolution mass spectrum. The isoform responsible for HMT metabolism was mainly CYP3A4/5. Second, the toxicity of HMT on L02 cells in the presence of the nicotinamide adenine dinucleotide phosphate system (NADPH) was significantly less than that without NADPH system. A CYP3A4/5 activity inhibition model was established by intraperitoneally injecting ketoconazole in mice and used to evaluate the role of CYP3A4/5 in HMT detoxification. In this model, the 14-day survival rate of the mice decreased to 17% after they were intragastrically treated with HMT, along with hepatic injury and increasing alanine aminotransferase (ALT) /aspartate aminotransferase (AST) levels. Overall, CYP3A4/5 mediated the metabolism and detoxification of HMT.

In vitro activity and clinical efficacy of macrolides, cefoperazone-sulbactam and piperacillin/piperacillin-tazobactam against Bordetella pertussis and the clinical manifestations in pertussis patients due to these isolates: A single-centre study in Zhejiang Province, China.

BACKGROUND AND OBJECTIVE: Macrolides are the recommended antibiotics for treating pertussis and preventing transmission. The causative bacterium, Bordetella pertussis, has high macrolide resistance and has recently circulated in China. The objective of this study was to find effective alternative antibiotics for treatment by assessing the in vitro activity and clinical efficacy of antibiotics against Bordetella pertussis. METHODS: Bordetella pertussis was confirmed by agglutination with specific antisera and mass spectrometry. The MICs of antibiotics against isolates were determined using the Etest method. Treatment outcomes were clinically and microbiologically evaluated. RESULTS: A total of 126 pertussis patients were diagnosed based on culture, 69.8% of whom were aged ≤6 months and 72.1% were treated with previous macrolides. Leucocytosis and lymphocytosis were observed in 29.4% and 54.8% of all patients, respectively. Both MIC50 and MIC90 of erythromycin, azithromycin, and clindamycin were >256mg/L, and 75.4% were highly macrolide resistant. The MIC90 of trimethoprim-sulfamethoxazole, ampicillin, ampicillin-sulbactam, cefuroxime, ceftriaxone and cefoperazone-sulbactam were 0.38mg/L, 0.25mg/L, 0.19mg/L, 12mg/L, 0.19mg/L and 0.047mg/L, respectively. The MICs of piperacillin in all of the isolations were <0.016mg/L. Of the patients treated with single cefoperazone-sulbactam or piperacillin-tazobactam, 30 of 32 (93.8%) had significantly improved clinical symptoms and 24 of 25 (96%) had negative culture results after 2 weeks of therapy. CONCLUSION: Macrolide resistance in Bordetella pertussis is a serious problem in Zhejiang Province, China. Piperacillin/piperacillin-tazobactam and cefoperazone-sulbactam have potent antibacterial activity in vitro and in vivo, and may become the alternative choice for treating pertussis caused by macrolide-resistant isolates.

Puerariae Lobatae Radix with chuanxiong Rhizoma for treatment of cerebral ischemic stroke by remodeling gut microbiota to regulate the brain-gut barriers.

The combination of Puerariae Lobatae Radix (PLR) and Chuanxiong Rhizoma (CXR) is commonly used to treat cerebrovascular diseases. This work aimed to clarify the mechanisms of their action in treating cerebral ischemic stroke from the perspective of gut microecology. The PLR and CXR combination effectively improved the neurological function, reduced the cerebral infarction and relieved the complications of cerebral ischemic stroke, including dyslipidemia, increased blood viscosity and thrombotic risk. Cerebral ischemic stroke triggered gut microbial disturbances by enriching pathogens and opportunistic microorganisms, including Bacteroides, Escherichia_Shigella, Haemophilus, Eubacterium_nodatum_group, Collinsella, Enterococcus, Proteus, Alistipes, Klebsiella, Shuttleworthia and Faecalibacterium. Cerebral ischemic stroke also increased the intestinal permeability, disrupted the gut barrier and caused intestinal microbial translocation. Occludin, claudin-5 and ZO-1 levels in the brain-gut barriers showed a high positive correlation. However, the combination remodeled the gut microecology by modulating endogenous bacteria whose effects may mitigate cerebral damage, such as Alloprevotella, Ruminococcaceae, Oscillospira, Lachnospiraceae_NK4B4_group, Akkermansia and Megasphaera, protected the brain-gut barriers by increasing claudin-5 and ZO-1 levels; and weakened the gut microbiota translocation by decreasing diamine oxidase, lipopolysaccharide and d-lactate. Although nimodipine effectively reduced the cerebral infarction, it did not relieve the gut microbiota dysbiosis and instead aggravated the gut barrier disruption and microbiota translocation. In conclusion, cerebral ischemic stroke caused gut microbiota dysbiosis, increased intestinal permeability, disrupted the gut barrier and triggered gut microbiota translocation. The PLR and CXR combination was an effective treatment for cerebral ischemic stroke that relieved the gut microbiota dysbiosis and brain-gut barriers disruption.

Asperuloside and Asperulosidic Acid Exert an Anti-Inflammatory Effect via Suppression of the NF-κB and MAPK Signaling Pathways in LPS-Induced RAW 264.7 Macrophages.

Hedyotis diffusa is a folk herb that is used for treating inflammation-related diseases in Asia. Previous studies have found that iridoids in H. diffusa play an important role in its anti-inflammatory activity. This study aimed to investigate the anti-inflammatory effect and potential mechanism of five iridoids (asperuloside (ASP), asperulosidic acid (ASPA), desacetyl asperulosidic acid (DAA), scandoside methyl ester (SME), and E-6-O-p-coumaroyl scandoside methyl ester (CSME)) that are presented in H. diffusa using lipopolysaccharide (LPS)-induced RAW 264.7 cells. ASP and ASPA significantly decreased the production of nitric oxide (NO), prostaglandin E2 (PGE2), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) in parallel with the inhibition of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), TNF-α, and IL-6 mRNA expression in LPS-induced RAW 264.7 cells. ASP treatment suppressed the phosphorylation of the inhibitors of nuclear factor-kappaB alpha (IκB-α), p38, extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase (JNK). The inhibitory effect of ASPA was similar to that of ASP, except for p38 phosphorylation. In summary, the anti-inflammatory effects of ASP and ASPA are related to the inhibition of inflammatory cytokines and mediators via suppression of the NF-κB and mitogen-activated protein kinase (MAPK) signaling pathways, which provides scientific evidence for the potential application of H. diffusa.