Electrochemical Cross-Coupling between N-(4-Hydroxyphenyl)-sulfonamides and 2-Naphthols: Synthesis of 2,2'-Bis(arenol)s.

We herein present an electrochemical method for the dehydrogenative cross-coupling of N-(4-hydroxyphenyl)-sulfonamides and 2-naphthols. This transformation provides a direct and scalable approach to a wide range of C1-symmetric 2,2'-bis(arenol)s with moderate to high yields under mild conditions. Preliminary attempts with the asymmetric variant of this reaction were also performed with ≤55% ee for the synthesis of 2,2'-bis(arenol)s. Control experiments were conducted to propose a plausible mechanism for the reaction.

Drug-Herb Synergistic Interactions between Clopidogrel and Natural Medicine.

INTRODUCTION: Natural medicine (NM) has been used since ancient times for therapeutic purposes worldwide. Presently, the combination of clopidogrel and NM with a reasonable synergistic effect has gained increasing acceptance in clinical therapeutics. METHODS: Here, we have performed a comprehensive retrieval of literature published in both English and Chinese databases until August 1, 2022, studying the synergistic interactions of clopidogrel and NM through pharmacokinetic/pharmacodynamic (PK-PD) analyses. We retrieved 7, 3, and 5 studies on PK analysis and 3, 3, and 8 studies on PD analysis for the interaction of clopidogrel with single herbal medicines, bioactive compounds, and herbal prescriptions, respectively. Most studies on NM have been found to mainly focus on preclinical observations, and there have been fewer clinical PK analyses. RESULTS: A potential drug-herb interaction has been observed to occur when clopidogrel and NM were metabolized by an enzyme network comprising P-gp, CES1, and CYP450. In contrast, most PD studies have focused on clinical observations, and few preclinical findings have been reported. Some cases have suggested that the combination of the two types of drugs would alter the antiplatelet efficacy and adverse effects. Studies on PK, however, have shown significant or slightly varying results for the drug prototype and its metabolites. CONCLUSION: In the combination therapies, the interaction between clopidogrel and NM was found to alter antiplatelet aggregation pathways and P2Y12 receptor function.

Design, synthesis and anti-tumor efficacy evaluation of novel 1,3-diaryl propane-based polyphenols obtained from Claisen rearrangement reaction.

Polyphenols such as resveratrol, honokiol and nordihydroguaiaretic acid are widely existing in nature products or synthetic compounds with interesting biological activities. Inspired by their structural feature, a total of 49 1,3-diaryl propane-based polyphenols were designed and synthesized through Claisen rearrangement reaction. New compounds were initially assessed for their anti-proliferative activities against various cancer cell lines (PC-3, U87MG, U251, HCT116) at a concentration of 50 µM, and the results guided the SAR of this series of compounds. Further screening of selected compounds against seven cancer cell lines (three additional colon cancer cell lines namely COLO205, HT29 and SW480 were chosen) led to the identification of two advanced leads 2t and 3t with IC50 values ranging from 8.2 ± 0.1 to 19.3 ± 1.9 µM. Both compounds also showed promising anti-proliferative activities against COLO205 in dose- and time-dependent manners. Furthermore, 2t and 3t exhibited good anti-tumor efficacy in COLO205 xenografted mice model with TGI values ranging from 38% to 58%. These results warrant the further investigation of this series of compounds.

Lead Drug Discover Strategies from Natural Medicines Based on Network Pharmacology.

The need for therapeutics to overcome development of existing diseases research to discover new lead agents. In the face of public health challenges worldwide, natural medicines play a pivotal role in innovative lead drug discovery. Network pharmacology can easily construct complicated poly-pharmacology network based on lead compound, biological function, and bioactive target proteins, which meets the overall feature of natural medicines, and enable to elucidate the action mechanism at molecule-protein level with systematic view. In this work, we first summarized the recent progress delineating lead drug development and its interaction with natural medicines. Second, we focused on the relationship between natural medicines and network pharmacology. Additionally, we discussed current issues and potential prospects for the lead drug discover from natural medicines by network pharmacology. Further investigations should be focus on relevant structural analysis for biological experiment, also the dynamic and quantitative network development. In summary, it is a rational approach for innovative lead drug discovery, and with the development of structure and biology research, this approach makes it a very powerful method for the lead molecules in a high-throughput manner from a comprehensive and powerful special multi-compound to target protein/disease poly pharmacology network.

Simultaneous determination of phenols in the four main original plants of the famous traditional Chinese medicine Shihu by pressurized capillary electrochromatography.

A rapid pressurized capillary electrochromatography (pCEC) method has been established for the simultaneous analysis of 11 phenols in the four main original plants of the famous traditional Chinese medicine (TCM) Shihu. The effects of wavelength, mobile phase, flow rate, pH value, concentration of buffer, and applied voltage were systematically studied. The investigated 11 phenols could be isolated in 35 min on a reversed-phase EP-100-20/45-3-C18 capillary column using the established method. To apply the established pCEC method, all phenols except tristin (11) were detected in the four Dendrobium plants. A total of 10 components were detected in D. huoshanense, 6 components in D. nobile, 3 components in D. chrysotoxum, and 4 components in D. fimbriatum. The consistent evaluation revealed that the similarities among the four original plants of Shihu were 38.2-86.0% based on the 11 polyphenols and 92.5-97.7% based on the pCEC fingerprints. These further suggested that the components of the four original plants of TCM Shihu might be significantly different. Further investigation should be conducted to confirm and evaluate if the four species could be used as the same medicine with the same amount according to Chinese Pharmacopoeia (ChP).

Synthesis and Discovery of Ligustrazine-Heterocycle Derivatives as Antitumor Agents.

Ligustrazine (TMP) is a natural pyrazine alkaloid extracted from the roots of Ligusticum Chuanxiong Hort, which has the potential as an antitumor agent. A series of 33 ligustrazine-heterocycle (TMPH) derivatives were designed, synthesized, and investigated via antitumor screening assays, molecular docking analysis, and prediction of drug-like properties. TMP was attached to other heterocyclic derivatives by an 8-12 methylene alkyl chain as a linker to obtain 33 TMPH derivatives. The structures were confirmed by 1H-NMR, 13C-NMR, and high-resolution mass spectroscopy spectral (HR-MS) data. The antiproliferative activity against human breast cancer MCF-7, MDA-MB-231, mouse breast cancer 4T1, mouse fibroblast L929, and human umbilical vein endothelial HUVEC cell lines was evaluated by MTT assay. Compound 12-9 displayed significant inhibitory activity with IC50 values in the low micromolar range (0.84 ± 0.02 µM against the MDA-MB-231 cell line). The antitumor effects of compound 12-9 were further evaluated by plate cloning, Hoechst 33 342 staining, and annexin V-FITC/PI staining. The results indicated that compound 12-9 inhibited the proliferation and apoptosis of breast cancer cells. Furthermore, molecular docking of compound 12-9 into the active site of the Bcl-2, CASP-3, and PSMB5 target proteins was performed to explore the probable binding mode. The 33 newly synthesized compounds were predicted to have good drug-like properties in a theoretical study. Overall, these results indicated that compound 12-9 inhibited cell proliferation through PSMB5 and apoptosis through Bcl-2/CASP-3 apoptotic signaling pathways and had good drug-like properties. These results provided more information, and key precursor lead derivatives, in the search for effective bioactive components from Chinese natural medicines.

ACSL4 promotes colorectal cancer and is a potential therapeutic target of emodin.

BACKGROUND: Colorectal cancer (CRC) is an important death-related disease in the world and new therapeutic strategies are urgently needed to reduce mortality. Several studies have demonstrated that emodin, the main ingredient of Rheum palmatum, fights cancer but its potential anti-tumor effect on CRC is still unknown. PURPOSE: The present study is aimed to explore the potential anti-tumor effects of emodin against CRC and the underlying molecular mechanism. METHODS: CRC-related datasets were screened according to filter criteria in the GEO database and TCGA database. By using screened differentially expressed genes, GO, KEGG and survival analysis were carried out. The expressions of ACSL4, VEGFR1, and VEGFR2 were examined by immunohistochemistry and western blot. Then, pcDNA-ACSL4, pcDNA-VEGFR1, and pcDNA-VEGFR2 were used to overexpress ACSL4, VEGFR1, and VEGFR2, while ACSL4 siRNA was used to silence ACSL4 expression in HCT116 cells. CCK-8 assay and transwell migration assay were used to detect the cell proliferation and invasion. A docking simulation assay and an MST assay were performed to explore the potential mode of emodin binding to ACSL4. The HCT116 cells and CRC mouse model were established to investigate the effects of emodin on CRC. RESULTS: The ACSL4, VEGFR1, and VEGFR2 expression were upregulated in CRC tissues and ACSL4 was associated with a shorter survival time in CRC patients. ACSL4 downregulation reduced cell proliferation and invasion, while ACSL4 exhibited a positive correlation with the levels of VEGFR1, VEGFR2, and VEGF. In HCT116 cells, emodin reduced cell proliferation and invasion by inhibiting ACSL4, VEGFR1, and VEGFR2 expression and VEGF secretion. Docking simulation and MST assay confirmed that emodin can directly bind to ACSL4 target. Moreover, ACSL4 overexpression abolished the inhibitory effect of emodin on VEGF secretion and VEGFR1 and VEGFR2 expression, but VEGFR1 and VEGFR2 overexpression did not affect the inhibitory effect of emodin on ACSL4 expression and VEGF secretion. Furthermore, emodin reduced the mortality and tumorigenesis of CRC mice and reduced ACSL4, VEGFR1, VEGFR2 expression, and VEGF content. CONCLUSION: Our findings indicate that emodin inhibits proliferation and invasion of CRC cells and reduces VEGF secretion and VEGFR1 and VEGFR2 expression by inhibiting ACSL4. This emodin-induced pathway offers insights into the molecular mechanism of its antitumor effect and provides a potential therapeutic strategy for CRC.

Quantitative Determination of Quercitrin Levels in Rat Plasma Using UHPLC-MS/MS and its Application in a Pharmacokinetic Study after the Oral Administration of Polygoni cuspidati Folium Capsules.

BACKGROUND: Quercitrin is widely found in herbal medicines, and it is particularly important in the design of new therapeutic agents. Because of its wide range of biological activities, methods for detecting quercitrin and its pharmacokinetics in biological samples must be investigated. OBJECTIVES: To develop and validate a sensitive and reliable ultra-high-performance liquid chromatography- tandem mass spectrometry (UHPLC-MS/MS) method for the quantitative determination of quercitrin levels in rat plasma, and test its application in a pharmacokinetic investigation after the oral administration of Polygoni cuspidati folium capsules (HC). METHODS: First, a rapid analytical method implementing UHPLC-MS/MS for the quantification of quercitrin levels in rat plasma was developed and validated. The analyte and internal standard (IS) tinidazole were extracted from rat plasma via protein precipitation with 800 µL of methanol and 50 µL of 1% formic acid solution. Chromatographic separation was performed using an Agilent ZORBAX C18 column within 4 min. Mass spectrometry was performed for quantification using a triple-quadrupole mass spectrometer employing electrospray ionization in the negative ion mode. The MRM transitions for quercitrin and IS were m/z 447.2→229.9 and m/z 246.0→125.8, respectively. The UHPLC-MS/MS method for the quantitative determination of quercitrin levels in rat plasma was then applied to investigate its pharmacokinetics after the oral administration of HC in rats. RESULTS: The developed UHPLC-MS/MS method for detecting quercitrin in rat plasma was linear over the range of 0.1-160 ng/mL. The linear regression equation was Y = (0.7373 ± 0.0023)X - (0.0087 ± 0.0021) (r2 = 0.9978). The intra- and interday precision values were within 7.8%, and the recoveries of quercitrin and IS exceeding 67.3%. The UHPLC-MS/MS method was successfully applied to characterize the pharmacokinetic profile of quercitrin in eight rats after the oral administration of HC. The experimentally obtained values were fit to a one-compartment, first-order pharmacokinetic model, and they appeared to fit the concentration-time curve. CONCLUSION: Quercitrin was proven to be stable during sample storage, preparation, and the analytical procedures. The pharmacokinetic parameters suggested that quercitrin may be present in the peripheral tissues of rats.

A Network Pharmacology Approach for Exploring the Mechanisms of Panax notoginseng Saponins in Ischaemic Stroke.

BACKGROUND: Panax notoginseng saponins (PNS) have been deemed effective herb compounds for treating ischaemic stroke (IS) and improving the quality of life of IS patients. This study aimed to investigate the underlying mechanisms of PNS in the treatment of IS based on network pharmacology. METHODS: PNS were identified from the Traditional Chinese Medicine System Pharmacology (TCMSP) database, and their possible targets were predicted using the PharmMapper database. IS-related targets were identified from the GeneCards database, OMIM database, and DisGeNET database. A herb-compound-target-disease network was constructed using Cytoscape, and protein-protein interaction (PPI) networks were established with STRING. GO enrichment and KEGG pathway analysis were performed using DAVID. The binding of the compounds and key targets was validated by molecular docking studies using AutoDock Vina. The neuroprotective effect of TFCJ was substantiated in terms of oxidative stress (superoxide dismutase, glutathione peroxidase, catalase, and malondialdehyde) and the levels of IGF1/PI3K/Akt pathway proteins. RESULTS: A total of 375 PNS targets and 5111 IS-related targets were identified. Among these targets, 241 were common to PNS, and IS network analysis showed that MAPK1, AKT1, PIK3R1, SRC, MAPK8, EGFR, IGF1, HRAS, RHOA, and HSP90AA1 are key targets of PNS against IS. Furthermore, GO and KEGG enrichment analysis indicated that PNS probably exert therapeutic effects against IS by regulating many pathways, such as the Ras, oestrogen, FoxO, prolactin, Rap1, PI3K-Akt, insulin, PPAR, and thyroid hormone signalling pathways. Molecular docking studies further corroborated the experimental results.The network pharmacology results were further verified by molecular docking and in vivo experiments. CONCLUSIONS: The ameliorative effects of PNS against IS were predicted to be associated with the regulation of the IGF1-PI3K-Akt signalling pathway. Ginsenoside Re and ginsenoside Rb1 may play an important role in the treatment of IS.

Iodide-promoted transformations of imidazopyridines into sulfur-bridged imidazopyridines or 1,2,4-thiadiazoles.

A NaI-promoted sequential double carbon-sulfur bond formation was developed to afford sulfur-bridged imidazopyridines, using Deoxofluor as the sulfur source and requiring only 15 min at room temperature. Using this process, imidazo[1,5-a]pyridines could also be transformed to 1,2,4-thiadiazoles in the presence of ammonium salt with the formation of both carbon-sulfur and nitrogen-sulfur bonds. This mechanistically unique method is distinguished by its wide substrate scope, lack of requirement for transition metals and mild conditions.

Unraveling the Action Mechanism of Buyang Huanwu Tang (BYHWT) for Cerebral Ischemia by Systematic Pharmacological Methodology.

BACKGROUND: Buyang Huanwu Tang (BYHWT) and relevant Traditional Chinese medicine (TCM) has its unique advantages in the treatment of cerebral ischemia. However, its pharmacological mechanism has not been fully explained. OBJECTIVE: Base on the multi-component, also the entire disease network targets, the present study sets out to identify major bioactive ingredients, key disease targets, and pathways of BYHWT against cerebral ischemia disease by systematic pharmacological methodology. METHODS: Both the bioactive compounds from the BYHWT and the positive drugs against cerebral ischemia were fully investigated. The binding targets of the positive drugs were then obtained. A virtual screening protocol was then used to highlight the compound-target interaction and network was constructed to visualize the compound-target binding effect after docking analysis. Moreover, the targets enrichment analysis for biological processes and pathways were performed to further explore the function of bio-targets protein gene and its role in the signal pathway. RESULTS: A total of 382 active ingredients of the BYHWT and 23 candidate disease targets were identified. Virtual screening results indicated that multiple bioactive compounds targeted multiple proteins. Each compound acts on one or more targets. The mechanisms were linked to 20 signaling pathways, and the key mechanism was related to serotonergic synapse, calcium signaling pathway and camp signaling pathways. CONCLUSION: The present study explored the bioactive ingredients and mechanisms of BYHWT against cerebral ischemia by systematic pharmacological methodology. The novel methodology would provide a reference for the lead discovery of precursors, disease mechanism and material base for TCM.

Catechins from oolong tea improve uterine defects by inhibiting STAT3 signaling in polycystic ovary syndrome mice.

BACKGROUND: It is showed that inflammation is causative factor for PCOS, leading to a decline in ovarian fertility. Previous studies have reported that tea consumption can reduce the incidence of ovarian cancer. We speculate that catechins from oolong tea (Camellia sinensis (L.) O. Kuntze) may have a potential therapeutic effect on PCOS. This study aims to investigate the effects of oolong tea catechins on the uterus of polycystic ovary syndrome (PCOS) mice induced by insulin combined with human chorionic gonadotropin (hCG). METHODS: Sixty female mice were divided into 6 groups (n = 10): model, model + Metformin 200 mg/kg, model + catechins 25 mg/kg, model + catechins 50 mg/kg, and model + catechins 100 mg/kg. Another forty female mice were divided into 4 groups (n = 10): control, control + catechins 100 mg/kg, model, and model + catechins 100 mg/kg. Ovarian and uterine weight coefficients, sex hormone levels, glucose metabolism and insulin resistance, and ovarian and uterine pathology were examined. Changes in NF-κB-mediated inflammation, MMP2 and MMP9 expressions, and STAT3 signaling were evaluated in the uterus of mice. RESULTS: Catechins could effectively reduce the ovarian and uterine organ coefficients, reduce the levels of E2, FSH and LH in the blood and the ratio of LH/FSH, and improve glucose metabolism and insulin resistance in PCOS mice induced by insulin combined with hCG. In addition, catechins could significantly down-regulated the expression of p-NF-κB p65 in the uterus and the protein expressions of the pro-inflammatory factors (IL-1ß, IL-6, and TNF-α). The expressions of mmp2 and mmp9 associated with matrix degradation in uterine tissue were also significantly down-regulated by catechins. Further, catechins significantly reduced the expression of p-STAT3 and increased the expression of p-IRS1 and p-PI3K in the uterus of PCOS mice. CONCLUSION: Catechins from oolong tea can alleviate ovarian dysfunction and insulin resistance in PCOS mice by inhibiting uterine inflammation and matrix degradation via inhibiting p-STAT3 signaling.

[A systematic pharmacological investigation of pharmacologically active ingredients in Toujie Quwen granules for treatment of COVID-19].

OBJECTIVE: To explore the pharmacologically active ingredients in Toujie Quwen granules (TJQW) for treatment of coronavirus disease 2019 (COVID-19) in light of systemic pharmacology. METHODS: We performed database search, literature mining and drug-like index screening to identify the bioactive components in TJQW, the positive drugs for disease treatment and their therapeutic targets. The core disease target was investigated based on the cross-linking interaction of the bioactive components, positive drug and potential disease target, and the target proteins at the key nodes were analyzed by GO and KEGG analyses. Based on the therapeutic targets for COVID-19, virtual screening was conducted to screen the compounds in TJQW and construct the network cross-linking the key bioactive molecules in TJQW, key node targets of the disease, and the related biological pathways. RESULTS: We identified 159 compounds in TJQW and obtained 18 core proteins based on the cross-linking of the bioactive components, positive drugs and disease targets. The key node targets consisted of 22 targets including the latest 4 COVID-19 proteins. Virtual screening results showed that at least 14 compounds could bind with the core disease target proteins. The material basis of TJQW for COVID-19 treatment was explained in multi-pathway, multi-component and multi-target perspectives. In terms of the structural characteristics of the compounds, we screened the top 30 molecules with strong binding with the target proteins, among which flavonoids were the predominant components. CONCLUSIONS: This investigation reveals the therapeutic mechanism of TJQW for COVID-19 involving multiple components, targets and pathways from the perspective of key bioactive molecules, disease key node targets and related biological pathways. We screened 30 active precursors from TJQW, which provides reference for the clinical application and further development of TJQW.

Emodin Inhibits the Proliferation of MCF-7 Human Breast Cancer Cells Through Activation of Aryl Hydrocarbon Receptor (AhR).

Natural products have proved to be a promising source for the development of potential anticancer drugs. Emodin, a natural compound from Rheum palmatum, is used to treat several types of cancers, including lung, liver, and pancreatic. However, there are few reports regarding its use in the treatment of breast cancer. Thus, the therapeutic effect and mechanism of emodin on MCF-7 human breast cancer cells were investigated in this study. Morphological observations and cell viability were evaluated to determine the anti-proliferation activity of emodin. Network pharmacology and molecular docking were performed to screen the potential targets. Western blot analysis was used to explore a potential antitumor mechanism. The results showed that emodin (50-100 µmol/L) could significantly inhibit the proliferation of MCF-7 cells in a time and dose-dependent manner. Furthermore, virtual screening studies indicated that emodin was a potent aryl hydrocarbon receptor (AhR) agonist in chemotherapy for breast cancer. Finally, when MCF-7 cells were treated with emodin (100 µmol/L) for 24 h, the AhR and cytochrome P450 1A1 (CYP1A1) protein expression levels were significantly upregulated compared with the control group. Our study indicated that emodin exhibited promising antitumor activity in MCF-7 cells, likely through activation of the AhR-CYP1A1 signaling pathway. These findings lay a foundation for the application of emodin in breast cancer treatment.

MicroRNA 27b promotes cardiac fibrosis by targeting the FBW7/Snail pathway.

Our study aspires to understand the impact of miR-27b on myocardial fibrosis as well as its functional mechanism. 12 days post the ligation of coronary artery in rats, the expression of miR-27b in the peri-infarction region was elevated. Treating cultivated rat neonatal cardiac fibroblasts (CFs) with angiotensin II (AngII) also enhanced the miR-27b expression. Forced expression of miR-27b promoted the proliferation and collagen production in rat neonatal CFs, as revealed by cell counting, MTT assay, and quantitative reverse transcription-polymerase chain reaction. FBW7 was found to be the miR-27b's target since the overexpression of miR-27b reduced the transcriptional level of FBW7. The enhanced expression of FBW7 protein abrogated the effects of miR-27b in cultured CFs, while the siRNA silence of FBW7 promoted the pro-fibrosis activity of AngII. As to the mechanism, we found that the expression of FBW7 led to the degradation of Snail, which is an important regulator of cardiac epithelial-mesenchymal transitions. Importantly, inhibition of miR-27b abrogated the coronary artery ligation (CAL) induced cardiac fibrosis in vivo, suggesting that it might be a potential target for the treatment of fibrosis associated cardiac diseases.

[Mechanism of Gardenia jasminoides against cholestasis based on network pharmacology].

To screen the active ingredients of Gardenia jasminoides and potential targets,and investigate the mechanisms against cholestasis based on network pharmacology technology. Twenty-one active components of G. jasminoides were retrieved and the target sites were screened by using Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform( TCMSP). Cytoscape3. 2. 1 was used to construct the component-target network. Two hundred and eight targets related to cholestasis were searched and screened through Dis Ge NET,KEGG and OMIM databases. The key targets of G. jasminoides components and cholestasis were integrated and screened,and the component-target-disease network was constructed with Cytoscape 3. 2. 1 software to screen out the core network whose freedom degree was greater than the average value. The Clue GO plug-in of Cytoscape 3. 2. 1 software was used to analyze the biological processes and pathway enrichment of G. jasminoides in regulation of cholestasis. GO biological process analysis revealed 17 biological processes,involving 3 signaling biological processes related to cholestasis,i.e. acute inflammatory response,positive regulation of reactive oxygen species metabolic process,and nitric oxide biosynthetic process. KEGG-KEEG-305 terms and REACTOME pathways analysis revealed 17 regulatory pathways,involving 4 signaling pathways related to cholestasis,i.e. metabolism of xenobiotics by cytochrome P450,nuclear receptor transcription pathway,GPVI-mediated activation cascade and platelet activation. It was found that aqueous extract of G. jasminoides could improve serum biochemical abnormalities in ANIT-induced cholestasis rats. Aqueous extract of G. jasminoides could decrease the protein and mRNA expression levels of ESR1 in liver tissues,and increase the protein and mRNA expression levels of PPARG,NOS2,F2 R,NOS3,and NR3 C1. To sum up,the possible mechanisms of G. jasminoides against cholestasis may be related with the above three processes and four pathways.

MiR-9 promotes the phenotypic switch of vascular smooth muscle cells by targeting KLF5

Background/aim: Diabetic vascular smooth muscle cells (VSMCs) are characterized by increased proliferation and migration. Small noncoding microRNAs (miRNAs) have been considered critical modulators of the VSMC phenotypic switch after an environmental stimulus. However, microRNA in high glucose-induced proinflammation and its atherogenic effect is still ambiguous. Materials and methods: The technique of qRT-PCR was used to examine the expression of miR-9 in VSMCs. The downstream signaling protein relative to miR-9 regulation, Krüppel-like factor 5, and some marker genes of contractile VSMCs were analyzed by western blotting and qRT-PCR. Luciferase reporter assay was used to detect the expression of KLF5, which is regulated by miR-9. To examine the function of a miR-9 inhibitor in VSMC proliferation and migration, VSMC proliferation and migration assays were performed. Results: Reduced transcriptional levels of miR-9 and expression of specific genes of contractile VSMCs were observed in the SMC cell line C-12511 treated with high glucose and SMCs, which were isolated from db/db mice. Moreover, the activity of KLF5 3'-UTR was dramatically reduced by a miR-9 mimic and increased by a miR-9 inhibitor. The proliferation and migration of SMCs were reduced by the miR-9 mimic. Conclusion: miR-9 inhibits the proliferation and migration of SMC by targeting KLF5 in db/db mice.

[Effect of geniposidic acid on hepato-enteric circulation in cholestasis rats through Sirt1-FXR signaling pathway].

To investigate the effects of geniposidic acid( GPA) on hepato-enteric circulation in cholestasis rats,and to explore the mechanism based on the sirtuin 1( Sirt1)-farnesol X receptor( FXR) pathway,sixty SD rats were randomly divided into 6 groups:blank control group,ANIT model group,ursodeoxycholic acid group( 100 mg·kg~(-1)·d-1 UDCA),and GPA high,medium and low( 100,50 and 25 mg·kg~(-1)·d-1) dosage groups,10 rats in each group. Corresponding drugs were intragastrically( ig) administered for10 days. After administration on day 8,all rats except blank rats were administered with 65 mg·kg~(-1)α-naphthalene isothiocyanate( ANIT) once. After the last administration,the serum levels of alanine aminotransferase( ALT),glutamine oxalacetate aminotransferase( AST),gamma-glutamyltransferase( γ-GGT),alkaline phosphatase( ALP),total bilirubin( TB) and total bile acid( TBA)were measured,and the mRNA transcription levels of Sirt1,FXR,multidrug resistant associated protein 2( MRP2),bile salt export pump( BSEP),sodium taurocholate contractible polypeptide( NTCP) in liver and apical sodium bile acid transporter( ASBT),ileum bile acid binding protein( IBABP) in ileum were detected by reverse transcription-polymerase chain reaction( RT-PCR). The protein expression levels of Sirt1,FXR and NTCP were detected by Western blot; the expression of MRP2,BSEP in liver and ASBT,IBABP in ileum were determined by immunofluorescence three staining. Primary rat hepatocytes were cultured in vitro to investigate the inhibitory effect of GPA on a potent and selective Sirt1 inhibitor( EX 527),and the mRNA and protein expression levels of Sirt1 and FXR were detected by RT-PCR and Western blot. GPA significantly decreased the levels of ALT,AST,γ-GGT,ALP,TB,TBA in serum( P<0.01) and improved the pathological damage of liver tissues in ANIT-induced cholestasis rats; significantly increased the mRNA and protein expression levels of Sirt1,FXR,MRP2,BSEP,NTCP in liver and ASBT,IBABP in ileum( P< 0.01). In vitro primary hepatocytes experiment indicated that the gene and protein expression levels of FXR and Sirt1 were noticeably improved by GPA in primary hepatocytes inhibited by EX-527( P<0.01). It was found that the improvement of GPA was in a dose-dependent manner. GPA could improve bile acid hepatointestinal circulation and play a liver protection and cholagogu role in cholestasis rats induced by ANIT.The mechanism may be that GPA activated FXR by regulating Sirt1,a key regulator of oxidative stress injury,and then the activated FXR could regulate protein of bile acid hepato-enteric circulation.

Distinctive Roles of Sirtuins on Diabetes, Protective or Detrimental?

Dysregulation of metabolic pathways leads to type 2 diabetes, characteristic of high glucose concentration caused by insulin resistance. The histone deacetylases sirtuins exhibit remarkable enzymatic activities. Accumulating evidence indicates that sirtuins can be pharmacologically activated to ameliorate diabetes. Here, we evaluated different roles of sirtuins (SIRT1-SIRT7) in diabetes progression and described their involvement in metabolic pathways of skeletal muscle, adipose tissue and liver. The nuclear sirtuins, SIRT1, SIRT6, and SIRT7, regulate the activity of key transcription factors and cofactors in almost all tissues with the cellular responses to energy demands. The mitochondrial sirtuins, SIRT3, SIRT4, and SIRT5, regulate the activity of mitochondrial enzymes in response to fasting and calorie restriction. Moreover, genetic polymorphisms of SIRT1 and SIRT2 have been reported to associate with diabetes development. It's worth noting that SIRT1, SIRT2, SIRT3, and SIRT6 are positive regulators of insulin resistance in most cases. In the opposite, SIRT4 and SIRT7 inhibit insulin secretion and fatty acid oxidation. Identification of SIRT1 activators for diabetes has gained wide attention, such as metformin, resveratrol, and resveratrol derivatives. Randomized, prospective, and large-scale clinical trials are warrant to uncover the responsibilities of SIRTs modulators on diabetes progress.

The Herb-Drug Interaction of Clopidogrel and Xuesaitong Dispersible Tablet by Modulation of the Pharmacodynamics and Liver Carboxylesterase 1A Metabolism.

OBJECTIVE: Clopidogrel and Xuesaitong dispersible tablet (XST) have been clinically proven to be effective for treating cardiocerebrovascular disease. The present study was to investigate the herb-drug interaction of Clopidogrel and XST by modulation of the pharmacodynamics and liver Carboxylesterase 1A(CES1A) metabolism. METHODS: 30 male SD rats were randomly divided into a control group (equal volumes of saline, 6 rats for mRNA analysis), a clopidogrel group (clopidogrel with dose 30 mg/kg), and a combination group (clopidogrel and XST, with dose 30 and 50 mg/kg respectively, each group continuous administration once daily for 30 days). The clopidogrel and combination group comprised 12 rats, with 6 designated for mRNA analysis and 6 for the pharmacokinetic study. The 2-bromo-3'-methoxyacetophenone- (MPB-) derivatized clopidogrel active thiol metabolite (CAMD) was measured by UHPLC-MS/MS for pharmacokinetics (n=6). The expression of CES1A mRNA was examined with real-time RT-PCR (n=6). Molecular simulation was used to investigate the inhibition effect of XST on the CES1A protein. The CAMD pharmacodynamics and CES1A metabolism were investigated to evaluated the herb-drug interaction. RESULTS: Clopidogrel and XST coadministration appreciably increased the Cmax, AUC, and MRT of CAMD. However, the expression of CES1A mRNA was decreased accordingly. It also indicated that the bioactive components in XST had good interaction with the CES1A metabolism target by molecular simulation. The animal study indicated that clopidogrel and XST coadministration produced significant herb-drug interactions at active CAMD pharmacokinetic and CES1A metabolic enzyme aspect. CONCLUSION: 30-days dose of coadministration altered hepatic CES1A protein and resulted in reduced plasma levels of active CAMD. both the decreased CES1A mRNA expression and the inhibition on the protein were due to the combination of XST, which accordingly upregulated the pharmacokinetics of plasma active CAMD.