[Structural Chinese medicine: new research field on pharmacodynamic substance basis of traditional Chinese medicine].

The research on the pharmacodynamic substance basis of traditional Chinese medicine(TCM) is a key scientific issue for the inheritance and development of TCM. At present, a large number of remarkable achievements have been made in the field of chemical components in Chinese medicine, however, another important aspect, namely the physical structure and mode of action of the multi-component assembly of TCM, has not been clearly understood and deeply studied. From the bottleneck of restricting material ba-sic research, we objectively analyzed the common cause of the existing problems. Based on the new discoveries and advances of active substances from TCM emerging in recent years, we extracted and summarized the concept of structural Chinese medicine, elaborated the basic ideas, main features and research modes, hoping to provide theoretical and practical references for the study on the pharmacodynamic substance basis and other research fields of TCM.

Xanthatin inhibits STAT3 and NF-κB signalling by covalently binding to JAK and IKK kinases.

Aberrant activation of the signal transducer and activator of transcription 3 (STAT3) and the nuclear factor-κB (NF-κB) signalling pathways is associated with the development of cancer and inflammatory diseases. JAKs and IKKs are the key regulators in the STAT3 and NF-κB signalling respectively. Therefore, the two families of kinases have been the major targets for developing drugs to regulate the two signalling pathways. Here, we report a natural compound xanthatin from the traditional Chinese medicinal herb Xanthium L. as a potent inhibitor of both STAT3 and NF-κB signalling pathways. Our data demonstrated that xanthatin was a covalent inhibitor and its activities depended on its α-methylene-γ-butyrolactone group. It preferentially interacted with the Cys243 of JAK2 and the Cys412 and Cys464 of IKKß to inactivate their activities. In doing so, xanthatin preferentially inhibited the growth of cancer cell lines that have constitutively activated STAT3 and p65. These data suggest that xanthatin may be a promising anticancer and anti-inflammation drug candidate.

Elaboration in type, primary structure, and bioactivity of polysaccharides derived from mollusks.

Over the past decades, numerous Mollusca species have received more attention in development and utilization as valuable bio-resources. Many efforts have been focused on investigating mollusk polysaccharides because of their rich content, ease of extraction, diversified sorts, specific structure, various biofunctions and potent activities. To date, many mollusks, especially species of gastropods, bivalves, or cephalopods, have been reported containing polysaccharide compounds in tissues with abundant amount, and most of polysaccharides are obtainable through combining techniques of extraction, separation and purification. The polysaccharides isolated from mollusks appeared with various structural and physicochemical characteristics, ranged from neutral polysaccharides and sulfated polysaccharides, to GAGs series (including Hep/HS, CS/DS, HA and similarities), even to heterogeneous glycan with high molecular weight. This review article provides comprehensive knowledge of recent researches on type classification, tissue origins and possible biofunctions of various polysaccharides from mollusks. The highlights were placed in structure variation including molecular weight, sulfation pattern, linkages and monomer compositions for repeating unit, and primary molecular construction of the mollusks polysaccharides. In addition, this article covers general information on exhibition of mollusks polysaccharide extracts or preparations in the various bioactivities, such as anticoagulant, antiatherogenic, antioxidant, immunomodulatory, antivirus and antitumor activities, which would reveal their possible potentials in medical application. Furthermore, the article presents a brief overview on several challenges and future scope in this field.

Small molecule IVQ, as a prodrug of gluconeogenesis inhibitor QVO, efficiently ameliorates glucose homeostasis in type 2 diabetic mice.

Gluconeogenesis is a major source of hyperglycemia in patients with type 2 diabetes mellitus (T2DM), thus targeting gluconeogenesis to suppress glucose production is a promising strategy for anti-T2DM drug discovery. In our preliminary in vitro studies, we found that a small-molecule (E)-3-(2-(quinoline-4-yl)vinyl)-1H-indol-6-ol (QVO) inhibited the hepatic glucose production (HGP) in primary hepatocytes. We further revealed that QVO suppressed hepatic gluconeogenesis involving calmodulin-dependent protein kinase kinase ß- and liver kinase B1-adenosine monophosphate-activated protein kinase (AMPK) pathways as well as AMPK-independent mitochondrial function-related signaling pathway. To evaluate QVO's anti-T2DM activity in vivo, which was impeded by the complicated synthesis route of QVO with a low yield, we designed and synthesized 4-[2-(1H-indol-3-yl)vinyl]quinoline (IVQ) as a prodrug with easier synthesis route and higher yield. IVQ did not inhibit the HGP in primary hepatocytes in vitro. Pharmacokinetic studies demonstrated that IVQ was quickly converted to QVO in mice and rats following administration. In both db/db and ob/ob mice, oral administration of IVQ hydrochloride (IVQ-HCl) (23 and 46 mg/kg every day, for 5 weeks) ameliorated hyperglycemia, and suppressed hepatic gluconeogenesis and activated AMPK signaling pathway in the liver tissues. Furthermore, IVQ caused neither cardiovascular system dysfunction nor genotoxicity. The good druggability of IVQ has highlighted its potential in the treatment of T2DM and the prodrug design for anti-T2DM drug development.

Tumor-targeting efficacy of a BF211 prodrug through hydrolysis by fibroblast activation protein-α.

BF211, a bufalin (BF) derivative, exhibits stronger anti-cancer activity than BF but with potential cardiotoxicity. Fibroblast activation protein-α (FAPα) is a membrane-bound protease specifically expressed by carcinoma-associated fibroblasts, thus has been used for the selective delivery of anticancer agents. In this study, we used a FAPα-based prodrug strategy to synthesize a dipeptide (Z-Gly-Pro)-conjugated BF211 prodrug named BF211-03. BF211-03 was hydrolyzed by recombinant human FAPα (rhFAPα) and cleaved by homogenates of human colon cancer HCT-116 or human gastric cancer MGC-803 xenografts. In contrast, BF211-03 showed good stability in plasma and in the homogenates of FAPα-negative normal tissues, such as heart and kidney. In HCT-116 and MGC-803 cells with low levels of FAPα expression, BF211-03 displayed a lower in vitro cytotoxicity than BF211 with approximately 30 to 40-fold larger IC50 values, whereas in human breast cancer MDA-MB-435 cells with high levels of FAPα expression, the IC50 value difference between BF211-03 and BF211 was small (approximately 4-fold). Although the cytotoxicity of BF211-03 against tumor cells was dramatically decreased by the chemical decoration, it was restored after cleavage of BF211-03 by rhFAPα or tumor homogenate. In HCT-116 tumor-bearing nude mice, doubling the dose of BF211-03, compared with BF211, caused less weight loss, but showing similar inhibitive effects on tumor growth. Our results suggest that BF211-03 is converted to active BF211 in tumor tissues and exhibits anti-tumor activities in tumor-bearing nude mice. FAPα-targeted BF211-03 displays tumor selectivity and may be useful as a targeting agent to improve the safety profile of cytotoxic natural products for use in cancer therapy.

LX2343 alleviates cognitive impairments in AD model rats by inhibiting oxidative stress-induced neuronal apoptosis and tauopathy.

Alzheimer's disease (AD) is a progressive neurodegenerative disease leading to the irreversible loss of brain neurons and cognitive abilities, and the vicious interplay between oxidative stress (OS) and tauopathy is believed to be one of the major players in AD development. Here, we demonstrated the capability of the small molecule N-(1,3-benzodioxol-5-yl)-2-[5-chloro-2-methoxy(phenylsulfonyl)anilino]acetamide (LX2343) to ameliorate the cognitive dysfunction of AD model rats by inhibiting OS-induced neuronal apoptosis and tauopathy. Streptozotocin (STZ) was used to induce OS in neuronal cells in vitro and in AD model rats that were made by intracerebroventricular injection of STZ (3 mg/kg, bilaterally), and Morris water maze test was used to evaluate the cognitive dysfunction in ICV-STZ rats. Treatment with LX2343 (5-20 µmol/L) significantly attenuated STZ-induced apoptosis in SH-SY5Y cells and mouse primary cortical neurons by alleviating OS and inhibiting the JNK/p38 and pro-apoptotic pathways. LX2343 was able to restore the integrity of mitochondrial function and morphology, increase ATP biosynthesis, and reduce ROS accumulation in the neuronal cells. In addition, LX2343 was found to be a non-ATP competitive GSK-3ß inhibitor with IC50 of 1.84±0.07 µmol/L, and it potently inhibited tau hyperphosphorylation in the neuronal cells. In ICV-STZ rats, administration of LX2343 (7, 21 mg·kg-1·d-1, ip, for 5 weeks) efficiently improved their cognitive deficits. LX2343 ameliorates the cognitive dysfunction in the AD model rats by suppressing OS-induced neuronal apoptosis and tauopathy, thus highlighting the potential of LX2343 for the treatment of AD.

Thamnolia vermicularis extract improves learning ability in APP/PS1 transgenic mice by ameliorating both Aß and Tau pathologies.

Considering the complicated pathogenesis of Alzheimer's disease (AD), multi-targets have become a focus in the discovery of drugs for treatment of this disease. In the current work, we established a multi-target strategy for discovering active reagents capable of suppressing both Aß level and Tau hyperphosphorylation from natural products, and found that the ethanol extract of Thamnolia vermicularis (THA) was able to improve learning ability in APP/PS1 transgenic mice by inhibiting both Aß levels and Tau hyperphosphorylation. SH-SY5Y and CHO-APP/BACE1 cells and primary astrocytes were used in cell-based assays. APP/PS1 transgenic mice [B6C3-Tg(APPswe, PS1dE9)] were administered THA (300 mg·kg-1·d-1, ig) for 100 d. After the administration was completed, the learning ability of the mice was detected using a Morris water maze (MWM) assay; immunofluorescence staining, Congo red staining and Thioflavine S staining were used to detect the senile plaques in the brains of the mice. ELISA was used to evaluate Aß and sAPPß contents, and Western blotting and RT-PCR were used to investigate the relevant signaling pathway regulation in response to THA treatment. In SH-SY5Y cells, THΑ (1, 10, 20 µg/mL) significantly stimulated PI3K/AKT/mTOR and AMPK/raptor/mTOR signaling-mediated autophagy in the promotion of Aß clearance as both a PI3K inhibitor and an AMPK indirect activator, and restrained Aß production as a suppressor against PERK/eIF2α-mediated BACE1 expression. Additionally, THA functioned as a GSK3ß inhibitor with an IC50 of 1.32±0.85 µg/mL, repressing Tau hyperphosphorylation. Similar effects on Aß accumulation and Tau hyperphosphorylation were observed in APP/PS1 transgenic mice treated with THA. Furthermore, administration of THA effectively improved the learning ability of APP/PS1 transgenic mice, and markedly reduced the number of senile plaques in their hippocampus and cortex. The results highlight the potential of the natural product THA for the treatment of AD.

Possible target-related proteins and signal network of bufalin in A549 cells suggested by both iTRAQ-based and label-free proteomic analysis.

Bufalin (BF) exhibited antiproliferation and antimigration effects on human A549 lung cancer cells. To search its target-related proteins, protein expression profiles of BF-treated and control cells were compared using two quantitative proteomic methods, iTRAQ-based and label-free proteomic analysis. A total of 5428 proteins were identified in iTRAQ-based analysis while 6632 proteins were identified in label-free analysis. The number of common identified proteins of both methods was 4799 proteins. By application of 1.20-fold for upregulated and 0.83-fold for downregulated cutoff values, 273 and 802 differentially expressed proteins were found in iTRAQ-based and label-free analysis, respectively. The number of common differentially expressed proteins of both methods was 45 proteins. Results of bioinformational analysis using Metacore(TM) showed that the two proteomic methods were complementary and both suggested the involvement of oxidative stress and regulation of gene expression in the effects of BF, and fibronectin-related pathway was suggested to be an important pathway affected by BF. Western blotting assay results confirmed BF-induced change in levels of fibronectin and other related proteins. Overexpression of fibronectin by plasmid transfection ameliorated antimigration effects of BF. Results of the present study provided information about possible target-related proteins and signal network of BF.

Structural Basis for Small Molecule NDB (N-Benzyl-N-(3-(tert-butyl)-4-hydroxyphenyl)-2,6-dichloro-4-(dimethylamino) Benzamide) as a Selective Antagonist of Farnesoid X Receptor α (FXRα) in Stabilizing the Homodimerization of the Receptor.

Farnesoid X receptor α (FXRα) as a bile acid sensor plays potent roles in multiple metabolic processes, and its antagonist has recently revealed special interests in the treatment of metabolic disorders, although the underlying mechanisms still remain unclear. Here, we identified that the small molecule N-benzyl-N-(3-(tert-butyl)-4-hydroxyphenyl)-2,6-dichloro-4-(dimethylamino) benzamide (NDB) functioned as a selective antagonist of human FXRα (hFXRα), and the crystal structure of hFXRα ligand binding domain (hFXRα-LBD) in complex with NDB was analyzed. It was unexpectedly discovered that NDB induced rearrangements of helix 11 (H11) and helix 12 (H12, AF-2) by forming a homodimer of hFXRα-LBD, totally different from the active conformation in monomer state, and the binding details were further supported by the mutation analysis. Moreover, functional studies demonstrated that NDB effectively antagonized the GW4064-stimulated FXR/RXR interaction and FXRα target gene expression in primary mouse hepatocytes, including the small heterodimer partner (SHP) and bile-salt export pump (BSEP); meanwhile, administration of NDB to db/db mice efficiently decreased the gene expressions of phosphoenolpyruvate carboxykinase (PEPCK), glucose 6-phosphatase (G6-pase), small heterodimer partner, and BSEP. It is expected that our first analyzed crystal structure of hFXRα-LBD·NDB will help expound the antagonistic mechanism of the receptor, and NDB may find its potential as a lead compound in anti-diabetes research.

Small molecule LX2343 ameliorates cognitive deficits in AD model mice by targeting both amyloid ß production and clearance.

AIM: Streptozotocin (STZ) is widely used to induce oxidative damage and to impair glucose metabolism, apoptosis, and tau/Aß pathology, eventually leading to cognitive deficits in both in vitro and in vivo models of Alzheimer's disease (AD). In this study, we constructed a cell-based platform using STZ to induce stress conditions mimicking the complicated pathologies of AD in vitro, and evaluated the anti-amyloid effects of a small molecule, N-(1,3-benzodioxol-5-yl)-2-[5-chloro-2-methoxy(phenylsulfonyl)anilino]acetamide (LX2343) in the amelioration of cognitive deficits in AD model mice. METHODS: Cell-based assays for screening anti-amyloid compounds were established by assessing Aß accumulation in HEK293-APPsw and CHO-APP cells, and Aß clearance in primary astrocytes and SH-SY5Y cells after the cells were treated with STZ in the presence of the test compounds. Autophagic flux was observed using confocal laser scanning microscopy. APP/PS1 transgenic mice were administered LX2343 (10 mg·kg-1·d-1, ip) for 100 d. After LX2343 administration, cognitive ability of the mice was evaluated using Morris water maze test, and senile plaques in the brains were detected using Thioflavine S staining. ELISA assay was used to evaluate Aß and sAPPß levels, while Western blot analysis was used to measure the signaling proteins in both cell and animal brains. RESULTS: LX2343 (5-20 µmol/L) dose-dependently decreased Aß accumulation in HEK293-APPsw and CHO-APP cells, and promoted Aß clearance in SH-SY5Y cells and primary astrocytes. The anti-amyloid effects of LX2343 were attributed to suppressing JNK-mediated APPThr668 phosphorylation, thus inhibiting APP cleavage on one hand, and inhibiting BACE1 enzymatic activity with an IC50 value of 11.43±0.36 µmol/L, on the other hand. Furthermore, LX2343 acted as a non-ATP competitive PI3K inhibitor to negatively regulate AKT/mTOR signaling, thus promoting autophagy, and increasing Aß clearance. Administration of LX2343 in APP/PS1 transgenic mice significantly ameliorated cognitive deficits and markedly ameliorated the Aß pathology in their brains. CONCLUSION: LX2343 ameliorates cognitive dysfunction in APP/PS1 transgenic mice via both Aß production inhibition and clearance promotion, which highlights the potential of LX2343 in the treatment of AD.

Bufalin derivative BF211 inhibits proteasome activity in human lung cancer cells in vitro by inhibiting ß1 subunit expression and disrupting proteasome assembly.

AIM: Bufalin is one of the active components in the traditional Chinese medicine ChanSu that is used to treat arrhythmia, inflammation and cancer. BF211 is a bufalin derivative with stronger cytotoxic activity in cancer cells. The aim of this study was to identify the putative target proteins of BF211 and the signaling pathways in cancer cells. METHODS: A549 human lung cancer cells were treated with BF211. A SILAC-based proteomic analysis was used to detect the protein expression profiles of BF211-treated A549 cells. Cellular proteasome activities were examined using fluorogenic peptide substrates, and the binding affinities of BF211 to recombinant proteasome subunit proteins were evaluated using the Biacore assay. The expression levels of proteasome subunits were determined using RT-PCR and Western blotting, and the levels of the integral 26S proteasome were evaluated using native PAGE analysis. RESULTS: The proteomic analysis revealed that 1282 proteins were differentially expressed in BF211-treated A549 cells, and the putative target proteins of BF211 were associated with various cellular functions, including transcription, translation, mRNA splicing, ribosomal protein synthesis and proteasome function. In A549 cells, BF211 (5, 10, and 20 nmol/L) dose-dependently inhibited the enzymatic activities of proteasome. But BF211 displayed a moderate affinity in binding to proteasome ß1 subunit and no binding affinity to the ß2 and ß5 subunits. Moreover, BF211 (0.1, 1, and 10 nmol/L) did not inhibit the proteasome activities in the cell lysates. BF211 (5, 10, and 20 nmol/L) significantly decreased the expression level of proteasome ß1 subunit and the levels of integral 26S proteasome in A549 cells. Similarly, knockdown of the ß1 subunit with siRNA in A549 cells significantly decreased integral 26S proteasome and proteasome activity. CONCLUSION: BF211 inhibits proteasome activity in A549 cells by decreasing ß1 subunit expression and disrupting proteasome assembly.

A cascaded QSAR model for efficient prediction of overall power conversion efficiency of all-organic dye-sensitized solar cells.

A cascaded model is proposed to establish the quantitative structure-activity relationship (QSAR) between the overall power conversion efficiency (PCE) and quantum chemical molecular descriptors of all-organic dye sensitizers. The cascaded model is a two-level network in which the outputs of the first level (JSC, VOC, and FF) are the inputs of the second level, and the ultimate end-point is the overall PCE of dye-sensitized solar cells (DSSCs). The model combines quantum chemical methods and machine learning methods, further including quantum chemical calculations, data division, feature selection, regression, and validation steps. To improve the efficiency of the model and reduce the redundancy and noise of the molecular descriptors, six feature selection methods (multiple linear regression, genetic algorithms, mean impact value, forward selection, backward elimination, and +n-m algorithm) are used with the support vector machine. The best established cascaded model predicts the PCE values of DSSCs with a MAE of 0.57 (%), which is about 10% of the mean value PCE (5.62%). The validation parameters according to the OECD principles are R(2) (0.75), Q(2) (0.77), and Qcv2 (0.76), which demonstrate the great goodness-of-fit, predictivity, and robustness of the model. Additionally, the applicability domain of the cascaded QSAR model is defined for further application. This study demonstrates that the established cascaded model is able to effectively predict the PCE for organic dye sensitizers with very low cost and relatively high accuracy, providing a useful tool for the design of dye sensitizers with high PCE.

Physalin B not only inhibits the ubiquitin-proteasome pathway but also induces incomplete autophagic response in human colon cancer cells in vitro.

AIM: To investigate the effects of physalin B insolated from Physalis divericata on human colon cancer cells in vitro and its anticancer mechanisms. METHODS: Human HCT116 colon cancer cell line was tested. Cell viability and apoptosis were detected, and relevant proteins were measured using Western blot analyses. Autophagosomes were observed in stable GFP-LC3 HCT116 cells. Localization of autophagosomes and lysosomes was evaluated in GFP-LC3/RFP-LAMP1-co-transfected cells. Microtubules and F-actin microfilaments were observed with confocal microscope. Mitochondrial ROS (mito-ROS) was detected with flow cytometry in the cells stained with MitoSox dye. RESULTS: Physalin B inhibited the viability of HCT116 cells with an IC50 value of 1.35 µmol/L. Treatment of the cells with physalin B (2.5-10 µmol/L) induced apoptosis and the cleavage of PARP and caspase-3. Meanwhile, physalin B treatment induced autophagosome formation, and accumulation of LC3-II and p62, but decreased Beclin 1 protein level. Marked changes of microtubules and F-actin microfilaments were observed in physalin B-treated cells, which led to the blockage of co-localization of autophagosomes and lysosomes. Physalin B treatment dose-dependently increased the phosphorylation of p38, ERK and JNK in the cells, whereas the p38 inhibitor SB202190, ERK inhibitor U0126 or JNK inhibitor SP600125 could partially reduce physalin B-induced PARP cleavage and p62 accumulation. Moreover, physalin B treatment dose-dependently increased mito-ROS production in the cells, whereas the ROS scavenger NAC could reverse physalin B-induced effects, including incomplete autophagic response, accumulation of ubiquitinated proteins, changes of microtubules and F-actin, activation of p38, ERK and JNK, as well as cell death and apoptosis. CONCLUSION: Physalin B induces mito-ROS, which not only inhibits the ubiquitin-proteasome pathway but also induces incomplete autophagic response in HCT116 cells in vitro.

Bigelovin inhibits STAT3 signaling by inactivating JAK2 and induces apoptosis in human cancer cells.

AIM: To study the function and mechanism of bigelovin, a sesquiterpene lactone from the flower of Chinese herb Inula hupehensis, in regulating JAK2/STAT3 signaling and cancer cell growth. METHODS: HepG2 cells stably transfected with the STAT3-responsive firefly luciferase reporter plasmid (HepG2/STAT3 cells), and a panel of human cancer cell lines were used to identify active compounds. Cell viability was measured using MTT assay. Western blotting was used to detect protein expression and phosphorylation. Kinase assays were performed and the reaction between bigelovin and thiol-containing compounds was analyzed with LC-MS. RESULTS: Bigelovin (1-50 µmol/L) dose-dependently inhibited the IL-6-induced STAT3 activation in HepG2/STAT3 cells (IC50=3.37 µmol/L) and the constitutive STAT3 activation in A549 and MDA-MB-468 cells. Furthermore, bigelovin dose-dependently inhibited JAK2 phosphorylation in HeLa and MDA-MB-468 cells, as well as the enzymatic activity of JAK2 in vitro (IC50=44.24 µmol/L). Pretreatment of the cells with DTT (500 µmol/L) or GSH (500 µmol/L) eliminated the inhibitory effects of bigelovin on the IL-6-induced and the constitutive STAT3 activation. The results in LC-MS analysis suggested that bigelovin might react with cysteine residues of JAK2 leading to inactivation of JAK2. Bigelovin (5 and 20 µmol/L) had no effects on the signaling pathways of growth factors EGF, PDGF or insulin. Finally, bigelovin suppressed the cell viability and induced apoptosis in 10 different human cancer cell lines, particularly those with constitutively activated STAT3. CONCLUSION: Bigelovin potently inhibits STAT3 signaling by inactivating JAK2, and induces apoptosis of a variety of human cancer cells in vitro.

Methylation and its role in the disposition of tanshinol, a cardiovascular carboxylic catechol from Salvia miltiorrhiza roots (Danshen).

AIM: Tanshinol is an important catechol in the antianginal herb Salvia miltiorrhiza roots (Danshen). This study aimed to characterize tanshinol methylation. METHODS: Metabolites of tanshinol were analyzed by liquid chromatography/mass spectrometry. Metabolism was assessed in vitro with rat and human enzymes. The major metabolites were synthesized for studying their interactions with drug metabolizing enzymes and transporters and their vasodilatory properties. Dose-related tanshinol methylation and its influences on tanshinol pharmacokinetics were also studied in rats. RESULTS: Methylation, preferentially in the 3-hydroxyl group, was the major metabolic pathway of tanshinol. In rats, tanshinol also underwent considerable 3-O-sulfation, which appeared to be poor in human liver. These metabolites were mainly eliminated via renal excretion, which involved tubular secretion mainly by organic anion transporter (OAT) 1. The methylated metabolites had no vasodilatory activity. Entacapone-impaired methylation did not considerably increase systemic exposure to tanshinol in rats. The saturation of tanshinol methylation in rat liver could be predicted from the Michaelis constant of tanshinol for catechol-O-methyltransferase (COMT). Tanshinol had low affinity for human COMT and OATs; its methylated metabolites also had low affinity for the transporters. Tanshinol and its major human metabolite (3-O-methyltanshinol) exhibited negligible inhibitory activities against human cytochrome P450 enzymes, organic anion transporting polypeptides 1B1/1B3, multidrug resistance protein 1, multidrug resistance-associated protein 2, and breast cancer resistance protein. CONCLUSION: Tanshinol is mainly metabolized via methylation. Tanshinol and its major human metabolite have low potential for pharmacokinetic interactions with synthetic antianginal agents. This study will help define the risk of hyperhomocysteinemia related to tanshinol methylation.

Discovery of a novel inhibitor of NAD(P)(+)-dependent malic enzyme (ME2) by high-throughput screening.

AIM: Malic enzymes are oxidative decarboxylases with NAD(+) or NAD(P)(+) as cofactor that catalyze the conversion of L-malate to pyruvate and CO2. The aim of this study was to discover and characterize a potent inhibitor of human NAD(P)(+)-dependent malic enzyme 2 (ME2). METHODS: Recombinant human ME2-His-Tag fusion protein was overexpressed in E coli and purified with Ni-NTA resin. A high-throughput screening (HTS) assay was developed to find ME2 inhibitors. Detergent Brij-35 was used to exclude false positives. The characteristics of the inhibitor were analyzed with enzyme kinetics analysis. A thermal shift assay for ME2 was carried out to verify the binding of the inhibitor with the enzyme. RESULTS: An HTS system for discovering ME2 inhibitors was established with a Z' factor value of 0.775 and a signal-to-noise ratio (S/N) of 9.80. A library containing 12 683 natural products was screened. From 47 hits, NPD387 was identified as an inhibitor of ME2. The primary structure-activity relationship study on NPD387 derivatives showed that one derivative NPD389 was more potent than the parent compound NPD387 (the IC50 of NPD389 was 4.63 ± 0.36 µmol/L or 5.59 ± 0.38 µmol/L, respectively, in the absence or presence of 0.01% Brij-35 in the assay system). The enzyme kinetics analysis showed that NPD389 was a fast-binding uncompetitive inhibitor with respect to the substrate NAD(+) and a mixed-type inhibitor with respect to the substrate L-malate. CONCLUSION: NPD389 is a potent ME2 inhibitor that binds to the enzyme in a fast-binding mode, acting as an uncompetitive inhibitor with respect to the substrate NAD(+) and a mixed-type inhibitor with respect to the substrate L-malate.

Arctigenin enhances swimming endurance of sedentary rats partially by regulation of antioxidant pathways.

AIM: Arctigenin, a phenylpropanoid dibenzylbutyrolactone lignan found in traditional Chinese herbs, has been determined to exhibit a variety of pharmacological activities, including anti-tumor, anti-inflammation, neuroprotection, and endurance enhancement. In the present study, we investigated the antioxidation and anti-fatigue effects of arctigenin in rats. METHODS: Rat L6 skeletal muscle cell line was exposed to H2O2 (700 µmol/L), and ROS level was assayed using DCFH-DA as a probe. Male SD rats were injected with arctigenin (15 mg·kg(-1)·d(-1), ip) for 6 weeks, and then the weight-loaded forced swimming test (WFST) was performed to evaluate their endurance. The levels of antioxidant-related genes in L6 cells and the skeletal muscles of rats were analyzed using real-time RT-PCR and Western blotting. RESULTS: Incubation of L6 cells with arctigenin (1, 5, 20 µmol/L) dose-dependently decreased the H2O2-induced ROS production. WFST results demonstrated that chronic administration of arctigenin significantly enhanced the endurance of rats. Furthermore, molecular biology studies on L6 cells and skeletal muscles of the rats showed that arctigenin effectively increased the expression of the antioxidant-related genes, including superoxide dismutase (SOD), glutathione reductase (Gsr), glutathione peroxidase (GPX1), thioredoxin (Txn) and uncoupling protein 2 (UCP2), through regulation of two potential antioxidant pathways: AMPK/PGC-1α/PPARα in mitochondria and AMPK/p53/Nrf2 in the cell nucleus. CONCLUSION: Arctigenin efficiently enhances rat swimming endurance by elevation of the antioxidant capacity of the skeletal muscles, which has thereby highlighted the potential of this natural product as an antioxidant in the treatment of fatigue and related diseases.

Comparison of Thoracoscopy-Guided Thoracic Paravertebral Block and Ultrasound-Guided Thoracic Paravertebral Block in Postoperative Analgesia of Thoracoscopic Lung Cancer Radical Surgery: A Randomized Controlled Trial.

INTRODUCTION: Ultrasound-guided thoracic paravertebral block (UTPB) is widely used for postoperative analgesia in thoracic surgery. However, it has many disadvantages. Thoracoscopy-guided thoracic paravertebral block (TTPB) is a new technique for thoracic paravertebral block (TPB). In this study, we compared the use of TTPB and UTPB for pain management after thoracoscopic radical surgery for lung cancer. METHODS: In total, 80 patients were randomly divided 1:1 into the UTPB group and the TTPB group. The surgical time of TPB, the success rate of the first puncture, block segment range, visual analog scale (VAS) scores at 2, 6, 12, 24, and 48 h post operation, and the incidence of postoperative adverse reactions were compared between the two groups. RESULTS: The surgical time of TPB was significantly shorter in the TTPB group than in the UTPB group (2.2 ± 0.3 vs. 5.7 ± 1.7 min, t = - 12.411, P < 0.001). The success rate of the first puncture and the sensory block segment were significantly higher in the TTPB group than in the UTPB group (100% vs. 76.9%, χ2 = 8.309, P < 0.001; 6.5 ± 1.2 vs. 5.1 ± 1.3 levels, t = - 5.306, P < 0.001, respectively). The VAS scores were significantly higher during rest and coughing at 48 h post operation than at 2, 6, 12, and 24 h post operation in the TTPB group. The VAS scores were significantly lower during rest and coughing at 12 and 24 h post operation in the TTPB group than in the UTPB group (rest: 2.5 ± 0.4 vs. 3.4 ± 0.6, t = 7.325, P < 0.001; 2.5 ± 0.5 vs. 3.5 ± 0.6, t = 7.885, P < 0.001; coughing: 3.4 ± 0.6 vs. 4.2 ± 0.7, t = 5.057, P < 0.001; 3.4 ± 0.6 vs. 4.2 ± 0.8, t = 4.625, P < 0.001, respectively). No significant difference was observed in terms of postoperative adverse reactions between the two groups. CONCLUSIONS: Compared with UTPB, TTPB shows advantages, such as simpler and more convenient surgery, shorter surgical time, a higher success rate of the first puncture, wider block segments, and superior analgesic effect. TTPB can effectively reduce postoperative pain due to thoracoscopic lung cancer radical surgery. TRIAL REGISTRATION: https://www.chictr.org.cn , identifier ChiCTR2300072005, prospectively registered on 31/05/2023.

Causal associations between the gut microbiota and multiple myeloma: a two-sample Mendelian randomization study.

Background: Previous observational studies have indicated a potential association between the gut microbiota and multiple myeloma (MM). However, the relationship between the gut microbiota and MM remains unclear. This study aimed to ascertain the existence of a causal link between the gut microbiota and MM. Methods: To investigate the potential causal relationship between gut microbiota and MM, a two-sample Mendelian randomization (MR) analysis was conducted. Exposure data was obtained from the MiBioGen consortium, which provided genetic variants associated with 211 bacterial traits. MM outcome data was obtained from the FinnGen consortium. The selection of Single nucleotide polymorphisms estimates was performed through meta-analysis using inverse-variance weighting, and sensitivity analyses were conducted using weighted median, MR Egger, Simple mode, and MR-PRESSO. Results: The results of the study demonstrated a significant positive correlation between the genus Eubacterium ruminantium group and the risk of MM (OR 1.71, 95% CI 1.21 to 2.39). Conversely, the genus: Dorea (OR 0.46, 95% CI 0.24 to 0.86), Coprococcus1 (OR 0.47, 95% CI 0.22 to 1.00), RuminococcaceaeUCG014 (OR 0.57, 95% CI 0.33 to 0.99), Eubacterium rectale group (OR 0.37, 95% CI 0.18 to 0.77), and order: Victivallales (OR 0.62, 95% CI 0.41-0.94), class: Lentisphaeria (OR 0.62, 95% CI 0.41 to 0.94), exhibited a negative association with MM. The inverse variance weighting analysis provided additional support for these findings. Conclusion: This study represents an inaugural exploration of MR to investigate the connections between gut microbiota and MM, thereby suggesting potential significance for the prevention and treatment of MM.