Tartary buckwheat root polysaccharides ameliorate non-alcoholic fatty liver disease via the IL6-SOCS3-SREBP1c pathway.

Our previous study demonstrated that Tartary buckwheat root polysaccharides (TBRP) could reduce insulin resistance in diabetes mellitus by inhibiting SOCS3-stimulated IRS1 protein degradation. However, whether TBRP has the efficiency to treat non-alcoholic fatty liver disease (NAFLD) is still undetermined. This investigation aimed to examine the effects of TBRP on a high-fat diet (HFD)-triggered NAFLD, and elucidate the underlying molecular mechanisms. Briefly, TBRP toxicity in hepatoma (BEL7404) and pancreatic cancer (BxPC3) cells and zebrafish embryos developmental models, were evaluated in-vitro and in-vivo, respectively. TBRP inhibited cellular lipid accumulation by suppressing fat synthesis, furthermore, it improved body weight gain, liver weight, liver-to-body weight ratio, serum lipids triglyceride, total cholesterol, ALT, LDL-C, HDL-C, and AST levels in the NAFLD mice model. Additionally, TBRP treatment also lowered the nitric oxide content. The qPCR assay revealed that mRNA expression of TNF, IL1ß, and IL6 was also markedly reduced in TBRP-treated NAFLD mice. The expression of SOCS3, SREBP1c, and STAT3 was elucidated by western blot analysis, which indicated that TBRP markedly decreased the gene expression for de novo fat synthesis by the SOCS3-SREBP1c pathway. These findings reveal that TBRP ameliorates NAFLD via the IL6-SOCS3-SREBP1c signaling pathway and therefore, may represent a promising approach for NAFLD treatment.

The prevention effect of probiotics against eczema in children: an update systematic review and meta-analysis.

Accumulated evidences support the fetus's intestinal flora unbalance is associated with the development of allergic diseases. Probiotic supplements in pregnancy and childhood might prevent atopic diseases. The aim of this systematic review and meta-analysis was to evaluate the effect of probiotic supplementation during pregnancy and early infancy in preventing eczema, atopic eczema, and other allergic diseases. We also explored whether different probiotic strains or intervention objects affected the antiallergic effect of probiotics and the prevention atopy effect of the long-term period. Fixed-effect models were used, and random-effects models where significant heterogeneity was present. Results were expressed as odds ratios (ORs) with a 95% confidence interval (CI). Twenty-one studies were included in the meta-analysis. The probiotics group had a significantly lower risk of eczema and atopic eczema compared to controls, especially those treated with probiotic combinations. Mothers' probiotics intake significantly contributed to reducing the risk of eczema as well as atopic eczema. What's more, probiotics seemed effective on eczema prevention ≤2 years of age, but against atopic eczema after 1 of age year. No significant difference in terms of prevention of asthma, rhinitis, wheeze, allergic diseases and sensation. In brief, a probiotic supplement is expected to become a novel potential strategy for infant eczema and atopic eczema.

Identification and characterization of the chemical components of Iris tectorum Maxim. and evaluation of their nitric oxide inhibitory activity.

RATIONALE: Iris tectorum Maxim. is a traditional medicinal herb that is commonly used to treat inflammatory conditions. The present study investigated the fragmentation patterns of isoflavone glycosides and their qualitative analysis. In addition, lipopolysaccharide (LPS)-induced RAW264.7 macrophages were used to evaluate the anti-inflammatory properties of I. tectorum Maxim. samples collected at different time points during the year. METHODS: High-performance liquid chromatography/quadrupole time-of-flight tandem mass spectrometry (HPLC/QTOF-MS/MS) and HPLC with diode-array detection were employed for qualitative and quantitative analysis. The fragmentation patterns of the isoflavones were observed in negative electrospray ionization mode with collision-induced dissociation (CID). Their anti-inflammatory activity was assessed via nitric oxide (NO) production in LPS-treated RAW264.7 macrophages. RESULTS: A total of 15 chemical components were observed and tentatively identified using HPLC/QTOF-MS/MS. At low collision energy, the relative abundances of the aglycone radical anions Y0 - , [Y0 - H]-• , [Y0 - CH3 ]-• and [Y0 - H- CH2 ]-• were used for the structural characterization of tectoridin and tectorigenin-4'-O-ß-D-glucoside. The radical ions [Y0 - CH3 ]-• and [Y0 - H - 2CH3 ]-• were also employed to differentiate between iristectorin A and iristectorin B based upon their high-energy CID spectra. Levels of 9.02 mg/g of tectoridin and 1.04 mg/g of tectorigenin were found in samples collected in June, which exhibited 69.7% NO inhibitory activity. CONCLUSIONS: The characteristic fragmentation patterns enabled us to reliably identify isoflavone glycosides. The results of the quantitative determination and NO inhibitory activity offer insight into the optimal I. tectorum Maxim. harvesting time.

Traditional Chinese medicine for psoriasis vulgaris: A Protocol of a prospective, multicenter cohort study.

INTRODUCTION: The incidence of psoriasis vulgaris is increasing worldwide. Chronic recurrence of the disease, as well as accompanying cardiovascular disease, metabolic syndrome, and depression has affected the physical and mental health of these patients. Psoriasis vulgaris is a difficult and major disease in the dermatology field. Short-term curative effects using conventional therapy for psoriasis vulgaris has made major strides. However, traditional Chinese medicine (TCM) treatment has long-term curative advantages for psoriasis vulgaris but lacks the scientific and clinical evidence for its use. This study intends to demonstrate and provide scientific and clinical evidence for the use of TCM to delay the recurrence of psoriasis vulgaris. METHODS AND ANALYSIS: This will be a prospective, multicenter cohort study. We intend to recruit 1521 psoriasis vulgaris patients from 14 hospitals in Beijing, Tianjin, and Hebei. Treatment will be based on the diagnosis specifications and clinical practice guidelines of TCM and conventional therapy. During inclusion and the subsequent follow-up period, doctors through electronic case reports will collect different therapeutic TCM regimens and conventional therapy that were administered. Information on life condition, skin lesions at each visit, World Health Organization Quality of Life Instruments, Zung Self-rating Anxiety Scale, Zung Self-assessment of Depression, laboratory examinations, incidence of new rash and recurrence during the remission and recurrence stages will be recorded. ETHICS AND DISSEMINATION: The clinical trial protocol for this study was approved by the ethics committee of the Beijing hospital of TCM affiliated to capital medical university (Ethics number: 2019BL02-010-02). We will publish and present our results at national and international conferences and in peer-reviewed journals specialized in dermatology. TRIAL REGISTRATION: This protocol has been registered in clinicaltrials. gov (ChiCTR1900021629).

Preparation and in vitro evaluation of multi-target-directed selenium-chondroitin sulfate nanoparticles in protecting against the Alzheimer's disease.

The purpose of this study was to ascertain the effect of selenium-chondroitin sulfate nanoparticles (CS@Se) on multi-target-directed therapy for the treatment of Alzheimer's disease (AD). CS@Se nanoparticles were successfully synthesized, and their therapeutic effects were studied in in vitro AD models. CS@Se effectively inhibited amyloid-ß (Aß) aggregation and protected SH-SY5Y cells from Aß1-42-induced cytotoxicity. Moreover, CS@Se significantly decreased okadaic acid-induced actin cytoskeleton instability in SH-SY5Y cells. In addition, CS@Se decreased the levels of reactive oxygen species (ROS) and malondialdehyde (MDA) and increased the levels of glutathione peroxidase (GSH-Px). The Western blot results indicated that CS@Se attenuated the hyperphosphorylation of tau (Ser396/Ser404) by regulating the expression of GSK-3ß. In summary, this study demonstrated that CS@Se could inhibit the aggregation of Aß, reduce damage to the cytoskeleton, mitigate oxidative stress and attenuate the hyperphosphorylation of tau protein. CS@Se might be a potent multi-functional agent for the treatment of AD and thus warrants further research and evaluation.

Efficacy and safety of pioglitazone for treatment of plaque psoriasis: a systematic review and meta-analysis of randomized controlled trials.

Background: A growing number of studies have shown that thiazolidinediones (TZD) can be antipsoriatic. Pioglitazone is a representative of the class of antidiabetic drugs known as TZD. TZD can activate nuclear peroxisome proliferator-activated receptors (PPAR)-c. PPARs are expressed on epidermal keratinocytes and exert their effects by promoting the terminal differentiation of keratinocytes, inhibiting epidermal growth, and reducing inflammatory responses. These observations suggest that TZD have potential benefits in the treatment of cutaneous and metabolic pathologies associated with psoriasis.Objective: A systematic review and meta-analysis was carried out to evaluate the efficacy of combined pioglitazone treatment. We point out three controversial side effects from administration of pioglitazone in psoriasis: elevated liver enzymes, weight gain, and nausea.Study selection: Randomized, single blind, or double blind, published studies of pioglitazone compared with placebo given to patients with plaque psoriasis for 10 weeks or 12 weeks were considered for inclusion in this review. The primary outcomes were 75% or greater improvement in the Psoriasis Area and Severity Index score from baseline (PASI 75) with pioglitazone.Data collection and analysis: The systematic literature search was conducted in the PubMed, Embase, Google Scholar, and Cochrane databases from inception up to December 20 2018. Data analysis was done using Revman 5.3 Haymarket, London, United Kingdom.Main results: We included six studies (three publications of pioglitazone only; three publications of pioglitazone combination therapy) comprising a total of 294 patients (n = 149 with pioglitazone only and n = 145 with pioglitazone combination therapy) in the analysis. There was a significant PASI 75 response, in the pioglitazone only subgroup as compared to placebo (OR = 8.74, 95% CI 3.76-20.31, p < .00001), and the pioglitazone combination subgroup as compared to placebo (OR = 4.64, 95% CI 2.03-10.60, p < .00001), others, the total of pioglitazone as compared to placebo (OR = 6.37, 95% CI 3.55-11.43, p < .00001), and tests of subgroup differences show: p = .29, I2 = 9.5%. The incidence rate of elevated liver enzymes (OR = 3.70, 95% CI 0.56-24.31, p = .99), weight gain (OR = 1.44, 95% CI 0.60-3.47, p = .41), and nausea (OR = 0.76, 95% CI 0.23-2.49, p = .65) were not significantly different compared with the control group.Conclusion: Pioglitazone has efficacy for the treatment of plaque psoriasis. There is no significant difference between patients treated with pioglitazone only or in combination with other therapies. The incidence rate of side effects associated with pioglitazone treatment such as elevated liver enzymes, weight gain, and nausea were not significantly different compared with the control group.

Identification and biochemical analyses of selective CB2 agonists.

Cannabinoid CB1 and CB2 receptors are activated by Δ9-tetrahydrocannabinol, a psychoactive component of marijuana. The cannabinoid CB1 receptor is primarily located in the brain and is responsible for the psychoactive side effects, whereas the cannabinoid CB2 receptor is located in immune cells and is an attractive target for immune-related maladies. We identify small molecules that selectively bind to the cannabinoid CB2 receptor and can be further developed into therapeutics. The affinity of three molecules, ABK5, ABK6, and ABK7, to the cannabinoid CB2 receptor was determined with radioligand competition binding. The potency of G-protein coupling was determined with GTPγS binding. The three compounds bound selectively to the cannabinoid CB2 receptor, and no binding to the cannabinoid CB1 receptor was detected up to 10 µM. Immunoblotting studies show that the amount of ERK1/2 and MEK phosphorylation increased in a Gi/o-dependent manner. Furthermore, an immune cell line (Jurkat cells) was treated with ABK5, and as a result, inhibited cell proliferation. These three compounds are novel cannabinoid CB2 receptor agonists and hold promise to be further developed to treat inflammation and the often-associated pain.

Leveraging the IncuCyte Technology for Higher-Throughput and Automated Chemotaxis Assays for Target Validation and Compound Characterization.

Chemotaxis is the directional movement of cells in response to a chemical stimulus and is vital for many physiological processes, including immune responses, tumor metastasis, wound healing, and blood vessel formation. Therefore, modulation of chemotaxis is likely to be of therapeutic benefit. Hence, a high-throughput means to conduct chemotaxis assays is advantageous for lead evaluation and optimization in drug discovery. In this study, we have validated a novel approach for a higher-throughput, label-free, image-based IncuCyte chemotaxis assay encompassing various cell types, including T cells, B cells, mouse Th17, immature and mature dendritic cells, monocyte THP-1, CCRF-CEM, monocytes, neutrophils, macrophages, and MDA-MB-231. These assays enable us to visualize chemotactic cell migration in real time and perform kinetic cell motility studies on an automated platform, thereby allowing us to incorporate the quantitative studies of cell migration behavior into a routine drug discovery screening cascade.

Development of a RapidFire mass spectrometry assay and a fluorescence assay for the discovery of kynurenine aminotransferase II inhibitors to treat central nervous system disorders.

Kynurenine aminotransferases convert kynurenine to kynurenic acid and play an important role in the tryptophan degradation pathway. Kynurenic acid levels in brain have been hypothesized to be linked to a number of central nervous system (CNS) disorders. Kynurenine aminotransferase II (KATII) has proven to be a key modulator of kynurenic acid levels in brain and, thus, is an attractive target to treat CNS diseases. A sensitive, high-throughput, label-free RapidFire mass spectrometry assay has been developed for human KATII. Unlike other assays, this method is directly applicable to KATII enzymes from different animal species, which allows us to select proper animal model(s) to evaluate human KATII inhibitors. We also established a coupled fluorescence assay for human KATII. The short assay time and kinetic capability of the fluorescence assay provide a useful tool for orthogonal inhibitor validation and mechanistic studies.

Discovery, synthesis, and molecular pharmacology of selective positive allosteric modulators of the δ-opioid receptor.

Allosteric modulators of G protein-coupled receptors (GPCRs) have a number of potential advantages compared to agonists or antagonists that bind to the orthosteric site of the receptor. These include the potential for receptor selectivity, maintenance of the temporal and spatial fidelity of signaling in vivo, the ceiling effect of the allosteric cooperativity which may prevent overdose issues, and engendering bias by differentially modulating distinct signaling pathways. Here we describe the discovery, synthesis, and molecular pharmacology of δ-opioid receptor-selective positive allosteric modulators (δ PAMs). These δ PAMs increase the affinity and/or efficacy of the orthosteric agonists leu-enkephalin, SNC80 and TAN67, as measured by receptor binding, G protein activation, ß-arrestin recruitment, adenylyl cyclase inhibition, and extracellular signal-regulated kinases (ERK) activation. As such, these compounds are useful pharmacological tools to probe the molecular pharmacology of the δ receptor and to explore the therapeutic potential of δ PAMs in diseases such as chronic pain and depression.

Identification of selective agonists and positive allosteric modulators for µ- and δ-opioid receptors from a single high-throughput screen.

Hetero-oligomeric complexes of G protein-coupled receptors (GPCRs) may represent novel therapeutic targets exhibiting different pharmacology and tissue- or cell-specific site of action compared with receptor monomers or homo-oligomers. An ideal tool for validating this concept pharmacologically would be a hetero-oligomer selective ligand. We set out to develop and execute a 1536-well high-throughput screen of over 1 million compounds to detect potential hetero-oligomer selective ligands using a ß-arrestin recruitment assay in U2OS cells coexpressing recombinant µ- and δ-opioid receptors. Hetero-oligomer selective ligands may bind to orthosteric or allosteric sites, and we might anticipate that the formation of hetero-oligomers may provide novel allosteric binding pockets for ligand binding. Therefore, our goal was to execute the screen in such a way as to identify positive allosteric modulators (PAMs) as well as agonists for µ, δ, and hetero-oligomeric receptors. While no hetero-oligomer selective ligands were identified (based on our selection criteria), this single screen did identify numerous µ- and δ-selective agonists and PAMs as well as nonselective agonists and PAMs. To our knowledge, these are the first µ- and δ-opioid receptor PAMs described in the literature.

High-throughput high-content imaging assays for identification and characterization of selective AXL pathway inhibitors.

Receptor tyrosine kinases (RTKs) regulate a wide range of important biological activities, including cell proliferation, differentiation, migration, and apoptosis. Abnormalities in RTKs are involved in numerous diseases, including cancer and other proliferative disorders. AXL belongs to the TAM (Tyso3, AXL, and Mer) family of RTKs. The AXL signaling pathway represents an attractive target for the treatment of diseases, such as cancer. Using phospho-AKT as readout, a high-throughput 384-well cell-based assay was established in the NCI-H1299 human non-small cell lung carcinoma cell line to evaluate compound potency in inhibiting AXL pathway activation. In addition, a counter screen assay was established in the same cellular background to differentiate AXL kinase inhibitors from AXL receptor antagonists, which block the interaction of AXL and its natural ligand GAS6. These cell-based functional assays are useful tools in the identification and optimization of small molecules and biological reagents for potential therapeutics for the treatment of GAS6/AXL-related diseases.

Efficacy of binary combinations of botanical pesticides for rotifer elimination in microalgal cultivation.

Binary interactions of celangulin, matrine and toosendanin against the rotifer Brachionus plicatilis were studied. Types of interactions (antagonism, synergism and addition) were dependent on the biocides themselves and their ratios in combinations. Mixtures of matrine/toosendanin mainly produced addition owing to their similar modes of action aiming at the nervous system. Combinations of celangulin mixed with matrine or toosendanin at 1:9 exhibited synergism, which is attributed to the interference of matrine or toosendanin with the detoxification enzymes of celangulin. Both the synergistic combinations were inappropriate for rotifer extermination in Isochrysis sp. cultivation owing to the high phytotoxicity resulting from the absence of cell walls. However, the celangulin/toosendanin (1:9) mixture decreased rotifer reproduction without damaging cells of Chlorella and Nannochloropsis sp. Application of frequent, low doses of celangulin/toosendanin (1:9) mixture also reduced the dosage of biocides, thereby reducing the cost of exterminating rotifers, and indicating a considerable practical application in microalgal cultivation.

Development of novel, 384-well high-throughput assay panels for human drug transporters: drug interaction and safety assessment in support of discovery research.

Transporter proteins are known to play a critical role in affecting the overall absorption, distribution, metabolism, and excretion characteristics of drug candidates. In addition to efflux transporters (P-gp, BCRP, MRP2, etc.) that limit absorption, there has been a renewed interest in influx transporters at the renal (OATs, OCTs) and hepatic (OATPs, BSEP, NTCP, etc.) organ level that can cause significant clinical drug-drug interactions (DDIs). Several of these transporters are also critical for hepatobiliary disposition of bilirubin and bile acid/salts, and their inhibition is directly implicated in hepatic toxicities. Regulatory agencies took action to address transporter-mediated DDI with the goal of ensuring drug safety in the clinic and on the market. To meet regulatory requirements, advanced bioassay technology and automation solutions were implemented for high-throughput transporter screening to provide structure-activity relationship within lead optimization. To enhance capacity, several functional assay formats were miniaturized to 384-well throughput including novel fluorescence-based uptake and efflux inhibition assays using high-content image analysis as well as cell-based radioactive uptake and vesicle-based efflux inhibition assays. This high-throughput capability enabled a paradigm shift from studying transporter-related issues in the development space to identifying and dialing out these concerns early on in discovery for enhanced mechanism-based efficacy while circumventing DDIs and transporter toxicities.

GPCR assay automation for leveraging lead optimization.

The drug discovery process is dependent on the ability of screening efforts to identify and optimize lead compounds with therapeutic potential. Because G-protein-coupled receptors (GPCRs) account for the most "druggable" targets, the development of high-throughput, low-cost, and high-density GPCR assays to accommodate the increasing size of compound collections is significant. In this study, we report the application of an advanced LEADseeker robotic platform equipped with customized IT solutions for rapid data transfer to reduce assay cycles times for support of GPCR panel screening. The advantages of assay throughput, format, automation design, data management flow, and data reproducibility are discussed in terms of gains in productivity for lead optimization. The GPCR robotic platform demonstrates how automation technology can leverage traditional drug discovery processes by providing consistent and reliable data packages to expedite lead optimization efforts.

Design of an automated enhanced-throughput platform for functional characterization of positive allosteric modulator-induced leftward shifts in apparent agonist potency in vitro.

Prosecution of positive allosteric modulator (PAM) targets demands a specialized assay toolset. Many GPCR or ion channel targets are adaptable to functional assays whereby PAM efficacy can be inferred from left or rightward shifts in the concentration-response curves of orthosteric agonist. The inherent emphasis on throughput and occasional paucity of radioligands for a diverse array of allosteric modulator targets yields a need for an enhanced throughput agonist potency shift assay. Here, we describe a process by which such an assay was automated with robust, reproducible in vitro pharmacology. In direct comparison with a manual CRC shift assay, the enhanced throughput automated platform described here delivered near identical rank orders (r(2) = 0.75) at ~4-fold throughput/assay iteration. Correspondingly, average cycle time/plate decreased from 104 to 72 minutes. We also observed reductions in assay interference associated with compounds exhibiting ago-allosterism, which we attribute to preread compound incubation periods which are more precisely time-constrained under automation control. By leveraging automated laboratory technology, we have achieved meaningful throughput with no sacrifice of precision. Rather than to be target-class specific, the present process was specifically designed to serve as a platform template for a variety of cell-based functional allosteric modulation assays.

Characterization of the programmed cell death induced by metabolic products of Alternaria alternata in tobacco BY-2 cells.

Alternaria alternata has received considerable attention in current literature and most of the studies are focused on its pathogenic effects on plant chloroplasts, but little is known about the characteristics of programmed cell death (PCD) induced by metabolic products (MP) of A. alternata, the effects of the MP on mitochondrial respiration and its relation to PCD. The purpose of this study was to explore the mechanism of MP-induced PCD in non-green tobacco BY-2 cells and to explore the role of mitochondrial inhibitory processes in the PCD of tobacco BY-2 cells. MP treatment led to significant cell death that was proven to be PCD by the concurrent cytoplasm shrinkage, chromatin condensation and DNA laddering observed in the cells. Moreover, MP treatment resulted in the overproduction of reactive oxygen species (ROS), rapid ATP depletion and a respiratory decline in the tobacco BY-2 cells. It was concluded that the direct inhibition of the mitochondrial electron transport chain (ETC), alternative pathway (AOX) capacity and catalase (CAT) activity by the MP might be the main contributors to the MP-induced ROS burst observed in tobacco BY-2 cells. The addition of adenosine together with the MP significantly inhibited ATP depletion without preventing PCD; however, when the cells were treated with the MP plus CAT, ROS overproduction was blocked and PCD did not occur. The data presented here demonstrate that the ROS burst played an important role in MP-induced PCD in the tobacco BY-2 cells.

[Inhibition effects of salt stress on photosynthetic activity of Rumex K-1].

With Rumex K-1 seedlings as test materials, this paper studied the effects of different concentration (100-300 mmol x L(-1)) NaCl and KCl on their leaf photosynthetic activity and osmotic adjustment. The results showed that at the concentration of 200 mmol x L(-1), NaCl had greater inhibition effect on the leaf photosynthetic activity than KCl, but at 300 mmol x L(-1), the inhibition effect of KCl was much greater than NaCl. After treated with 300 mmol x L(-1) of KCl and NaCl, the leaf water potential was -0.93 MPa and -1.05 MPa, and the osmotic potential was -1.43 MPa and -1.10 MPa, respectively, indicating that the increased damage caused by 300 mmol x L(-1) of KCl was not from osmotic stress. Under the stress of 300 mmol KCl x L(-1), the leaf Na+ concentration decreased by 88.6%, compared with the control, while the supplement of 25 mmol NaCl x L(-1) could obviously alleviate the damage of KCl on leaf photosynthesis, which proved that the deficit of Na+ and the accumulation of K+ in Rumex K-1 leaves could be responsible to the enhanced damage caused by 300 mmol x L(-1) of KCl.