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Hepatocellular carcinoma (HCC) is a common type of cancer that shows great morbidity and mortality rates. However, there are limited available drugs to treat HCC. The present work focused on discovering the potential anti-HCC compounds from traditional Chinese medicine (TCM) by employing high-throughput sequencing-based high-throughput screening (HTS2) together with the liver cancer pathway-associated gene signature. HTS2 assay was adopted for identifying herbs. Protein-protein interaction (PPI) network analysis and computer-aided drug design (CADD) were used to identify key targets and screen the candidate natural products of herbs. Molecular docking, network pharmacology analysis, western blotting, immunofluorescent staining, subcellular fractionation experiment, dual-luciferase reporter gene assay, surface plasmon resonance (SPR) as well as nuclear magnetic resonance (NMR) were performed to validate the ability of compound binding with key target and inhibiting its function. Moreover, cell viability, colony-forming, cell cycle assay and animal experiments were performed to examine the inhibitory effect of compound on HCC. We examined the perturbation of 578 herb extracts on the expression of 84 genes from the liver cancer pathway, and identified the top 20 herbs significantly reverting the gene expression of this pathway. Signal transducer and activator of transcription 3 (STAT3) was identified as one of the key targets of the liver cancer pathway by PPI network analysis. Then, by analyzing compounds from top 20 herbs utilizing CADD, we found ginsenoside F2 (GF2) binds to STAT3 with high affinity, which was further validated by the results from molecular docking, SPR and NMR. Additionally, our results showed that GF2 suppresses the phosphorylation of Y705 of STAT3, inhibits its nuclear translocation, decreases its transcriptional activity and inhibits the growth of HCC in vitro and in vivo. Based on this large-scale transcriptional study, a number of anti-HCC herbs were identified. GF2, a compound derived from TCM, was found to be a chemical basis of these herbs in treating HCC. The present work also discovered that GF2 is a new STAT3 inhibitor, which is able to suppress HCC. As such, GF2 represents a new potential anti-HCC therapeutic strategy. [Display omitted] [ABSTRACT FROM AUTHOR]

Gynostemma pentaphyllum (Thunb.) Makino (G. pentaphyllum) is an oriental herb documented to treat many diseases, including obesity, hyperlipidemia, metabolic syndromes and aging. However, the anti-obesity mechanism of G. pentaphyllum remains poorly understood. To reveal the anti-obesity mechanism of G. pentaphyllum Extract (GPE) in High-Fat Diet (HFD)-induced obese mice through untargeted metabolomics, Real-Time Quantitative PCR (RT-qPCR), and immunohistochemical experiments. Additionally, to tentatively identify the active constituents through LC-MS/MS and molecular docking approaches. GPE was prepared using ethanol reflux and purified by HP-20 macroporous resins. The components of GPE were identified by Liquid Chromatography- Mass Spectrometry (LC-MS) system. Forty-two C57BL/6 J mice were randomly and evenly divided into six groups, with seven mice in each group: the control group, obese model group, Beinaglutide group (positive control), and GPE low, medium, and high-dose groups (50 mg/kg, 100 mg/kg, and 200 mg/kg of 80% ethanol extract). Body weight, liver weight, blood glucose, blood lipids, and liver histopathological changes were assessed. Untargeted metabolomics was employed to characterize metabolic changes in obese mice after GPE treatment. The expression of genes related to differential metabolites was verified using Real-Time Quantitative PCR (RT-qPCR) and immunohistochemical experiments. The constituents with anti-obesity effects from GPE were tentatively identified through molecular docking approaches. A total of 17 compounds were identified in GPE. GPE significantly lowered body weight, total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) in obese mice and reduced liver weight and hepatic steatosis. Serum metabolomics identified 20 potential biomarkers associated with GPE treatment in obese mice, primarily related to tryptophan metabolism. GPE treatment downregulated the expression of Slc6a19 and Tph1 and upregulated Ucp1 expression. Molecular docking illustrated that compounds such as 20(R)-ginsenoside Rg3, Araliasaponin I, Damulin B, Gypenoside L, Oleifolioside B, and Tricin7-neohesperidoside identified in GPE exhibited favorable interaction with Tph1. The extract of G. pentaphyllum can inhibit the absorption of tryptophan and its conversion to 5-HT through the Slc6a19/Tph1 pathway, upregulating the expression of Ucp1, thereby promoting thermogenesis in brown adipose tissue, facilitating weight loss, and mitigating symptoms of fatty liver. Triterpenoids such as Araliasaponin I, identified in GPE, could be the potential inhibitor of Tph1 and responsible for the anti-obesity activities. [Display omitted] • Gynostemma pentaphyllum (Thunb.) Makino demonstrated efficacy for weight loss. • This effect is mediated by inhibiting the Slc6a19/Tph1 pathway and activating Ucp1. • Araliasaponin I from the herb could be responsible for the anti-obesity activities. [ABSTRACT FROM AUTHOR]

A protocol for efficiently identifying ligands directly interacting with a target protein in complex extracts of medicinal herbs was proposed by combining an adapted 2D perfect-echo Carr-Purcell-Meiboom-Gill heteronuclear single quantum correlation (PE-CPMG HSQC) spectrum with metabolomic analysis. PE-CPMG HSQC can suppress the signal interference from the target protein, allowing more accurate peak quantification than conventional HSQC. Inspired from untargeted metabolomics, regions of interest (ROIs) are constructed and quantified for the mixture or complex extract samples with and without a target protein, and then a binding index (BI) of each ROI is determined. ROIs or corresponding peaks significantly perturbed by the presence of the target protein (BI ≥1.5) are detected as differential features, and potential binding ligands identified from the differential features can be equated with bioactive markers associated with the 'treatment' of the target protein. Quantifying ROI can inclusively report the ligand bindings to a target protein in fast, intermediate and slow exchange regimes on nuclear magnetic resonance (NMR) time scale. The approach was successfully implemented and identified Angoroside C, Cinnamic acid and Harpagoside from the extract of Scrophularia ningpoensis Hemsl. as ligands binding to peroxisome proliferator-activated receptor γ. The proposed 2D NMR-based approach saves excess steps for sample processing and has a higher chance of detecting the weaker ligands in the complex extracts of medicinal herbs. We expect that this approach can be applied as an alternative to mining the potential ligands binding to a variety of target proteins from traditional Chinese medicines and herbal extracts. [Display omitted] • Suppress protein interference and render cleaner background than conventional HSQC. • Be able to detect ligands with low-to-moderate affinities. • Simplify the sample processing and does not require the immobilization of proteins. [ABSTRACT FROM AUTHOR]

β-Polypeptides are known to adopt helical secondary structure in organic solvents, even for rather short chain lengths. It is investigated whether a short α-polypeptide with amino-acid side chains that enable β-peptides to adopt helical structures, can maintain or adopt stable helical structure in methanol or in water. The molecular dynamics simulations do not predict a particular fold, which indicates an essential role for the additional methylene moiety in the backbone of β-peptides regarding helix stability. [Copyright &y& Elsevier]

Objective: To determine the relationships between circulating sex hormone-binding globulin (SHBG) and oxidized low-density lipoprotein (ox-LDL), total oxidant status, total antioxidant capacity, oxidative stress index, malondialdehyde, and the high-density lipoprotein (HDL) inflammatory index in patients with polycystic ovary syndrome (PCOS) and to investigate the effect of oxidative stress on the expression of SHBG and its mechanism in HepG2 cells.Design: Cross-sectional study.Setting: University hospital.Patient(s): A total of 533 women with PCOS and 292 control women were included.Intervention(s): None.Main Outcome Measure(s): Circulating SHBG, hormones, and metabolic and oxidative stress indices were determined in all subjects. The effects of ox-LDL and ox-HDL on the mRNA and protein expression of SHBG and related transcription factors were observed in HepG2 cells.Result(s): The HDL inflammatory index, total oxidant status, oxidative stress index, and malondialdehyde levels were significantly higher in the three PCOS subgroups with different SHBG levels than in the controls. The ox-LDL and total antioxidant capacity were higher in the PCOS subgroups with SHBG levels <75th percentile compared with the controls or the PCOS subgroup with SHBG levels ≥75th percentile. In HepG2 cells, the SHBG concentration in the culture supernatant, the mRNA levels of SHBG and hepatocyte nuclear factor-4α (HNF-4α), and the protein levels of HNF-4α were significantly lower in ox-LDL- and ox-HDL-treated cells than in the control cells and lipoprotein-treated cells.Conclusion(s): Oxidative stress inhibits the expression and secretion of SHBG by downregulating HNF-4α in vitro and may be an important factor promoting the occurrence of hyperandrogenemia in PCOS. [ABSTRACT FROM AUTHOR]

The physicochemical properties and biological activities of polysaccharides depend on their structures. Monosaccharide composition analysis is indispensable for the structural characterization of polysaccharides and is helpful in the quality control of polysaccharide preparation. Here, using a model mixture and tamarind seed polysaccharide as examples, we demonstrated that a quantitative 2D NMR method, gsHSQC i (three gradient-selective Heteronuclear Single Quantum Coherence spectra acquired with incremented repetition times, i = 1, 2, 3) can directly quantify a variety of monosaccharides in solution with adequate precision and accuracy, requiring no derivatization, postprocessing steps and column separation. Both anomeric and non-anomeric signals of monosaccharides can be utilized for content determination. More accurate quantification of fructose in a mixture containing nine monosaccharides is obtained, which is difficult to achieve by quantitative 1D 1HNMR and the common PMP-HPLC method (high-performance liquid chromatography through pre-column derivatization with 1-phenyl-3-methyl-5-pyrazolone) due to the peak overlapping and the poor derivatization efficiency, respectively. The results also revealed that Na[Fe(EDTA)] can serve as a proper relaxation-enhancing agent for saccharide samples to save experimental time. We expect that this approach can be applied as an alternative to analyzing the monosaccharide composition and be helpful in interpreting the structure of polysaccharides. [Display omitted] • Direct quantification of a variety of monosaccharides in the mixture without derivatization and column separation • Measure the monosaccharide mixture directly without a pre-established standard curve for each monosaccharide • More non-anomeric signals can be utilized for quantification in gsHSQC i compared with qHNMR • Na[Fe(EDTA)] is used as a proper relaxation-enhancing agent to save experimental time • DMSO 2 is used as an internal reference for quantitation. [ABSTRACT FROM AUTHOR]

The anti-hyperlipidemic effects of crude crabapple extracts derived from Malus ‘Red jade’, Malus hupehensis (Pamp.) Rehd. and Malus prunifolia (Willd.) Borkh. were evaluated on high-fat diet induced obese (HF DIO) mice. The results revealed that some of these extracts could lower serum cholesterol levels in HF DIO mice. The same extracts were also parallelly analyzed by LC–MS in both positive and negative ionization modes. Based on the pharmacological results, 22 LC–MS variables were identified to be correlated with the anti-hyperlipidemic effects using partial least square discriminant analysis (PLS-DA) and independent samples t -test. Further, under the guidance of the bioactivity-correlated LC–MS signals, 10 compounds were targetedly isolated and enriched using UPLC-DAD-MS-SPE and identified/elucidated by NMR together with MS/MS as citric acid(1), p -coumaric acid(2), hyperoside(3), myricetin(4), naringenin(5), quercetin(6), kaempferol(7), gentiopicroside(8), ursolic acid(9) and 8-epiloganic acid(10). Among these 10 compounds, 6 compounds, hyperoside(3), myricetin(4), naringenin(5), quercetin(6), kaempferol(7) and ursolic acid(9), were individually studied and reported to indeed have effects on lowering the serum lipid levels. These results demonstrated the efficiency of this strategy for drug discovery. In contrast to traditional routes to discover bioactive compounds in the plant extracts, targeted isolation and identification of bioactive compounds in the crude plant extracts using UPLC-DAD-MS-SPE/NMR based on pharmacology-guided PLS-DA of LC–MS data brings forward a new efficient dereplicated approach to natural products research for drug discovery. [ABSTRACT FROM AUTHOR]