Untargeted foodomics for authenticating the organic farming of water spinach (Ipomoea aquatica).

This study aimed to conduct a comprehensive analysis of the primary and secondary metabolites of water spinach (Ipomoea aquatica) using hydrophilic interaction liquid chromatography coupled with Orbitrap high-resolution mass spectrometry (HILIC-Orbitrap-HRMS). Certified samples from two cultivars, Green stem water spinach (G) and White stem water spinach (W) cultivated using organic and conventional farming methods, were collected from the Hong Kong market. Multivariate analysis was used to differentiate water spinach of different cultivars and farming methods. We identified 12 metabolites to distinguish between G and W, 26 metabolites to identify G from organic farming and 8 metabolites to identify W from organic farming. Then, two metabolites, isorhamnetin and jasmonic acid, have been proposed to serve as biomarkers for organic farming (in both G and W). Our foodomics findings provide useful tools for improving the crop performance of water spinach under abiotic/biotic stressesand authentication of organic produce.

Insights into serum metabolic biomarkers for early detection of incident diabetic kidney disease in Chinese patients with type 2 diabetes by random forest.

Diabetic kidney disease (DKD) is a leading cause of end-stage renal disease (ESRD) worldwide. Early detection is critical for the risk stratification and early intervention of progressive DKD. Serum creatinine (sCr) and urine output are used to assess kidney function, but these markers are limited by their delayed changes following kidney pathology, and lacking of both sensitivity and accuracy. Hence, it is essential to illustrate potential diagnostic indicators to enhance the precise prediction of early DKD. A total of 194 Chinese individuals include 30 healthy participants (Stage 0) and 164 incidents with type 2 diabetes (T2D) spanning from DKD's Stage 1a to 4 were recruited and their serums were subjected for untargeted metabolomic analysis. Random forest (RF), a machine learning approach, together with univariate linear regression (ULR) and multivariate linear regression (MvLR) analysis were applied to characterize the features of untargeted metabolites of DKD patients and to identify candidate DKD biomarkers. Our results indicate that 2-(α-D-mannopyranosyl)-L-tryptophan (ADT), succinyladenosine (SAdo), pseudouridine and N,N,N-trimethyl-L-alanyl-L-proline betaine (L-L-TMAP) were associated with the development of DKD, in particular, the latter three that were significantly elevated in Stage 2-4 T2D incidents. Each of the four metabolites in combination with sCr achieves better performance than sCr alone with area under the receiver operating characteristic curve (AUC) of 0.81-0.91 in predicting DKD stages. An average of 3.9 years follow-up study of another cohort including 106 Stage 2-3 patients suggested that "urinary albumin-to-creatinine ratio (UACR) + ADT + SAdo" can be utilized for better prognosis evaluation of early DKD (average AUC = 0.9502) than UACR without sexual difference.

The Lignan-Rich Fraction from Sambucus williamsii Hance Exerts Bone Protective Effects via Altering Circulating Serotonin and Gut Microbiota in Rats.

Our previous study revealed that the bone anabolic effects of the lignan-rich fraction (SWCA) from Sambucus williamsii Hance was involved in modulating the metabolism of tryptophan in vivo and inhibiting serotonin (5-HT) synthesis in vitro. This study aimed to determine how SWCA modulates bone metabolism via serotonin in vivo. The effects of SWCA were evaluated by using 4-month-old Sprague-Dawley (SD) ovariectomized rats. The serum levels of 5-HT and kynurenine, the protein expressions of tryptophan hydroxylase 1 (TPH-1) and TPH-2, the genes and proteins related to the 5-HT signaling pathway as well as gut microbiota composition were determined. SWCA treatment alleviated bone loss and decreased serum levels of serotonin, which was negatively related to bone mineral density (BMD) in rats. It suppressed the protein expression of TPH-1 in the colon, and reversed the gene and protein expressions of FOXO1 and ATF4 in the femur in OVX rats, while it did not affect the TPH-2 protein expression in the cortex. SWCA treatment escalated the relative abundance of Antinobacteria and modulated several genera relating to BMD. These findings verified that the bone protective effects of lignans were mediated by serotonin, and provided evidence that lignans might be a good source of TPH-1 inhibitors.

Lactobacillus casei Strain Shirota Ameliorates Dextran Sulfate Sodium-Induced Colitis in Mice by Increasing Taurine-Conjugated Bile Acids and Inhibiting NF-κB Signaling via Stabilization of IκBα.

Inflammatory bowel disease (IBD) is a chronic progressive intestinal inflammatory disease, characterized by an altered gut microbiota composition and accompanying alterations in circulatory bile acids. Increasing evidence supports the beneficial effect of probiotics intake on health. Introduction of probiotics to the intestines can modulate gut microbiota composition and in turn regulate the host immune system and modify the inflammatory response. Probiotics can also improve intestinal barrier function and exhibit a positive impact on host physiological and pathological conditions via gut microbiota-derived metabolites. Previous studies have demonstrated that Lactobacillus casei strain Shirota (LcS) treatment could inhibit clinical manifestation of colitis in dextran sulfate sodium (DSS)-induced mice, however, the underlying mechanisms remain unknown. In this study, we employed the DSS-induced acute colitis mouse model to investigate the anti-inflammatory effects of LcS and related mechanisms. Administration of LcS ameliorated the severity of DSS-induced colitis and enhanced intestinal integrity via induction of mucin-2 and occludin expression in colons. Fecal microbiota analysis showed that LcS increased the relative abundance of beneficial bacterial species in colitic mice, whereas the relative abundance of pathobionts was reduced. Additionally, LcS treatment modulated circulating bile acid profiles in colitic mice. In mice treated with LcS, we identified increased levels of primary taurine-conjugated bile acids, including taurocholic acid (TCA) and taurochenodeoxycholic acid (TCDCA). LcS treatment also increased the levels of secondary taurine-conjugated bile acids, including taurodeoxycholic acid (TDCA) and tauroursodeoxycholic acid (TUDCA). Moreover, LcS treatment exhibited a suppressive effect on the hydroxylated primary bile acids α-muricholic acid (α-MCA) and ß-muricholic acid (ß-MCA). We further demonstrated that LcS treatment suppressed the expression of pro-inflammatory mediators interferon-gamma (IFN-γ) and nitric oxide (NO), and increased the expression of the anti-inflammatory mediator interleukin-10 (IL-10) in colon tissues, potentially as a result of altered bile acid profiles. Mechanistically, we showed that LcS treatment suppressed the activation of nuclear factor-kappa B (NF-κB) signaling via stabilization of inhibitor of NF-κB alpha (IκBα). Altogether, we have demonstrated the therapeutic effects of LcS in DSS-induced colitis, providing new insights into its effect on bile acid metabolism and the related anti-inflammatory mechanisms. Our findings provide support for the application of LcS in the treatment of IBD.

Biological effect of dysregulated LBX1 on adolescent idiopathic scoliosis through modulating muscle carbohydrate metabolism.

BACKGROUND CONTEXT: Abnormal energy metabolism such as lower body weight and body mass index (BMI) and less fat mass is widely reported in patients with adolescent idiopathic scoliosis (AIS) and has been implicated in deformity development. However, the underlying mechanism is largely unclear. LBX1 is one of the promising AIS predisposing genes validated by multicenter studies. PURPOSE: This study aimed to identify differentially expressed proteins (DEPs) relating to energy metabolism in AIS by using proteomic and metabolic analysis and to explore if the expression of these DEPs is associated with clinical parameters and modulated by LBX1. STUDY DESIGN: This is a cross-sectional study using clinical data and biological samples followed by basic study using a cellular model. PATIENT SAMPLE: Plasma samples were collected from Chinese girls with nonprogressive and progressive AIS (N=7 and 8, respectively) and age-matched healthy girls (N=50). Paraspinal muscle tissues were collected intraoperatively from concave and convex side of the apex of the major spinal curve in AIS (N=24) and either side from nonscoliosis patients (N=14). OUTCOME MEASURES: Radiological Cobb angle and basic anthropometric data of recruited subjects were measured. The DEPs and metabolites were compared in plasma using proteomics and metabolomics technique. The relative expression of selected genes was measured in muscles. METHODS: Plasma samples from AIS were collected at first clinical visit and were further divided into nonprogressive or progressive groups according to Cobb angle changes in 6-year follow-up. Age-matched healthy girls were recruited as control. High-performance liquid chromatography-mass spectrometry based proteomic analysis was carried out in three groups to identify DEPs and their annotated metabolic pathways. An independent cohort was used for validation by gas chromatography-mass spectrometry based metabolomic analysis. Paraspinal muscles were subjected to quantitative polymerase chain reaction (qPCR) followed by correlation analysis. Human skeletal muscle myoblast (HSMM) was used as the cellular model. RESULTS: The likelihood of aberrant galactose metabolism and glycolysis was found to be associated with AIS curve progression as evidenced by the thirteen DEPs and seven related metabolites according to proteomic and metabolomic analysis. Some of the DEPs showed significantly altered expression in AIS concave and convex sides paraspinal muscles compared with those in nonscoliosis control. Four DEPs were found significantly and negatively correlated with LBX1 in AIS convex side paraspinal muscles. Overexpressing LBX1 in HSMM cells led to increased expression of three DEPs and decreased expression of three DEPs, respectively. CONCLUSIONS: This is the first integrated proteomic and metabolomic analysis on AIS. Our findings show dysregulated galactose metabolism and glycolysis pathways in progressive group of AIS, suggesting the presence of abnormal energy metabolism at early stage of this disease, and their association with higher risk of progressing into more severe curvature. Evidence from ex vivo study with human muscle biopsies and in vitro study with human myoblast cells propose the possible effect of LBX1 on these two pathways in skeletal muscles. The present study provides new evidence of LBX1 function in AIS via modulating effect on the expression of energy metabolism related genes. This study might provide new insights into etiopathogenesis and development of novel treatment strategy targeting on abnormal body weight and BMI in patients with AIS. Additionally, the plasma proteomic and metabolomic studies suggested new candidates as biomarkers for establishing predictive model for AIS onset/progression.

Water Extract of Rhizoma Drynaria Selectively Exerts Estrogenic Activities in Ovariectomized Rats and Estrogen Receptor-Positive Cells.

Our previous study demonstrated that the bone protective actions of herbal medicine Rhizoma Drynariae (Gusuibu, RD) were mainly mediated by flavonoid phytoestrogens via estrogen receptors, raising concerns about the safety of using RD as it may induce estrogen-like risk-benefit profile and interact with other ER ligands, such as selective estrogen receptor modulators (SERMs), when coadministered. The present study evaluated the estrogenic activities of RD and its potential interaction with tamoxifen, a SERM, in estrogen-sensitive tissues by using mature ovariectomized (OVX) rats and ER-positive cells. Similar to but weaker than tamoxifen, RD at its clinical dose dramatically ameliorated OVX-induced changes in bone and dopamine metabolism-related markers in OVX rats. However, tamoxifen, but not RD, induced uterotrophic effects. No significant alteration in mammary gland was observed in OVX rats treated with RD, which was different from the inhibitory actions of tamoxifen. The two-way ANOVA results indicated the interactions between RD and tamoxifen in the bone, brain, and uterus of OVX rats while RD did not alter their responses to tamoxifen. Our results demonstrate that RD selectively exerts estrogenic actions in a different manner from tamoxifen. Moreover, RD interacts with tamoxifen without altering its effects in OVX rats.

Prenylated Isoflavonoids-Rich Extract of Erythrinae Cortex Exerted Bone Protective Effects by Modulating Gut Microbial Compositions and Metabolites in Ovariectomized Rats.

Flavonoids, found in a wide variety of foods and plants, are considered to play an important role in the prevention and treatment of osteoporosis. Our previous studies demonstrated that Erythrina cortex extract (EC) rich in prenylated isoflavonoids exerted bone protective effects in ovariectomized (OVX) rats. The present study aimed to investigate the interactions of gut microbiota with the EC extract to explore the underlying mechanisms involved in its beneficial effects on bone. Sprague-Dawley female rats of 3-months-old were ovariectomized and treated with EC extract for 12 weeks. EC extract reversed ovariectomy-induced deterioration of bone mineral density and bone microarchitecture as well as downregulated cathepsin K (Ctsk) and upregulated runt-related transcription factor 2 (Runx2) and alkaline phosphatase (ALP) in the tibia of OVX rats. Its protective effects on bone were correlated with changes in microbial richness and the restorations of several genera. EC increased the serum circulating levels of acetate and propionate in OVX rats. We conclude that the bone protective effects of EC extract were associated with the changes in microbial compositions and serum short chain fatty acids (SCFAs) in OVX rats.

Water extract of Er-xian decoction selectively exerts estrogenic activities and interacts with SERMs in estrogen-sensitive tissues.

ETHNOPHARMACOLOGICAL RELEVANCE: The increasing use of "kidney"-nourishing Traditional Chinese Medicine (TCM) like Er-xian decoction (EXD) for management of menopausal symptoms and osteoporosis has aroused concerns about their safety, and whether they interact with prescription drugs as both of them act via estrogen receptors (ERs) and regulate serum estradiol. AIM OF THE STUDY: The present study aimed to evaluate whether EXD selectively exerted estrogenic activities and interacted with Selective Estrogen Receptor Modulators (SERMs). MATERIALS AND METHODS: In vivo, mature ovariectomized (OVX) rats were administrated with EXD or combined treatment of EXD and SERMs for 12 weeks. The tissue-selective effect of EXD and its interaction of SERMs were studied in four estrogen sensitive tissues, bone, brain, breast and uterus. In vitro, the interaction of extracts of EXD-treated serum and SERMs in four ER-positive cell lines. RESULTS: In OVX rats, EXD selectively alleviated estrogen deficiency-induced changes in the bone and brain without inducing any estrogenic effects in the breast or uterus. Two-way ANOVA indicated the presence of interactions between EXD and SERMs in OVX rats but EXD did not significantly alter the tissue responses to SERMs in the bone, breast or brain. Indeed, the combined use of EXD and SERMs appeared to suppress the estrogenic effect of raloxifene and tamoxifen in the uterus. Extract of EXD-treated serum directly stimulated cell proliferation or differentiation in human osteosarcoma MG-63, neuroblastoma SHSY5Y, breast cancer MCF-7, and endometrial Ishikawa cells. Two-way ANOVA revealed that EXD-treated serum interacted with SERMs at various concentrations and altered the effects of tamoxifen in MG-63 and MCF-7 cells. CONCLUSIONS: EXD exerted estrogenic effects in a tissue-selective manner and interacted with SERMs. Combined treatment of EXD and SERMs did not hamper the beneficial effects of SERMs on the bone or brain but appeared to moderate the estrogenic effect of SERMs in the uterus.

The lignan-rich fraction from Sambucus Williamsii Hance ameliorates dyslipidemia and insulin resistance and modulates gut microbiota composition in ovariectomized rats.

Menopausal women are susceptible to have high risk of cardiovascular diseases, type II diabetes and osteoporosis due to the metabolic disorder caused by estrogen deficiency. Accumulating evidence supports that gut microbiota is a key regulator of metabolic diseases. Our previous metabolomics study interestingly demonstrated that the anti-osteoporotic effects of lignan-rich fraction (SWCA) from Sambucus wialliamsii Hance were related to the restoration of a series of lipid and glucose metabolites. This study aims to investigate how SWCA modulates lipid and glucose metabolism and the underlying mechanism. Our results show that oral administration of SWCA (140 mg/kg and 280 mg/kg) for 10 weeks alleviated dyslipidemia, improved liver functions, prevented glucose tolerance and insulin actions, attenuated system inflammation and improved intestinal barrier in OVX rats. It also induced a high abundance of Actinobacteria, and restored microbial composition. We are the first to report the protective effects of the lignan-rich fraction from S. williamsii on dyslipidemia and insulin resistance. Our findings provide strong evidence for the application of this lignan-rich fraction to treat menopausal lipid disorder and insulin resistance-related diseases.

Mass spectrometry-based untargeted metabolomics approach for differentiation of beef of different geographic origins.

Beef is a common staple food in many countries, and there is a growing concern over misinformation of beef products, such as false claims of origin, species and production methods. In this study, we used a mass spectrometry-based metabolomics approach to study the metabolite profiles of beef samples purchased from local retailers in Hong Kong. Using multivariate analysis, beef samples from different a) geographical origins, namely the United States (US), Japan and Australia, and b) feeding regimes could be differentiated. We identified twenty-four metabolites to distinguish beef samples from different countries, ten metabolites to identify Angus beef samples from others and seven metabolites to discriminate Australian beef produced by the organic farming from that produced using other farming modes. Based on results of this study, it is concluded that metabolomics provides an efficient strategy for tracing and authenticating beef products to ensure their quality and to protect consumer rights.

Schwann cell-specific Pten inactivation reveals essential role of the sympathetic nervous system activity in adipose tissue development.

There is increasing evidence that the sympathetic nervous system (SNS) plays an important role in adipose tissue development. However, the underlying molecular mechanism(s) associated with this remains unclear. SNS innervation of white adipose tissue (WAT) is believed to be necessary and sufficient to elicit WAT lipolysis. In this current study, mice with Schwann cell (SC)-specific inactivation of phosphatase and tensin homolog (Pten) displayed enlarged inguinal white adipose tissue (iWAT). This serendipitous observation implicates the role of SCs in mediating SNS activity associated with mouse adipose tissue development. Mice with SC-specific Pten inactivation displayed enlarged iWAT. Interestingly, the SNS activity in iWAT of SC-specific Pten-deficient mice was reduced as demonstrated by decreased tyrosine hydroxylase (TH) expression level and neurotransmitters, such as norepinephrine (NE) and histamine (H). The lipolysis related protein, phosphorylated hormone sensitive lipase (pHSL), was also decreased. As expected, AKT-associated signaling pathway was hyperactivated and hypothesized to induce enlarged iWAT in SC-specific Pten-deficient mice. Moreover, preliminary experiments using AKT inhibitor AZD5363 treatment ameliorated the enlarged iWAT condition in SC-specific Pten-deficient mice. Taken together, SCs play an essential role in the regulation of SNS activity in iWAT development via the AKT signaling pathway. This novel role of SCs in SNS function allows for better understanding into the genetic mechanisms of peripheral neuropathy associated obesity.

Selective Estrogen Receptor Modulator-Like Activities of Herba epimedii Extract and its Interactions With Tamoxifen and Raloxifene in Bone Cells and Tissues.

Herba epimedii (HEP), a kidney-tonifying herb, has been commonly used alone or in formula for strengthening kidney function and treating bone disorders. Its bone protective activity has been demonstrated to be via estrogen receptor (ERs). HEP activates the phosphorylation of ERα in an estrogen response element- (ERE-) dependent manner. We examined the bone protective effects of HEP and its potential interactions with Selective Estrogen Receptor Modulators (SERMs, such as tamoxifen and raloxifene) as they act via the same ERs. Six-month-old mature Sprague Dawley sham-operated (Sham) or ovariectomized (OVX) rats were treated with either vehicle, 17ß-estradiol (1.0 mg/kg.day), tamoxifen (Tamo, 1.0 mg/kg.day), raloxifene (Ralo, 3.0 mg/kg.day), HEP (0.16 g/kg.day), or its combinations with respective SERMs (HEP + Tamo; HEP + Ralo) for 12 weeks. HEP and SERMs as well as their combinations significantly restored changes in bone mineral density (BMD), trabecular bone properties, and bone turnover biomarkers induced by ovarian sex hormone deficiency in ovariectomized rats. Besides the increase in serum estradiol, inhibition on follicle stimulating hormone (FSH) might also be involved in the osteoprotective activities of HEP and SERMs. HEP interacted with SERMs to protect bones from ovarian sex hormone deficiency without altering SERMs' bone protective activities. HEP neither induced changes in uterus weight nor altered the uterotrophic activity of SERMs in OVX rats. In human osteosarcoma MG-63 cells, HEP-treated serum (HEP-Ts) significantly promoted alkaline phosphatase (ALP) activity like the crude HEP extract did but did not stimulate ERE activity. Our study also reported that biologically activated HEP interacted with SERMs to promote ALP activity without altering the action of SERMs at most of the concentrations tested in MG-63 cells. HEP exerted bone protective activity and the use of HEP did not alter the bone protective activities of SERMs when they were used simultaneously in an estrogen-deficient rat model.

Qualitative and quantitative analysis of sesquiterpene lactones in Centipeda minima by UPLC-Orbitrap-MS & UPLC-QQQ-MS.

An ultra-high performance liquid chromatography coupled with high-resolution Orbitrap mass spectrometry (UPLC-Orbitrap-MS) method has been used to identify sesquiterpene lactones in the methanolic extract of Centipeda minima. Fifteen sesquiterpene lactones were tentatively identified based on retention time and accurate mass of external standards or exact accurate mass searching (within 2 ppm) by comparison of some previous isolated sesquiterpene lactones. Meanwhile, a rapid, sensitive, precise, and reliable ultra-high performance liquid chromatography coupled with triple quadrupole mass spectrometry (UPLC-QQQ-MS) method has been developed to evaluate the quality of Centipeda minima through a simultaneous determination of five sesquiterpene lactones, namely brevilin A, arnicolide C, arnicolide D, microhelenin C, minimolide F. Chromatographic separation was achieved on a Waters Acquilty UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 µm) with a mobile phase consisting of a) 0.1% formic acid and b) a mixture of acetonitrile and methanol 50:50 v/v under an isocratic elution (42:58) manner. Positive electrospray ionization mode with multiple reaction monitoring was applied for the detection of the five sesquiterpene lactones. Method validation for linearity, accuracy and precision was also carried out. Finally, the method was successfully used for the analysis of 10 batches of Centipeda minima samples collected in China. Brevilin A and arnicolide D were the dominant sesquiterpene lactones in Centipeda minima and could be proposed as suitable markers for the quality control of Centipeda minima.

Dihydrochalcone-derived polyphenols from tea crab apple (Malus hupehensis) and their inhibitory effects on α-glucosidase in vitro.

Three dihydrochalcone-derived polyphenols, huperolides A-C (1-3), along with thirteen known compounds (4-16) were isolated from the leaves of Malus hupehensis, the well-known tea crab apple in China. Their chemical structures were elucidated by extensive spectroscopic analysis including NMR (HSQC, HMBC, 1H-1H COSY and ROESY), HRMS and CD spectra. Huperolide A is a polyphenol with a new type of carbon skeleton, while huperolides B and C are a couple of atropisomers, which were isolated from natural sources for the first time. The antihyperglycemic effects of the isolated compounds were evaluated based on assaying their inhibitory activities against α-glucosidase. As a result, phlorizin (4), 3-hydroxyphloridzin (5), 3-O-coumaroylquinic acid (12) and ß-hydroxypropiovanillone (15) showed significant concentration-dependent inhibitory effects on α-glucosidase. Therefore, those compounds might be responsible for the antihyperglycemic effect of this herb, and are the most promising compounds to lead discovery of drugs against diabetes.

HBx-K130M/V131I Promotes Liver Cancer in Transgenic Mice via AKT/FOXO1 Signaling Pathway and Arachidonic Acid Metabolism.

Chronic hepatitis B viral (HBV) infection remains a high underlying cause for hepatocellular carcinoma (HCC) worldwide, while the genetic mechanisms behind this remain unclear. This study elucidated the mechanisms contributing to tumor development induced by the HBV X (HBx) gene of predominantly Asian genotype B HBV and its common HBx variants. To compare the potential tumorigenic effects of K130M/V131I (Mut) and wild-type (WT) HBx on HCC, the Sleeping Beauty (SB) transposon system was used to deliver HBx Mut and WT into the livers of fumarylacetoacetate hydrolase (Fah)-deficient mice and in the context of transformation related protein 53 (Trp53) deficiency. From our results, HBx Mut had a stronger tumorigenic effect than its WT variant. Also, inflammation, necrosis, and fibrosis were evident in HBx experimental animals. Reduction of forkhead box O1 (FOXO1) with increased phosphorylation of upstream serine/threonine kinase (AKT) was detected under HBx Mut overexpression. Thus, it is proposed that HBx Mut enhances disease progression by reducing FOXO1 via phosphorylation of AKT. At the metabolomic level, HBx altered the expression of genes that participated in arachidonic acid (AA) metabolism, as a result of inflammation via accumulation of proinflammatory factors such as prostaglandins and leukotriene in liver. Taken together, the increased rate of HCC observed in chronic hepatitis B patients with K130M/V131I-mutated X protein, may be due to changes in AA metabolism and AKT/FOXO1 signaling. IMPLICATIONS: Our findings suggested that HBx-K130M/V131I-mutant variant promoted HCC progression by activating AKT/FOXO1 pathway and inducing stronger inflammation in liver via AA metabolism.

Altered metabolites in guinea pigs with allergic asthma after acupoint sticking therapy: New insights from a metabolomics approach.

BACKGROUND: Clinical evidence gathered in Chinese communities suggested that acupoint sticking therapy could be an alternative treatment for asthma-related diseases. However, its underlying mechanism is still poorly understood. AIM/HYPOTHESIS: In this study, we aimed to investigate the mechanism of the anti-inflammatory effect of acupoint sticking application with 'Treatment of Winter Disease in Summer' (TWDS) prescription by using metabolomics. METHODS: Allergic asthma in guinea pig was sensitized and challenged by ovalbumin (OVA). Histopathological evaluation of the lung tissue was performed by hematoxylin and eosin (H&E) staining and Masson's trichrome staining. The levels of Th2 cytokine and IgE level in serum were measured using enzyme-linked immunoassay (ELISA). The mRNA expression levels of IL-4, IL-5, IL-13 and orosomucoid-like 3 (ORMDL3) were measured using quantitative reverse transcription polymerase chain reaction (RT-qPCR). Proteins of NF-κB signaling pathway were measured using western blot. The serum metabolomics profiles were obtained by using ultra-performance liquid chromatography combined with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOF-MS). RESULTS: The overall results confirmed that AST with TWDS prescription had a significant protective effect against OVA-induced allergic asthma in guinea pig. This treatment not only attenuated airway inflammation and collagen deposition in the airway, but also decreased the levels of IL-4, IL-5, IL-13 and IgE in serum. In addition, metabolomics results indicated that metabolisms of phospholipid, sphingolipid, purine, amino acid and level of epinephrine were restored back to the normal control level. Moreover, results of the gene expression of ORMDL3 in lung tissues indicated that AST using TWDS could alter the sphingolipid metabolism. Further western blotting analysis also showed that its anti-inflammatory mechanism was by decreasing the phosphorylation of p65 and IκB. CONCLUSION: The study demonstrated that metabolomics provides a better understanding of the actions of TWDS acupoint sticking therapy on OVA-induced allergic asthma.

Clinical and histopathological features of chronic hepatitis B virus infected patients with high HBV-DNA viral load and normal alanine aminotransferase level: A multicentre-based study in China.

BACKGROUND: The aim of this study is to reveal the clinical and histopathological features of HBsAg-positive and HBeAg-positive chronic hepatitis B infected patients with high level of HBV DNA, from 17 hospitals and medical centres in China, with alanine aminotransferase levels within the lower region of normal range versus those with levels within the upper region of normal range and to investigate the clinical risk factors for the requirement of treatment through the examination of liver biopsy. METHODS: Liver biopsy was performed on high level of HBV DNA of 455 patients with HBsAg-positive and HBeAg-positive chronic hepatitis B infection and persistently normal alanine aminotransferase level. Liver necroinflammation and fibrosis were graded per the Knodell histological activity index and Ishak's fibrosis score, respectively. Univariate analysis of the clinical parameters versus necroinflammation and fibrosis was carried out. RESULTS: Of the subjects in this multicentre-based study, 5.49% and 10.11% had significant necroinflammation with Knodell histological activity index ≥ 9 and hepatic fibrosis stages with Ishak scores ≥ 3, respectively. The subjects were stratified into three age groups (30-39, 40-49 and ≥ 50 years), and our data clearly suggested that age, particularly in the age group over 50, was an independent predictor of liver necroinflammation and fibrosis. Lower HBV-DNA viral levels were found in patients with Knodell histological activity index ≥ 9 or advanced fibrosis (Ishak scores ≥ 3). CONCLUSION: Our results showed that histological changes in liver tissues were observed in a significant proportion of patients with persistently normal alanine aminotransferase level. According to the data evaluation results, liver biopsy is advisable for HBeAg-positive chronic hepatitis B infected patients aged older than 40 and high HBV-DNA viral load in China.

A Metabolomics Study on the Bone Protective Effects of a Lignan-Rich Fraction From Sambucus Williamsii Ramulus in Aged Rats.

The lignan-rich fraction (SWR) of Sambucus Williamsii Ramulus, a folk herbal medicine in China for treatment of bone diseases, has previously reported to exert protective effects on bone without exerting uterotrophic effects in ovariectomized (OVX) mice. The aim of the present study was to identify the potential metabolites and the associated metabolic pathways that contribute to the beneficial effects of SWR on bone in vivo. Aged female Sprague Dawley rats (9 months old) were either sham-operated or ovariectomized for 12 weeks, before receiving treatment for another 12 weeks with the following treatment groups (n = 12 each): vehicle (Sham), vehicle (OVX), Premarin (130 µg/kg) or low (57 mg/kg), medium (114 mg/kg), and high (228 mg/kg) doses of SWR. The results showed that SWRH significantly suppressed bone loss, improved bone micro-architecture and increased bone strength on tibia without stimulating uterus weight gain in OVX rats. Premarin exerted similar bone protective effects as SWRH but elicited uterotrophic effects in OVX rats. The metabolic profiles of serum samples were analyzed by using ultra-performance liquid chromatography quadrupole time-of flight mass spectrometry and gas chromatography time-of flight mass spectrometry, and the metabolites that were significantly altered were identified by multivariate statistical analysis. Our study indicated that SWRH effectively restored the changes of 26 metabolites induced by estrogen-deficiency in OVX rats, which related to lipids, amino acids, tryptophan metabolisms, and anti-oxidative system. A subsequent validation showed that the serum level of superoxide dismutase and catalase were indeed up-regulated, while the serotonin level in a tryptophan hydroxylase 1 (TPH1) high expressing cells (rats RBL-2H3 cells) was down regulated after treatment with SWR. The results also suggested that the gut-microbiota may play an important role on the bone protective effects of SWR. The current study provides insight for understanding the unique mechanism of actions of SWR that might be involved in achieving bone protective effects in vivo.

Rapid differentiation of Ganoderma species by direct ionization mass spectrometry.

In this study, direct ionization mass spectrometry (DI-MS) has been developed for rapid differentiation of Ganoderma (known as Lingzhi in Chinese), a very popular and valuable herbal medicine. Characteristic mass spectra can be generated by DI-MS directly from the raw herbal medicines with the application of a high voltage and solvents. Rapid differentiation of the Ganoderma species that are officially stated in the Chinese pharmacopoeia from easily confused Ganoderma species could be achieved based on this method, as the acquired DI-MS spectra showed that ganoderic acids, the major active components of Ganoderma, could be found only in the official Ganoderma species but not in the confused Ganoderma species. In addition, classification of wild and cultivated Ganoderma and potential differentiation of Ganoderma from different geographical locations could be accomplished based on principal component analysis (PCA) or hierarchical clustering analysis (HCA). The method is rapid, simple and reproducible, and can be further extended to analysis of other herbal medicines.

Comparative Evaluation of Chemical Profiles of Pyrrosiae Folium Originating from Three Pyrrosia Species by HPLC-DAD Combined with Multivariate Statistical Analysis.

Pyrrosiae Folium (PF) is a commonly used Chinese herb medicine originating from three Pyrrosia species for the treatment of urinary infection and urolithiasis. According to Chinese medicine practice, different specie origins led to some variations in the therapeutic effects of PF. To ensure the safety and efficacy of PF in clinical practice, it is necessary to establish a reliable and integrative method to distinguish PF occurring from the three species. In the present paper, a HPLC-DAD method was developed and applied to simultaneously analyze five major compounds in PF. Afterwards, multivariate statistical analyses including principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA) were applied for specie discrimination and integrative quality evaluation based on quantitative data. The chemical determination and pattern recognition results of 35 batches of PF samples indicated that PF samples from three species showed different chemical profiles and could be discriminated clearly. In conclusion, the present method is rapid and reliable for the quality assessment and species discrimination of PF.