Metabolic map of osthole and its effect on lipids.

1. Osthole, a coumarin compound from plants, is a promising agent for the treatment of metabolic diseases, including hyperglycemia, fatty liver, and cancers. Studies indicate that the peroxisome proliferator-activated receptors (PPAR) α and γ are involved in the pharmacological effects of osthole. The in vitro and in vivo metabolism of osthole and its biological activity are not completely understood. 2. In this study, ultra-performance chromatography electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOFMS)-based metabolomics was used to determine the metabolic pathway of osthole and its influence on the levels of endogenous metabolites. Forty-one osthole metabolites, including 23 novel metabolites, were identified and structurally elucidated from its metabolism in vitro and in vivo. Recombinant cytochrome P450s (CYPs) screening showed that CYP3A4 and CYP3A5 were the primary enzymes contributing to osthole metabolism. 3. More importantly, osthole was able to decrease the levels of lysophosphatidylethanolamine (LPE) and lysophosphatidylcholine (LPC) in the plasma, which explains in part its modulatory effects on metabolic diseases. 4. This study gives the insights about the metabolic pathways of osthole in vivo, including hydroxylation, glucuronidation, and sulfation. Furthermore, the levels of the lipids regulated by osthole indicated its potential effects on adipogenesis. These data contribute to the understanding of the disposition and pharmacological activity of osthole in vivo.

Simulated Gastric Acid Promotes the Horizontal Transfer of Multidrug Resistance Genes across Bacteria in the Gastrointestinal Tract at Elevated pH Levels.

The assessment of factors that can promote the transmission of antibiotic resistance genes (ARGs) across bacteria in the gastrointestinal tract is in great demand to understand the occurrence of infections related to antibiotic-resistant bacteria (ARB) in humans. However, whether acid-resistant enteric bacteria can promote ARG transmission in gastric fluid under high-pH conditions remains unknown. This study assessed the effects of simulated gastric fluid (SGF) at different pH levels on the RP4 plasmid-mediated conjugative transfer of ARGs. Moreover, transcriptomic analysis, measurement of reactive oxygen species (ROS) levels, assessment of cell membrane permeability, and real-time quantitative assessment of the expression of key genes were performed to identify the underlying mechanisms. The frequency of conjugative transfer was the highest in SGF at pH 4.5. Antidepressant consumption and certain dietary factors further negatively impacted this situation, with 5.66-fold and 4.26-fold increases in the conjugative transfer frequency being noted upon the addition of sertraline and 10% glucose, respectively, compared with that in the control group without any additives. The induction of ROS generation, the activation of cellular antioxidant systems, increases in cell membrane permeability, and the promotion of adhesive pilus formation were factors potentially contributing to the increased transfer frequency. These findings indicate that conjugative transfer could be enhanced under certain circumstances in SGF at elevated pH levels, thereby facilitating ARG transmission in the gastrointestinal tract. IMPORTANCE The low pH of gastric acid kills unwanted microorganisms, in turn affecting their inhabitation in the intestine. Hence, studies on the factors that influence antibiotic resistance gene (ARG) propagation in the gastrointestinal tract and on the underlying mechanisms are limited. In this study, we constructed a conjugative transfer model in the presence of simulated gastric fluid (SGF) and found that SGF could promote the dissemination of ARGs under high-pH conditions. Furthermore, antidepressant consumption and certain dietary factors could negatively impact this situation. Transcriptomic analysis and a reactive oxygen species assay revealed the overproduction of reactive oxygen species as a potential mechanism by which SGF could promote conjugative transfer. This finding can help provide a comprehensive understanding of the bloom of antibiotic-resistant bacteria in the body and create awareness regarding the risk of ARG transmission due to certain diseases or an improper diet and the subsequent decrease in gastric acid levels.

A rapid identification method for common astigmatid species based on multiplex polymerase chain reaction.

Astigmatid mites are economically significant pests of stored products and sources of inhalant allergens causing allergic rhinitis and asthma worldwide. The morphological identification of astigmatid mites at the species level is often a difficult task due to their small size, phenotypic similarity and lack of diagnostic characters. We used multiplex polymerase chain reaction (PCR) to identify astigmatid mite species, which could complement the morphological data for the species-specific identification of mites. Internal ribosomal transcribed spacer (ITS) sequences (i.e., partial 18S, the full length of ITS1-5.8S-ITS2 and partial 28S) from eight astigmatid species (Acarus siro, Tyrophagus putrescentiae, Suidasia nesbitti, Dermatophagoides pteronyssinus, Dermatophagoides farinae, Lepidoglyphus destructor, Chortoglyphus arcuatus and Gohieria fuscus) were obtained by DNA extraction and then sequenced after PCR amplification. Specific primers were designed in the ITS2 region manually. Results revealed that an identification method for eight common astigmatid species was established based on multiplex PCR, which should be effective for the identification of other species of mites by redesigning species-specific primers in future experiments.

Absorption, distribution, metabolism, and excretion of [14C]Mefuparib (CVL218), a novel PARP1/2 inhibitor, in rats.

INTRODUCTION: Mefuparib (CVL218) is a novel second-generation poly-ADP-ribose polymerase (PARP) inhibitor for cancer treatment. CVL218 can easily enter the brain. However, the transport mechanism by which CVL218 crosses the blood-brain barrier (BBB) is unknown. METHODS: (1) [14C] CVL218 metabolism in rats was traced by a liquid scintillation counter and oxidative combustion. (2) Metabolic profiles and metabolites were identified by UHPLC-ß-RAM/UHPLC-Fraction Collector and UHPLC-Q Exactive Plus MS. (3) The partition coefficient Kp,uu,brain value was simulated by two strategies. One strategy was using ACD and GastroPlus Software based on the results of intravenous administration pharmacokinetics and plasma protein-binding studies. The reliability was confirmed by comparison with another strategy (brain/plasma distribution study). RESULTS: (1) Rapid drug elimination was observed 24 h after intragastric administration. The total cumulative excretion in urine and feces within 168 h accounted for 97.15% of the dose. The cumulative radioactive dose recovery in bile was 41.87% within 72 h. The drug-related substances were extensively distributed to the tissues within 48 h. (2) M8 was the major metabolite in plasma, urine, feces and bile. (3) CVL218 exhibited high brain protein-binding rate (88.16%). The Kp,uu,brain value (8.42) simulated by the simple software strategy was similar to that of the brain/plasma distribution study (7.01). CONCLUSIONS: CVL218 is a fast-metabolizing drug and is mainly excreted in feces. The B/P ratio prediction and observation data for CVL218 were consistent. Furthermore, the Kp,uu,brain value indicated that penetration through the BBB might be mediated by uptake transporters.

BRCA1-Associated Protein Is a Potential Prognostic Biomarker and Is Correlated With Immune Infiltration in Liver Hepatocellular Carcinoma: A Pan-Cancer Analysis.

BACKGROUND: BRCA1-associated protein (BRAP) is a critical gene that regulates inflammation-related signaling pathway and affects patients' prognosis in esophageal squamous cell carcinoma (ESCC). However, its roles in different cancers remain largely unknown. METHODS: BRAP expression in human pan-cancer was analyzed via the Genotype-Tissue Expression (GTEx) and The Cancer Genome Atlas (TCGA) database. Pearson correlation analysis was used to evaluate the association between BRAP expression with mismatch repair (MMR) gene mutation and DNA methyltransferase. We evaluated the influence of BRAP on clinical prognosis by univariate survival analysis. Moreover, the correlation between BRAP and tumor immune infiltration was analyzed via the Tumor Immune Evaluation Resource (TIMER) database. Pearson correlation analysis was used to investigate the correlation between BRAP expression and immune checkpoint genes expression. RESULTS: BRAP is abnormally overexpressed and significantly correlated with MMR gene mutation level and DNA methyltransferase expression in human pan-cancer. Univariate survival analysis showed that BRAP was significant with patients' overall survival (OS) in six cancer types, disease-free interval (DFI) in three cancer types, and progression-free interval (PFI) in two cancer types. Remarkably, increased BRAP expression was strongly correlated with patients' poor prognosis in liver hepatocellular carcinoma (LIHC), whether OS (P < 0.0001, hazard ratio (HR) = 1.1), DFI (P = 0.00099, HR = 1.06), or PFI (P = 0.00025, HR = 1.07). Moreover, a positive relationship was found between BRAP expression and immune infiltrating cells including B cell, CD4 + T cell, CD8 + T cell, dendritic cell, macrophage cell, and neutrophil cell in colon adenocarcinoma (COAD), kidney renal clear cell carcinoma (KIRC), and LIHC. Additionally, BRAP expression showed strong correlations with immune checkpoint genes in LIHC. CONCLUSION: BRAP expression is increased in human pan-cancer samples compared with normal tissues. Overexpression of BRAP is correlated with poor prognosis and immune infiltration in multiple cancers, especially in LIHC. These findings suggest that BRAP may be used as a potential molecular biomarker for determining prognosis and immune infiltration in LIHC.

Genome-wide analysis of prognostic-related lncRNAs, miRNAs and mRNAs forming a competing endogenous RNA network in lung squamous cell carcinoma.

PURPOSE: As a type of cancer with the highest morbidity and mortality, lung squamous cell carcinoma (LUSC) has a very poor prognosis. Long-non-coding RNA (lncRNA) has recently attracted attentions because it can play the role of competing endogenous RNA (ceRNA) to inhibit microRNA (miRNA) functions. In this study, we aimed to find prognosis-related lncRNAs, miRNAs and mRNAs and construct a prognosis-related ceRNA network. METHODS: The original LUSC RNA-sequencing data and miRNA profiles data were downloaded from the cancer genome atlas (TCGA) database. Differentially expressed lncRNAs, miRNAs and mRNAs were then identified between patients with lymph node metastasis and no lymph node metastasis. Univariate Cox regression analysis was performed to find the survival-associated lncRNAs, miRNAs and mRNAs. Subsequently, prognostic-related ceRNA network was established. By multivariate Cox regression analysis, three lncRNA signatures and three mRNA signatures were developed and used for predicting LUSC patients' survival. RESULTS: A total of 224 lncRNAs, 160 miRNAs, 913 mRNAs were identified between samples with lymph node metastasis and no lymph node metastasis. Univariate Cox regression analysis showed that, among them, 28 lncRNAs, 8 miRNAs, 105 mRNAs were significantly associated with patients' overall survival time. Further pathway and enrichment analysis suggested that these mRNAs were associated with the regulation of transmembrane transport, regulation of blood circulation, plasma lipoprotein particle organization. Then we constructed a survival-related ceRNA network including 9 lncRNAs, 8 miRNAs and 23 mRNAs. Additionally, a multivariate Cox regression analysis demonstrated that three lncRNAs (AL161431.1, LINC02389, APCDD1L.DT) and three mRNAs (KLK6, SLITRK5, CCDC177) had a significant prognostic value. Risk score indicated that lncRNA signature and mRNA signature could independently predict overall survival in LUSC patients. CONCLUSION: The current study provided a better understanding of the ceRNA network in the progression of LUSC and laid a theoretical foundation for LUSC prognosis.

Influences of different developmental periods of taurine supplements on synaptic plasticity in hippocampal CA1 area of rats following prenatal and perinatal lead exposure.

BACKGROUND: Previous study has demonstrated that dietary taurine supplement protected rats from impairments of synaptic plasticity induced by postnatal lead exposure. However, little is known about the role of taurine in the presence of prenatal and perinatal lead exposure. We investigated the possible effect of taurine supplement on prenatal and perinatal lead-induced synaptic plasticity deficit and determined developmental periods critical for the effect of taurine. RESULTS: In the present study, taurine was administrated to prenatal and perinatal lead-exposed rats in different developmental periods: from prenatal to weaning (Lead+PW-Tau), from weaning to life (Lead+WL-Tau), and from prenatal to life (Lead+PL-Tau). We examined the input-output (I/O) function, paired-pulse facilitation (PPF) and the long-term potentiation (LTP) of field excitatory postsynaptic potential (fEPSP) in the hippocampal CA1 area of rats on postnatal days 18-25 (P18-25) or days 60-75 (P60-75). We found that (1) on P18-25, taurine had no evident effect on I/O functions and PPF ratios of lead-exposed rats but caused a 12.0% increase in the LTP amplitudes of these animals; (2) on P60-75, taurine significantly elevated lead depressed I/O functions and PPF ratios in Lead+PW-Tau and Lead+PL-Tau rats, but failed in Lead+WL-Tau rats. The amplitudes of LTP of lead-exposed rats were all significantly increased by additional taurine supplement in any developmental period compared with untreated rats. Thus, taurine appeared to have the most effect during the prenatal and lactation periods and its effects on younger rats would not be manifest until the adult life; and (3) the level of lead deposition in hippocampus was evidently reduced by additional treatment of taurine in lead-exposed rats, compared with untreated rats. CONCLUSION: Taurine supplement can protect the adult rats from synaptic plasticity deficits following prenatal and perinatal lead exposure, and the protective effects are critical for the prenatal and lactation periods of lead-exposed rats.

Inhibitory effect of Cd2+ on glycine-induced chloride current in rat hippocampal neurons.

The effects of cadmium (Cd(2+)) on glycine-induced Cl(-) current (I(Gly)) were investigated in acutely dissociated rat hippocampal CA1 neurons using the conventional whole-cell patch-clamp technique in this study. We found that Cd(2+) reversibly and concentration-dependently, reduced the amplitudes of I(Gly), with an IC(50) of 1.27 mM and Hill coefficient of 0.45. The depression of I(Gly) by Cd(2+) was independent of membrane voltage between -60 and +40 mV and did not involve a shift in the reversal potential of the current. A non-competitive inhibition was suggested by a double reciprocal plot of the effects of Cd(2+) on the concentration-response curve of the I(Gly). Since intracellular dialysis with 3mM Cd(2+) failed to modify I(Gly), it was suggested that the site of action of Cd(2+) is extracellular. The suppression of I(Gly) by Zn(2+) was unaffected by 3mM Cd(2+), which indicated that Zn(2+) and Cd(2+) bind to independent sites on glycine receptor. The results show that Cd(2+) decreases I(Gly) in acutely dissociated rat hippocampal neurons and the depression of I(Gly) by Cd(2+) may contribute to worsen the neurotoxicological impairment.

Inhibition of delayed rectifier K+ currents by copper in acutely dissociated rat hippocampal CA1 neurons.

A growing number of research results demonstrate that copper is an important trace element to life. In this study, whole-cell recording made from acutely dissociated rat hippocampal CA1 neurons was employed to investigate the actions of copper (Cu(2+)) on the delayed rectifier K(+) currents (I(K)). External application of various concentrations of Cu(2+) (1-1000microM) reduced the amplitude of I(K) in a dose-dependent manner with an IC(50) value of 100microM and a Hill coefficient of 0.4. 300microM of Cu(2+) depolarized the I(K) activation curves by 12.5mV and hyperpolarized the I(K) state-inactivation curves by 17.4mV, respectively. At this concentration, Cu(2+) also significantly increased the value of the fast decay time constant (tau(1)), but had no effect on the I(K) recovery from inactivation. These results suggest that relevant concentrations of copper at physiological and pathological level can influence the neuronal excitability of rat hippocampal CA1 neurons by voltage-gated delayed rectifier K(+) channels, and such actions are likely involved in the pathophysiology of Cu-related Wilson's disease.

[Trial study on DENG Tie-tao's coronary heart disease capsules in improving patients' quality of life].

OBJECTIVE: To assess the efficacy of the Coronary Heart Disease (CHD) Capsules worked out by Prof. Deng --in improving quality of life of CHD patients of qi deficiency with phlegm and blood stasis syndrome. METHOD: According to the WHO's diagnosis criteria of CHD, a total of 93 stable angina patients were divided into 3 groups using the single blinded method. The groups were evenly distributed into CHD Capsule treated group (CHDC), isosorbide dinitrate control group (ID), and Compound Prescription Danshen Droplet Pills control group (CPDDP). Two courses of treatment lasting for 6 months were given. During the courses of treatment, the following parameters were observed: clinical symptoms of angina pectoris, ECG change, treadmill exercise test, 36 items in short form of health survey (SF-36) and Seattle Angina Questionnaire (SAQ) scale. RESULTS: After 6 months of treatment, all the three groups showed good curative effect in angina pectoris, ECG and treadmill exercise test, differences between them had no statistical significance. The CHDC group showed a better result in nitro-glycerine stopping or alleviation rate and in improving symptoms than the other groups (P < 0.05). The general health, vitality, role-emotional, mental health and reported health transition in the CHDC group were significantly better than those in the control groups (P < 0.05). The scores in physiological functioning role, physiological function and pain alleviation were not different among the three groups. CONCLUSION: Prof. DENG Tie-tao's CHDC is effective in treating CHD with qi deficiency, phlegm and blood stasis and also in improving the quality of life. CHDC is more suitable to be used in long-term treatment than isosorbide dinitrate. The SF-36 and SAQ can be used to appraise the curative effect of traditional Chinese medicine agents for CHD angina pectoris.

[Optical properties of aerosol during haze-fog episodes in Beijing].

The purpose of this study is to investigate the optical properties of aerosol during haze-fog episodes in Beijing. The aerosol optical depth (AOD), Angstrom exponent (alpha), size distribution and single scattering albedo (omega) during haze-fog episodes were analyzed between 2002 and 2008 using AERONENT data. During haze-fog episodes, the aerosol optical depth showed a decreasing trend with wavelengths, and showed high values with an average 1.34 at 440 nm. The magnitude of Angstrom exponent was relatively high during haze-fog episodes and the mean values reached 1.11. The frequency distribution of alpha was up to 94% when alpha > 0.9, indicating the predominance of fine particles during haze-fog episodes in Beijing. The aerosol volume size distributions presented a bimodal structure (fine and coarse modes). The maxima (peaks) radius of fine mode showed an increasing trend with AOD, however, those of coarse mode showed a decreasing trend with AOD. The size distribution showed a distinct difference in dominant mode for the different AOD. The single scattering albedo showed an increasing trend with AOD during haze-fog episodes in Beijing. The mean value of omega was 0.89 at the four wavelengths and the omega exhibited a low sensitivity to wavelengths.

Effect of Huxin Formula on reverse cholesterol transport in ApoE-gene knockout mice.

OBJECTIVE: To observe the effect of Huxin Formula on expressions of the chief reverse cholesterol transport (RCT) associated genes, caveolin-1 and scavenger receptor-BI (SR-BI) in ApoE-gene knockout [ApoE (-/-)] mice. METHODS: Thirty ApoE (-/-) mice of 4-6 weeks old were randomly divided into three groups (A-C). After being fed with high-fat diet for 16 weeks, they were treated with HXF (1 mL/100 g), pravachol (0.3 mg/100 g), and saline in equal volume respectively for 16 weeks successively; in addition, a blank group was set up with 10 C57BL/6J mice of 6-week old received 16-week high-fat feeding and saline treatment. Animals were sacrificed at the termination of the experiment, their paraffin sections of aortic tissue were used to measure the size of plaque, expressions of cavolin-1 and SR-BI were detected by immunological histochemical method. RESULTS: As compared with the blank group, levels of caveolin-1 and SR-BI were increased in Groups A and B (P<0.01); but the increase in Group A was more significant than that in Group B (P<0.05). The plaque/aorta area ratio decreased significantly in Groups A and B, but showed insignificant difference between the two groups. CONCLUSION: HXF could obviously increase the expressions of RCT associated genes, caveolin-1 and SR-BI, promote the RCT process, so as to reduce the formation of aorta atherosclerotic plaque in ApoE (-/-) mice.

JNK1 contributes to metabotropic glutamate receptor-dependent long-term depression and short-term synaptic plasticity in the mice area hippocampal CA1.

Several recent reports implicate an important role played by c-Jun N-terminal kinases (JNKs) in long-term potentiation (LTP). However, little is known about how the isoforms of JNKs participate in synaptic plasticity. Here we showed that short-term synaptic plasticity was impaired in the hippocampal area CA1 of JNK1-deficient (JNK1-/-) mice; these mice showed normal LTP in response to a strong tetanus and no alteration of N-methyl-D-aspartate receptor-dependent long-term depression (LTD) in the hippocampus. However, LTD induced either by group I metabotropic glutamate receptors (mGluRs) agonist dihydroxyphenylglycine or by paired-pulse low-frequency stimulation was absent in both the JNK1-/- slices and in JNK inhibitor anthrax [1, 9-cd] pyrazol-6(2H)-1 (SP600125)-pretreated slices. Induction of mGluR-dependent LTD resulted in an increase in phosphorylation of JNK1 substrates, including p-c-Jun and p-ATF2 in wild-type (WT) mice, and these increases failed to occur in the JNK1-/- or SP600125-pretreated mice. These results demonstrated that JNK1 played a crucial role in the short-term synaptic plasticity and mGluR-dependent LTD, whereas hippocampus LTP was not affected by JNK1 deficiency.

[Different sensitivity of two chronic myeloid leukemia cell lines to interferon-alpha].

BACKGROUND & OBJECTIVE: The clinical observation has shown that interferon-alpha(IFN-alpha) is one of the most effective therapeutic agents for the malignancies of hemopoietic system and lymphoma. However, IFN-alpha can only induce about 70-80% of the patients with chronic myeloid leukemia (CML) to get hematological remission. The mechanism by which various CML cases respond differently to IFN-alpha is still unclear. METHODS: (1)The effects of IFN-alpha in different concentrations (100, 500, 1,000, 5,000, and 10,000 U/ml) on growth of the two CML cell lines were detected by MTT assay,semisolid colony formation and trypan-blue staining for the cells in liquid culture. (2)The cell apoptosis was examined by flow cytometry(FCM),fluorescence microscopy and gel electrophoresis analysis for DNA fragmentation in 48 hours after IFN-alpha (1,000 U/ml) induction of both KT-1/A3 and K562 cells. (3)The expression levels of bcr/abl chimeric genes were analyzed by relative quantitative RT-PCR at 48 hours after cultivation of both KT-1/A3 and K562 cells with IFN-alpha in 1,000 U/ml. RESULTS: (1)The growth inhibition of KT-1/A3 cells was dose-dependent in IFN-alpha from the concentration of 100 U/ml to 10,000 U/ml. (2)Having been induced with IFN-alpha in 1000 U/ml for 48 hours, the apoptosis rate of KT-1/A3 cells raised from 3.29% to 11.8% and the expression level of bcr/abl chimeric gene in this cell line declined to 66.7% as compared with those of the control. (3)The growth and apoptosis rate as well as bcr/abl gene expression level of K562 cells were not significantly affected by IFN-alpha. CONCLUSION: Different populations of CML cells shows different sensitivity to IFN-alpha.