Determination of aconitine,hypaconitine and mesaconitine in urine using hollow fiber liquid-phase microextraction combined with high-performance liquid chromatography

Hollow fiber liquid-phase microextraction (HF-LPME) coupled with high-performance liquid chromatography was used to simultaneously determine three Aconitum alkaloids, including aconitine (AC), hypaconitine (HA) and mesaconitine (MA) in human urine sample. Analytes were extracted from 5 mL urine sample containing 1.0 mmol/L NaOH into 1-octanol membrane phase impregnated in the pores of hollow fiber wall, and then back extracted into acidified aqueous solution in the lumen of the hollow fiber. After extraction, 10 μL of the acceptor phase was analyzed directly by HPLC. In this method, some important extraction parameters, such as organic solvent, extraction time, stirring rate, pH of donor phase and acceptor phase, temperature, and the volume of acceptor phase were optimized. This method provided 98- to 288-fold enrichment factors within 60 min of extraction and good repeatability with RSDs of 0.99–7.22%. The calibration curves were linear over the ranges of 16.0–128.0 μg/L for AC, 11.0–88.0 μg/L for HA and 8.1–64.8 μg/L for MA in human urine sample, with correlation coefficients of 0.9949, 0.9969 and 0.9904, respectively. Limits of detection were from 0.7 to 1.5 μg/L, and recoveries from spiked urine sample varied from 84.4% to 106.2% for AC, 77.3% to 85.6% for HA and 90.1% to 100.8% for MA.     

Quantitative proteomics study of breast cancer cell lines isolated from a single patient: Discovery of TIMM17A as a marker for breast cancer

The proteins involved in breast cancer initiation and progression are still largely elusive. To gain insights into these processes, we conducted quantitative proteomic analyses with 21T series of breast cell lines, which include a normal, primary tumor and a metastatic tumor that were isolated from a single patient. Stable isotope labeling of amino acid in cell culture followed by LC‐MS/MS analysis was performed and deregulated proteins were identified using statistical analysis. Gene ontology analysis revealed that proteins involved in metabolic processes were the most deregulated in both tumorigenesis and metastasis. Interaction network analysis indicated that ERBB2 signaling played a critical role in tumorigenesis. In addition to known markers such as ERBB2 and E‐cadherin, novel markers, including BRP44L, MTHFD2 and TIMM17A, were found to be overexpressed in 21T breast cancer cells and verified in additional breast cell lines. mRNA expression analysis as well as immunohistochemistry analysis in breast cancer tissues indicated that expression level of TIMM17A was directly correlated with tumor progression, and survival analysis suggested that TIMM17A was a powerful prognosis factor in breast cancer. More interestingly, overexpression and siRNA knockdown experiments indicated an oncogenic activity of TIMM17A in breast cancer. Our study provides a list of potential novel markers for breast cancer tumorigenesis and metastasis using a unique cell model. Further studies on TIMM17A as well as other markers on the list may reveal mechanisms that result in more effective therapeutics for cancer treatment.     

Chemiluminescent immunoassay of thyroxine enhanced by microchip electrophoresis

A homogeneous chemiluminescent immunoassay of thyroxine (T4) enhanced by microchip electrophoresis separation has been developed. The method deployed the competitive immunoreaction of T4 and horseradish peroxidase (HRP)-labeled T4 (HRP-T4) with anti-T4 mouse monoclonal antibody (Ab). HRP-T4 and the HRP-T4-Ab complex were separated and quantified by using microchip electrophoresis (MCE) with chemiluminescence (CL) detection. Highly sensitive CL detection was achieved by means of HPR-catalyzed luminol-H₂O₂ reaction. Due to the effective MCE separation, the CL analytical signal was less prone to sample matrix interference. Under the selected assay conditions, the MCE separation was accomplished within 60 s. The linear range for T4 was 5-250 nM with a detection limit of 2.2 nM (signal/noise ratio = 3). The current method was successfully applied for the quantification of T4 in human serum samples. It was demonstrated that the current MCE-CL-enhanced competitive immunoassay was quick, sensitive, and highly selective. It may serve as a tool for clinical analysis of T4 to assist in the diagnosis of thyroid gland functions.     

Rafetus swinhoei名称的历史考证与中文名更改为“黄斑巨鳖”的建议

Rafetusswinhoei是近年来才引起科学和保护界关注的极危物种,关于此物种的分类和命名,长期存在分歧和混淆。简要回顾了对R．swinhoei的科学认识过程,并对古籍进行考证后认为,中国古代所说的“鼋”,本应指此种而非Pdochelyscantorii。鉴于R．swinhoei已有的诸多中文名各自存在一定缺陷．建议采用中文名“黄斑巨鳖”。     

Conformational rearrangements to the intracellular open states of the LeuT and ApcT transporters are modulated by common mechanisms

Recent crystallographic studies revealed that five transporter families without much sequence similarities among them have similar structure folds to LeuT, a bacterial neurotransmitter:sodium symporter homolog. The LeuT fold is characterized by an internal twofold structural pseudosymmetry. The transport cycle of some members of each of these families is dependent on a sodium gradient across the membrane, whereas in some others the role of sodium is mimicked by proton. We report on the identification of common structure-dynamics elements of the transporters with LeuT fold, which are recognizable in the conformational transitions related to function. The findings from comparative computational modeling and simulation studies of LeuT, and ApcT from the amino acid-polyamine-organocation transporter family define the intramolecular mechanisms by which Na⁺ binding couples to the transport process, and single out the lead/active role of TM1a in the transition to inward-open conformation. These mechanistic insights are derived in the context of collaborative investigations of LeuT dynamics with both single-molecule fluorescence and simulations that have produced excellent agreement of the dynamic details, and are found to be generalizable across the transporter families and to transcend sequence and motif similarities.     

The phylogeny of Orthoptera inferred from mtDNA and description of Elimaea cheni （Tettigoniidae： Phaneropterinae） mitogenome

The complete 15,831 bp nucleotide sequence of the mitochondrial genome from Elimaea cheni(Phaneropterinae)was determined.The putative initiation codon for cox1 was TTA.The phylogeny of Orthoptera based on different mtDNA datasets were analyzed with maximum likelihood(ML)and Bayesian inference(BI).When all 37 genes(mtDNA)were analyzed simultaneously,the monophyly of Caelifera and Ensifera were recovered in the context of our taxon sampling.The phylogeny of Orthoptera was largely consistent with previous phylogenetie hypotheses.Rhaphidophoridae to be a sister group of Tettigoniidae,and the relationships among four subfamilies of Tettigoniidae were(Phaneropterinae+(Conocephalinae+(Bradyporinae+Tettigoniinae))).Pyrgomorphidae was the most basal group of Caelifera.The relationships among six acridid subfamilies were(Oedipodinae+(Acridinae+(Gomphocerinae+(Oxyinae+(Calliptaminae +Cyrtacanthaeridinae))))).However,we did not recover a monophyletic Grylloidea.Myrmecophilidae clustered into one clade with Gryllotalpidae instead of with Gryllidae.ML and BI analyses of all protein coding genes(using all nucleotide sequence data or excluding the third codon position,and amino acid sequences)revealed a topology identical to that of the entire mtDNA genome dataset.However,22 tRNAs genes excluding the DHU loop and T()C loop(TRNA),and two rRNA genes(RRNA)perform poorly when analyzed as single dataset.Our results suggest that the best phylogenetie inferences were ML and BI methods based on total mtDNA.Excluding tRNA genes,rRNA genes and the third codon position of protein coding genes from dataset and converting nucleotide sequences to amino acid sequences do not positively affect phylogenetic reconstruction.     

Short exposure to paclitaxel induces multipolar spindle formation and aneuploidy through promotion of acentrosomal pole assembly

Paclitaxel is a widely used microtubule drug and cancer medicine. Here we report that by short exposure to paclitaxel at a low dose, multipolar spindles were induced in mitotic cells without centrosome amplification. Both TPX2 depletion and Aurora-A overexpression antagonized the multipolarity. Live cell imaging showed that some paclitaxel-treated cells accomplished multipolar cell division and a portion of the daughter cells went on to the next round of mitosis. The surviving cells grew into clones with varied genome content. The results indicated that an aneuploidy population could be induced by short exposure to paclitaxel at a low dose, implicating potential side effects of paclitaxel.     

Co-expression module analysis reveals biological processes,genomic gain,and regulatory mechanisms associated with breast cancer progression

Background  

Gene expression signatures are typically identified by correlating gene expression patterns to a disease phenotype of interest. However, individual gene-based signatures usually suffer from low reproducibility and interpretability.     

Utilizing elementary mode analysis,pathway thermodynamics,and a genetic algorithm for metabolic flux determination and optimal metabolic network design

Background  

Microbial hosts offer a number of unique advantages when used as production systems for both native and heterologous small-molecules. These advantages include high selectivity and benign environmental impact; however, a principal drawback is low yield and/or productivity, which limits economic viability. Therefore a major challenge in developing a microbial production system is to maximize formation of a specific product while sustaining cell growth. Tools to rationally reconfigure microbial metabolism for these potentially conflicting objectives remain limited. Exhaustively exploring combinations of genetic modifications is both experimentally and computationally inefficient, and can become intractable when multiple gene deletions or insertions need to be considered. Alternatively, the search for desirable gene modifications may be solved heuristically as an evolutionary optimization problem. In this study, we combine a genetic algorithm and elementary mode analysis to develop an optimization framework for evolving metabolic networks with energetically favorable pathways for production of both biomass and a compound of interest.