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N. Cesbron A.-L. Royer Y. Guitton A. Sydor B. Le Bizec G. Dervilly-Pinel 《Metabolomics : Official journal of the Metabolomic Society》2017,13(8):99
Introduction
Collecting feces is easy. It offers direct outcome to endogenous and microbial metabolites.Objectives
In a context of lack of consensus about fecal sample preparation, especially in animal species, we developed a robust protocol allowing untargeted LC-HRMS fingerprinting.Methods
The conditions of extraction (quantity, preparation, solvents, dilutions) were investigated in bovine feces.Results
A rapid and simple protocol involving feces extraction with methanol (1/3, M/V) followed by centrifugation and a step filtration (10 kDa) was developed.Conclusion
The workflow generated repeatable and informative fingerprints for robust metabolome characterization.3.
Yun Yang Rui Guo Qi Chen Youxun Liu Pengfei Zhang Ziheng Zhang Xi Chen Tianyun Wang 《Biotechnology letters》2018,40(5):789-795
Objective
To promote targeting specificity of anti-CD47 agents, we have constructed a novel bispecific antibody fusion protein against EGFR and CD47, which may minimize the “off-target” effects caused by CD47 expression on red blood cells.Results
The novel bispecific antibody fusion protein, denoted as Bi-SP could simultaneously bind to EGFR and CD47 and exhibited potent phagocytosis-stimulation effects in vitro. Bi-SP treatment with a low dose more effectively inhibited tumor growth than either EGFR-targeting antibody, Pan or the SIRPα variant-Fc (SIRPαV-Fc) in the A431 xenograft tumor model. In addition, the treatment with Bi-SP produced less red blood cell (RBC) losses than the SIRPαV-Fc treatment, suggesting its potential use for minimizing RBC toxicity in therapy.Conclusions
Bi-SP with improved therapeutic index has the potential to treat CD47+ and EGFR+ cancers in clinics.4.
Renato de Souza Pinto Lemgruber Kaspar Valgepea Mark P. Hodson Ryan Tappel Sean D. Simpson Michael Köpke Lars K. Nielsen Esteban Marcellin 《Metabolomics : Official journal of the Metabolomic Society》2018,14(3):35
Introduction
Quantification of tetrahydrofolates (THFs), important metabolites in the Wood–Ljungdahl pathway (WLP) of acetogens, is challenging given their sensitivity to oxygen.Objective
To develop a simple anaerobic protocol to enable reliable THFs quantification from bioreactors.Methods
Anaerobic cultures were mixed with anaerobic acetonitrile for extraction. Targeted LC–MS/MS was used for quantification.Results
Tetrahydrofolates can only be quantified if sampled anaerobically. THF levels showed a strong correlation to acetyl-CoA, the end product of the WLP.Conclusion
Our method is useful for relative quantification of THFs across different growth conditions. Absolute quantification of THFs requires the use of labelled standards.5.
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Thijs Welle Anna T. Hoekstra Ineke A. J. J. M. Daemen Celia R. Berkers Matheus O. Costa 《Metabolomics : Official journal of the Metabolomic Society》2017,13(7):83
Introduction
Swine dysentery caused by Brachyspira hyodysenteriae is a production limiting disease in pig farming. Currently antimicrobial therapy is the only treatment and control method available.Objective
The aim of this study was to characterize the metabolic response of porcine colon explants to infection by B. hyodysenteriae.Methods
Porcine colon explants exposed to B. hyodysenteriae were analyzed for histopathological, metabolic and pro-inflammatory gene expression changes.Results
Significant epithelial necrosis, increased levels of l-citrulline and IL-1α were observed on explants infected with B. hyodysenteriae.Conclusions
The spirochete induces necrosis in vitro likely through an inflammatory process mediated by IL-1α and NO.7.
Young Chan Kim Cesar Lopez-Camacho Joanne E. Nettleship Nahid Rahman Michelle L. Hill Laura Silva-Reyes Georgina Ortiz-Martinez Gloria Figueroa-Aguilar María Antonieta Mar Héctor Vivanco-Cid Christine S. Rollier Nicole Zitzmann Martha Eva Viveros-Sandoval Raymond J. Owens Arturo Reyes-Sandoval 《Virology journal》2018,15(1):193
Background
Zika virus (ZIKV) has become a global threat with immediate need for accurate diagnostics, efficacious vaccines and therapeutics. Several ZIKV envelope (Env)-based vaccines have been developed recently. However, many commercially available ZIKV Env are based on the African lineage and produced in insect cells. Here, we sought to produce Asian-lineage ZIKV Env in mammalian cells for research and clinical applications.Methods
We designed various gene expression constructs to optimize the production of ZIKV using prM-Env and full or C-terminal truncations of Env; with or without a rat CD4 fusion partner to allow large-scale production of soluble protein in mammalian HEK293 cells. Protein expression was verified by mass spectrometry and western-blot with a pan-flavivirus antibody, a ZIKV Env monoclonal antibody and with immune sera from adenoviral (ChAdOx1) ZIKV Env-vaccinated mice. The resulting Env-CD4 was used as a coating reagent for immunoassay (ELISA) using both mouse and human seropositive sera.Results
Replacement of the C-terminus transmembrane Env domain by a rat CD4 and addition of prM supported optimal expression and secretion of Env. Binding between the antigens and the antibodies was similar to binding when using commercially available ZIKV Env reagents. Furthermore, antibodies from ZIKV patients bound ZIKV Env-CD4 in ELISA assays, whereas sera from healthy blood donors yielded minimal OD background. The serological outcomes of this assay correlated also with ZIKV neutralisation capacity in vitro.Conclusions
Results obtained from this study indicate the potential of the Asian-lineage Zika Env-CD4 and Env proteins in ELISA assays to monitor humoral immune responses in upcoming clinical trials as well as a sero-diagnostic tool in ZIKV infection.8.
Clenivaldo Alves Caixeta Marina Lara de Carli Noé Vital Ribeiro Júnior Felipe Fornias Sperandio Suely Nonogaki Denismar Alves Nogueira Alessandro Antônio Costa Pereira João Adolfo Costa Hanemann 《Mycopathologia》2018,183(5):785-791
Background
Paracoccidioidomycosis is a neglected tropical fungal infection with great predilection for adult men, indicating the participation of female hormone estrogen in preventing paracoccidioidomycosis development in women. Estrogen has an immunologic effect leading to polarization toward the Th2 immune response, which favors the disease evolution.Objectives
To evaluate estrogen and progesterone receptors in oral paracoccidioidomycosis lesions and to verify any association with tissue fungi counting in women and men.Methods
Thirty-two cases of chronic oral paracoccidioidomycosis were included. Immunohistochemical analyses for anti-estrogen receptor-α, anti-progesterone receptor and anti-Paracoccidioides brasiliensis antibodies were performed. The differences between women and men and the relations among the immunomarkers for each gender were also evaluated.Results
A significant positive correlation was observed between estrogen receptor-α and the amount of fungi in women. In addition, estrogen receptor-α was mildly expressed in the inflammatory cells of female patients, while progesterone receptor was expressed in both genders, with similar expression between women and men. Moreover, fungi counting revealed no differences between genders.Conclusions
Estrogen receptor-α was expressed only in women and showed a positive correlation with the amount of fungi in oral paracoccidioidomycosis, while progesterone receptor was observed in both genders and exhibited no correlation with estrogen receptor-α or fungi counting.9.
Background
The current literature establishes the importance of gene functional category and expression in promoting or suppressing duplicate gene loss after whole genome doubling in plants, a process known as fractionation. Inspired by studies that have reported gene expression to be the dominating factor in preventing duplicate gene loss, we analyzed the relative effect of functional category and expression.Methods
We use multivariate methods to study data sets on gene retention, function and expression in rosids and asterids to estimate effects and assess their interaction.Results
Our results suggest that the effect on duplicate gene retention fractionation by functional category and expression are independent and have no statistical interaction.Conclusion
In plants, functional category is the more dominant factor in explaining duplicate gene loss.10.
Rachel A. Spicer Christoph Steinbeck 《Metabolomics : Official journal of the Metabolomic Society》2018,14(1):16
Introduction
Data sharing is being increasingly required by journals and has been heralded as a solution to the ‘replication crisis’.Objectives
(i) Review data sharing policies of journals publishing the most metabolomics papers associated with open data and (ii) compare these journals’ policies to those that publish the most metabolomics papers.Methods
A PubMed search was used to identify metabolomics papers. Metabolomics data repositories were manually searched for linked publications.Results
Journals that support data sharing are not necessarily those with the most papers associated to open metabolomics data.Conclusion
Further efforts are required to improve data sharing in metabolomics.11.
Xin-Jian Wang Jiang Wang Yue-Ying Wang Yu-Jie Guo Bei-Bei Chu Guo-Yu Yang 《Biotechnology letters》2016,38(6):949-958
Objectives
To determine the effects of the Sus scrofa matrix attachment region (SusMAR) on transgene expression in HEK293T cells.Results
Three expression vectors with the MAR at different sites in the PiggyBac (PB) transposon vector backbone were compared: two MARs flanking the β-galactosidase (β-gal) expression cassette, and one at the upstream or downstream site. Bos taurus MAR (BosMAR) and a β-gal expression cassette without MARs were the positive and negative controls, respectively. Compared to the control, β-gal activity of all SusMAR and BosMAR vectors was significantly improved in the presence of PB transposase (PBase). However, only the downstream SusMAR and upstream BosMAR vectors showed increased expression in the absence of PBase. Expression was significantly increased in all vectors with the PBase group compared to those without the PBase group. Gene copy numbers were not increased compared to the negative control.Conclusions
SusMAR enhanced recombinant gene expression levels and stability in HEK293T cells, was not increase transgene copy number. These results could facilitate the development of vectors for stable production of therapeutic proteins.12.
Yangyang Wang Xinke Yan Shengyuan Zeng Ting Zhang Fengjuan Cheng Rongjuan Chen Changzhu Duan 《Biotechnology letters》2018,40(8):1181-1188
Objectives
To investigate the interaction of E3 ubiquitin ligase UHRF2 with p21 and the mechanism of UHRF2 in repairing DNA damage caused by hydroxyurea (HU) in HEK293 cells.Results
Western blotting indicated that the overexpression of UHRF2 reduced the level of p21, particularly in HEK293 cells. Immunoprecipitation and immunofluorescence staining reveled that UHRF2 combined with p21 in the nucleus. In addition, UHRF2 degraded p21 through ubiquitination and shortened the half-life of p21. UHRF2 could repair DNA damage caused by HU treatment, which was impaired by the inhibition of p21 in HEK293 cells.Conclusions
UHRF2 may negatively modulate p21 to regulate DNA damage response, suggesting a novel pathway of UHRF2 repairing DNA damage through the partial regulation of p21.13.
A. B. Araújo J. M. Furlan G. D. Salton T. Schmalfuss L. M. Röhsig L. M. R. Silla E. P. Passos A. H. Paz 《Biotechnology letters》2018,40(6):989-998
Objective
To compare four enzymatic protocols for mesenchymal stem cells (MSCs) isolation from amniotic (A-MSC) and chorionic (C-MSC) membranes, umbilical cord (UC-MSC) and placental decidua (D-MSC) in order to define a robust, practical and low-cost protocol for each tissue.Results
A-MSCs and UC-MSCs could be isolated from all samples using trypsin/collagenase-based protocols; C-MSCs could be isolated from all samples with collagenase- and trypsin/collagenase-based protocols; D-MSCs were isolated from all samples exclusively with a collagenase-based protocol.Conclusions
The trypsin-only protocol was least efficient; the collagenase-only protocol was best for C-MSCs and D-MSCs; the combination of trypsin and collagenase was best for UC-MSCs and none of tested protocols was adequate for A-MSCs isolation.14.
Background
In recent years the visualization of biomagnetic measurement data by so-called pseudo current density maps or Hosaka-Cohen (HC) transformations became popular.Methods
The physical basis of these intuitive maps is clarified by means of analytically solvable problems.Results
Examples in magnetocardiography, magnetoencephalography and magnetoneurography demonstrate the usefulness of this method.Conclusion
Hardware realizations of the HC-transformation and some similar transformations are discussed which could advantageously support cross-platform comparability of biomagnetic measurements.15.
Introduction
Untargeted metabolomics is a powerful tool for biological discoveries. To analyze the complex raw data, significant advances in computational approaches have been made, yet it is not clear how exhaustive and reliable the data analysis results are.Objectives
Assessment of the quality of raw data processing in untargeted metabolomics.Methods
Five published untargeted metabolomics studies, were reanalyzed.Results
Omissions of at least 50 relevant compounds from the original results as well as examples of representative mistakes were reported for each study.Conclusion
Incomplete raw data processing shows unexplored potential of current and legacy data.16.
Background
Chemical crosslinking is the most straightforward method to produce bispecific antibodies (BsAb) for arming ex vivo activated cytotoxic T lymphocytes. However, heterogeneous polymers are produced by chemical crosslinking. Currently, it is not known under what circumstances or to what extent further purification is needed.Results
In this study, we purified Traut’s Reagent-Sulfo-SMCC crosslinked anti-CD3?×?anti-HER2 by size-exclusion column chromatography and compared the capacity of the crude and the purified forms of the BsAb in enhancing cytokine-induced killer (CIK) cell-mediated cytotoxicity in vitro. We found that the purified BsAb assisted CIK cells more efficiently than the crude form only when the spontaneous cytotoxicity of the CIK cells was relatively low; otherwise, the two forms performed almost identically.Conclusions
For the CIK cells of low spontaneous cytotoxicity, purified BsAb is a more powerful substitute for crude BsAb in enhancing their killing efficacy. However, that purification of BsAb is not necessary for robust CIK cells. This phenomenon also corroborates that CIK-mediated cytotoxicity is highly dependent on cell contact.17.
Sven Zukunft Cornelia Prehn Cornelia Röhring Gabriele Möller Martin Hrabě de Angelis Jerzy Adamski Janina Tokarz 《Metabolomics : Official journal of the Metabolomic Society》2018,14(1):18
Introduction
Global metabolomics analyses using body fluids provide valuable results for the understanding and prediction of diseases. However, the mechanism of a disease is often tissue-based and it is advantageous to analyze metabolomic changes directly in the tissue. Metabolomics from tissue samples faces many challenges like tissue collection, homogenization, and metabolite extraction.Objectives
We aimed to establish a metabolite extraction protocol optimized for tissue metabolite quantification by the targeted metabolomics AbsoluteIDQ? p180 Kit (Biocrates). The extraction method should be non-selective, applicable to different kinds and amounts of tissues, monophasic, reproducible, and amenable to high throughput.Methods
We quantified metabolites in samples of eleven murine tissues after extraction with three solvents (methanol, phosphate buffer, ethanol/phosphate buffer mixture) in two tissue to solvent ratios and analyzed the extraction yield, ionization efficiency, and reproducibility.Results
We found methanol and ethanol/phosphate buffer to be superior to phosphate buffer in regard to extraction yield, reproducibility, and ionization efficiency for all metabolites measured. Phosphate buffer, however, outperformed both organic solvents for amino acids and biogenic amines but yielded unsatisfactory results for lipids. The observed matrix effects of tissue extracts were smaller or in a similar range compared to those of human plasma.Conclusion
We provide for each murine tissue type an optimized high-throughput metabolite extraction protocol, which yields the best results for extraction, reproducibility, and quantification of metabolites in the p180 kit. Although the performance of the extraction protocol was monitored by the p180 kit, the protocol can be applicable to other targeted metabolomics assays.18.
Jamie V. de Seymour Stephanie Tu Xiaoling He Hua Zhang Ting-Li Han Philip N. Baker Karolina Sulek 《Metabolomics : Official journal of the Metabolomic Society》2018,14(6):79
Introduction
Intrahepatic cholestasis of pregnancy (ICP) is a common maternal liver disease; development can result in devastating consequences, including sudden fetal death and stillbirth. Currently, recognition of ICP only occurs following onset of clinical symptoms.Objective
Investigate the maternal hair metabolome for predictive biomarkers of ICP.Methods
The maternal hair metabolome (gestational age of sampling between 17 and 41 weeks) of 38 Chinese women with ICP and 46 pregnant controls was analysed using gas chromatography–mass spectrometry.Results
Of 105 metabolites detected in hair, none were significantly associated with ICP.Conclusion
Hair samples represent accumulative environmental exposure over time. Samples collected at the onset of ICP did not reveal any metabolic shifts, suggesting rapid development of the disease.19.
Xiaofang Deng Zhuqing Zhao JiaJie Zhou JinZhao Chen Chenhao Lv Xinwei Liu 《Plant and Soil》2018,433(1-2):55-64
Aims
The objectives of this study were to determine the valence state of Se and major components in Se ore powder, examine its effect on Se enrichment in crops under different cultivation patterns, and assess the safety of edible parts of crops.Methods
Se ore powder from Enshi was subjected to compositional analysis by X-ray photoelectron spectroscopy and applied to rice (wetland) and soybean (dryland) fields before planting.Results
Se was mainly present in tetravalent and zero-valent forms at a 4:6 ratio in Se ore powder. Following soil application of Se ore powder, concentrations of As, Hg, Cr, and Pb in brown rice and soybean seed were below the limit of detection, and the Cd concentration was below the European Union food regulation limit. Organic Se accounted for more than 96% of total Se in both crops, including >91% Se-methionine. Compared with dryland, the flooded environment facilitated residual Se transformation into Fe/Mn oxide-bound Se, and thus increased the potential activity of Se in the soil.Conclusions
Se ore powder can be applied for the production of Se-rich agricultural products in the short-term, and possibilities of upgrading the Se ore product should be considered in the future.20.