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E Sweeney, A M Lovering, K E Bowker, A P MacGowan, S M Nelson
Chronic osteomyelitis is difficult to treat, with biofilm growth and the diffusion barrier to antibiotics presented by bone contributory factors. The aim of this study was to develop and evaluate an in vitro model of osteomyelitis. A bioluminescent strain of Staphylococcus aureus was grown in bone blocks made from bovine femur. Light output was insufficient for detection of bacterial cells within bone by 24 h and viable counting of crushed bone blocks was used to determine bacterial survival. Challenge of 72 h biofilms with gentamicin and daptomycin for 24 h demonstrated that only concentrations of ten times the clinical peak serum target levels (100 mg l-1 gentamicin and 1000 mg l-1 daptomycin) resulted in significant reductions in cell viability compared to controls...
February 15, 2019: Letters in Applied Microbiology
Nicolas Bery, Terence H Rabbitts
Protein-protein interactions (PPIs) are principle biological processes that control normal cell growth, differentiation, and homeostasis but are also crucial in diseases such as malignancy, neuropathy, and infection. Despite the importance of PPIs in biology, this target class has been very challenging to convert to therapeutics. In the last decade, much progress has been made in the inhibition of PPIs involved in diseases, but many remain difficult such as RAS-effector interactions in cancers. We describe here a protocol for using Bioluminescence Resonance Energy Transfer 2 (BRET2)-based RAS biosensors to detect and characterize RAS PPI inhibition by macromolecules and small molecules...
February 15, 2019: Current Protocols in Cell Biology
Diana Caridha, Mark Hickman, Lisa Xie, Franklyn Ngundam, Erin Milner, Amanda Schenk, Kirk Butler, Dylan Nugent, Patricia Lee, Norma Roncal, Susan Leed, Eve Hosford, Jangwoo Lee, Richard J Sciotti, Gregory Reichard, Chad Black, Mara Kreishman-Deitrick, Qigui Li, Brian Vesely
BACKGROUND: Rodent malaria models are extensively used to predict treatment outcomes in human infections. There is a constant need to improve and refine these models by innovating ways to apply new scientific findings and cutting edge technologies. In addition, and in accordance with the three R's of animal use in research, in vivo studies should be constantly refined to avoid unnecessary pain and distress to the experimental animals by using preemptive euthanasia as soon as the main scientific study objective has been accomplished...
February 15, 2019: Malaria Journal
Devendra H Dusane, Varun Lochab, Travis Jones, Casey W Peters, Devin Sindeldecker, Amitava Das, Sashwati Roy, Chandan K Sen, Vish V Subramaniam, Daniel J Wozniak, Shaurya Prakash, Paul Stoodley
Electroceutical wound dressings, especially those involving current flow with silver based electrodes, show promise for treating biofilm infections. However, their mechanism of action is poorly understood. We have developed an in vitro agar based model using a bioluminescent strain of Pseudomonas aeruginosa to measure loss of activity and killing when direct current was applied. Silver electrodes were overlaid with agar and lawn biofilms grown for 24 h. A 6 V battery with 1 kΩ ballast resistor was used to treat the biofilms for 1 h or 24 h...
February 14, 2019: Scientific Reports
Raquel Álvarez-Velilla, Maria Del Camino Gutiérrez-Corbo, Carmen Punzón, Maria Yolanda Pérez-Pertejo, Rafael Balaña-Fouce, Manuel Fresno, Rosa María Reguera
BACKGROUND: Visceral leishmaniasis is a neglected parasitic disease with no vaccine available and its pharmacological treatment is reduced to a limited number of unsafe drugs. The scarce readiness of new antileishmanial drugs is even more alarming when relapses appear or the occurrence of hard-to-treat resistant strains is detected. In addition, there is a gap between the initial and late stages of drug development, which greatly delays the selection of leads for subsequent studies. METHODOLOGY/PRINCIPAL FINDINGS: In order to address these issues, we have generated a red-shifted luminescent Leishmania infantum strain that enables long-term monitoring of parasite burden in individual animals with an in vivo limit of detection of 106 intracellular amastigotes 48 h postinfection...
February 14, 2019: PLoS Neglected Tropical Diseases
Rohit Srivastava, Anna Jablonska, Chengyan Chu, Lydia Gregg, Jeff Bulte, Raymond C Koehler, Piotr Walczak, Miroslaw Janowski
White matter damage persists in hypoxic-ischemic newborns even when treated with hypothermia. We have previously shown that intra-ventricular delivery of human glial progenitors (GPs) at the neonatal stage is capable of replacing abnormal host glia and rescuing the lifespan of dysmyelinated mice. However, such transplantation in the human brain poses significant challenges as related to high-volume ventricles and long cell migration distances. These challenges can only be studied in large animal model systems...
February 14, 2019: Stem Cells and Development
Jonathan P Evans, Boleslaw K Winiarski, Paul A Sutton, Lorenzo Ressel, Carrie A Duckworth, D Mark Pritchard, Daniel H Palmer, Christopher E Goldring, Neil R Kitteringham
Improving outcomes in colorectal cancer requires more accurate in vivo modelling of the disease in humans, allowing more reliable pre-clinical assessment of potential therapies. Novel imaging techniques are necessary to improve the longitudinal assessment of disease burden in these models, reducing the number of animals required for translational studies. This report describes the development of an immune-competent syngeneic orthotopic murine model of colorectal cancer, utilising caecal implantation of CT26 cells stably transfected with the luciferase gene into immune-competent BALB/c mice, allowing serial bioluminescent imaging of cancer progression...
February 13, 2019: Laboratory Animals
Matthew J Rubin, Marcus T Brock, Seth J Davis, Cynthia Weinig
The circadian clock facilitates coordination of the internal rhythms of an organism to daily environmental conditions, such as the light-dark cycle of one day. Circadian period length (the duration of one endogenous cycle) and phase (the timing of peak activity) exhibit quantitative variation in natural populations. Here, we measured circadian period and phase in June, July and September in three Arabidopsis thaliana recombinant inbred line populations. Circadian period and phase were estimated from bioluminescence of a genetic construct between a native circadian clock gene ( COLD CIRCADIAN RHYTHM RNA BINDING 2 ) and the reporter gene ( LUCIFERASE ) after lines were entrained under field settings...
February 12, 2019: G3: Genes—Genomes—Genetics
Nicholas Aj Dawson, Caroline Lamarche, Romy E Hoeppli, Peter Bergqvist, Vivian Fung, Emma McIver, Qing Huang, Jana Gillies, Madeleine Speck, Paul C Orban, Jonathan W Bush, Majid Mojibian, Megan K Levings
Chimeric antigen receptor (CAR) technology can be used to engineer the antigen-specificity of regulatory T cells (Tregs) and improve their potency as an adoptive cell therapy in multiple disease models. As synthetic receptors, CARs carry the risk of immunogenicity, particularly when derived from non-human antibodies. Using an HLA-A*02:01-specific CAR (A2-CAR) encoding a single-chain Fv derived from a mouse antibody, we developed a panel of 20 humanized (h)A2-CARs. Systematic testing demonstrated variations in expression, ability to bind HLA-A*02:01, and stimulate human Treg suppression in vitro...
February 12, 2019: JCI Insight
Jun Wang, Liying Zhu, Xinwei Chen, Ruofei Huang, Shili Wang, Pin Dong
Hypopharyngeal carcinoma is a common malignant tumor of the head and neck with a very poor prognosis; the median survival time for curatively treated patients was 17.2 months in India. However, cell-based gene therapy holds promise to improve patient outcomes. In this study, we investigated whether human bone marrow mesenchymal stem cells (BMSCs) possess potential homing capacity for hypopharyngeal carcinoma. To monitor the efficiency of BMSC transplantation therapy via reporter gene imaging, we employed a hybrid baculovirus vector containing the Luc-P2A-eGFP fusion or sodium iodide symporter (NIS) sequence under the control of the cytomegalovirus promoter...
February 12, 2019: Stem Cells and Development
Anna Davydova, Mariya Vorobyeva, Eugenia Bashmakova, Pavel Vorobjev, Olga Krasheninina, Alexey Tupikin, Marsel Kabilov, Vasilisa Krasitskaya, Ludmila Frank, Alya Venyaminova
Aptamers are short DNA and RNA fragments which bind their molecular targets with affinity and specificity comparable to those of antibodies. Here, we describe the selection of novel 2'-F-RNA aptamers against total human hemoglobin or its glycated form HbA1c. After SELEX and high-throughput sequencing of the enriched libraries, affinities and specificities of candidate aptamers and their truncated variants were examined by the solid-phase bioluminescent assay. As a result, we identified aptamers specific to both hemoglobins or only glycated HbA1c...
February 8, 2019: Analytical Biochemistry
Alyssa S Ball, Julia C van Kessel
In Vibrio species, quorum sensing controls gene expression for numerous group behaviors, including bioluminescence production, biofilm formation, virulence factor secretion systems, and competence. The LuxR/HapR master quorum sensing regulators activate expression of hundreds of genes in response to changes in population densities. The mechanism of transcription activation by these TetR-type transcription factors is unknown, though LuxR DNA binding sites that lie in close proximity to the -35 region of the promoter are required for activation at some promoters...
February 11, 2019: Molecular Microbiology
Sanjay Anand, Anton Yasinchak, Taylor Bullock, Mukul Govande, Edward V Maytin
Aim: Breast cancer (BCA) in women is a leading cause of mortality and morbidity; distant metastases occur in ~40% of cases. Here, as an alternative to ionizing radiation therapy and chemotherapy and their associated side effects, we explored a new combination approach using capecitabine (CPBN) and aminolevulinate-based photodynamic therapy (PDT). We had previously developed a combination PDT approach in which 5-fluorouracil (5FU), a differentiation-promoting agent, increases the levels of protoporphyrin IX (PpIX) in cancer cells when given as a neoadjuvant prior to aminolevulinic acid (ALA)...
2019: Journal of Cancer Metastasis and Treatment
Vít Herynek, Karolína Turnovcová, Andrea Gálisová, Ondřej Kaman, Dana Mareková, Jakub Koktan, Magda Vosmanská, Lucie Kosinová, Pavla Jendelová
Manganese-zinc ferrite nanoparticles were synthesized by using a hydrothermal treatment, coated with silica, and then tested as efficient cellular labels for cell tracking, using magnetic resonance imaging (MRI) in vivo. A toxicity study was performed on rat mesenchymal stem cells and C6 glioblastoma cells. Adverse effects on viability and cell proliferation were observed at the highest concentration (0.55 mM) only; cell viability was not compromised at lower concentrations. Nanoparticle internalization was confirmed by transmission electron microscopy...
February 2019: ChemistryOpen
Warren M Kadoya, Reyes Sierra-Alvarez, Bhumasamudram Jagadish, Stanley Wong, Leif Abrell, Eugene A Mash, Jim A Field
Explosives, pesticides, and pharmaceuticals contain toxic nitroaromatic compounds that may form even more toxic azo compounds if they encounter reducing conditions in the environment. We investigated the mechanism by which 4,4'-dimethoxyazobenzene forms in anaerobic sludge incubations of 4-nitroanisole, an analog for the insensitive munitions compound 2,4-dinitroanisole (DNAN). Because studies have reported the mechanism to involve the coupling of reduced nitroaromatic intermediates, specifically aromatic amines and nitrosoaromatics, by nucleophilic processes, we abiotically paired 10 mM 4-aminoanisole with 2 mM 4-nitrosoanisole in nitrogen-flushed microcosms...
February 2, 2019: Chemosphere
Rafael Franco, María Villa, Paula Morales, Irene Reyes-Resina, Ana Gutiérrez-Rodríguez, Jasmina Jiménez, Nadine Jagerovic, José Martínez-Orgado, Gemma Navarro
Preclinical work shows cannabidiol as a promising drug to manage neonatal hypoxic-ischemic brain damage (NHIBD). The molecular mechanism is not well defined but the beneficial effects of this phytocannabinoid are blocked by antagonists of both cannabinoid CB2 (CB2 R) and serotonin 5-HT1A (5-HT1AR) receptors that, in addition, may form heteromers in a heterologous expression system. Using bioluminescence energy transfer, we have shown a direct interaction of the two receptors that leads to a particular signaling in a heterologous system...
February 6, 2019: Neuropharmacology
Qiong Zhang, Yizhi Song, Ken Amor, Wei E Huang, Don Porcelli, Ian Thompson
Bioremediation is a sustainable and cost-effective means of contaminant detoxification. Although Cr(VI) is toxic at high concentrations, various microbes can utilise it as an electron accepter in the bioremediation process, and reduce it to the less toxic form Cr(III). During remediation, it is important to monitor the level of toxicity and effectiveness of Cr(VI) reduction in order to optimize the conditions. This study employed a whole-cell bioreporter Acinetobacter baylyi ADPWH-recA to access the degree of toxicity of different species of Cr over a range of initial concentrations...
January 21, 2019: Water Research
Bang Xiao, Wenjun Yang, Dong Lei, Jinfeng Huang, Yupeng Yin, Yiqing Zhu, Zhengwei You, Fang Wang, Shuhan Sun
Intrauterine adhesion (IUA) causing infertility and recurrent miscarriage of reproductive female mammals usually results from endometrium injury. Nevertheless, there is no efficient therapeutic method to avoid IUA. Bone marrow derived mesenchymal stem cells (BMSCs) are an important cell source for tissue regeneration. This study designs and explores the ability of BMSC-loaded elastic poly(glycerol sebacate) (PGS) scaffold to prevent IUA and compares the effect of PGS with poly(lactic-co-glycolic acid) (PLGA) and collagen scaffolds in resumption of damaged rat uteruses...
February 7, 2019: Advanced Healthcare Materials
Eszter Simon, Andrea Schifferli, Thomas B Bucher, Daniel Olbrich, Inge Werner, Etiënne L M Vermeirssen
Ecotoxicological screening of surface waters can involve multiple analyses using multiple bioassay and chemical analytical methods that require enriched samples to reach low concentrations. Such broad screening of the same sample necessitates sufficient sample volume-typically several liters-to produce a sufficient amount of enriched sample. Often, this is achieved by performing parallel solid-phase extractions (SPE) where extracts are combined into a pool-this is a laborious process. In this study, we first validated our existing SPE method for the chemical recovery of an extended set of compounds...
February 8, 2019: Analytical and Bioanalytical Chemistry
Katie M Parkins, Amanda M Hamilton, Veronica P Dubois, Suzanne M Wong, Paula J Foster, John A Ronald
Purpose: The combined use of anatomical magnetic resonance imaging (MRI), cellular MRI, and bioluminescence imaging (BLI) allows for sensitive and improved monitoring of brain metastasis in preclinical cancer models. By using these complementary technologies, we can acquire measurements of viable single cell arrest in the brain after systemic administration, the clearance and/or retention of these cells thereafter, the growth into overt tumours, and quantification of tumour volume and relative cancer cell viability over time...
2019: Contrast Media & Molecular Imaging
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