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Neftali Vazquez, Lilia Sanchez, Rebecca Marks, Eduardo Martinez, Victor Fanniel, Alma Lopez, Andrea Salinas, Itzel Flores, Jesse Hirschmann, Robert Gilkerson, Erin Schuenzel, Robert Dearth, Reginald Halaby, Wendy Innis-Whitehouse, Megan Keniry
BACKGROUND: Clustered regularly interspaced short palindromic repeat (CRISPR) RNA-guided adaptive immune systems are found in prokaryotes to defend cells from foreign DNA. CRISPR Cas9 systems have been modified and employed as genome editing tools in wide ranging organisms. Here, we provide a detailed protocol to truncate genes in mammalian cells using CRISPR Cas9 editing. We describe custom donor vector construction using Gibson assembly with the commonly utilized pcDNA3 vector as the backbone...
March 14, 2018: BMC Molecular Biology
Kevin Rey, Sukhbir Manku, Winnie Enns, Thea Van Rossum, Kevin Bushell, Ryan D Morin, Fiona S L Brinkman, Jonathan C Choy
BACKGROUND: The gut microbiota influences many immunological processes but how its disruption affects transplant rejection is poorly understood. METHODS: Interposition grafting of aortic segments was used to examine vascular rejection. The gut microbiota was disrupted in graft recipients using an antibiotic cocktail (ampicillin, vancomycin, metronidazole, neomycin sulfate) in their drinking water. RESULTS: Treatment of mice with antibiotics severely reduced total bacterial content in the intestine and disrupted the bacterial composition...
March 10, 2018: Transplantation
Ying-Chun Wan, Yan-Jie Liu, Chen Liu, Hui-Ting Ma, Hui-Fang Yu, Jia-Wei Kang, Chen-Long Gao, Zhong-Qiao Wu, Dan Zheng, Bin Lu
A new analytical method was developed to detect neomycin in complex biological samples using molecularly imprinted polymer to construct an optical sensor. Fluorescent neomycin-imprinted polymers (fMIPs) containing both imprinted cavity and boronate affinity site were synthesized on the surface of silica-modified quantum dots. The fMIPs exhibited high selectivity to neomycin by having two binding sites for the target analyte. Neomycin analogues (competing for imprinted cavity) and D-glucose (competing for the boronate affinity site) did not affect the selectivity of the fMIPs...
February 22, 2018: Journal of Pharmaceutical and Biomedical Analysis
Youngji Cho, Galeb Abu-Ali, Hiroki Tashiro, David I Kasahara, Traci A Brown, Jeffrey D Brand, Joel A Mathews, Curtis Huttenhower, Stephanie A Shore
Previous reports demonstrate that the microbiome impacts allergic airway responses, including airway hyperresponsiveness, a characteristic feature of asthma. Here we examined the role of the microbiome in pulmonary responses to a non-allergic asthma trigger, ozone. We depleted the microbiota of conventional mice with either a single antibiotic (ampicillin, metronidazole, neomycin, or vancomycin) or a cocktail of all four antibiotics given via the drinking water. Mice were then exposed to room air or ozone. In air-exposed mice, airway responsiveness was not different in antibiotic- versus control water-treated mice...
March 12, 2018: American Journal of Respiratory Cell and Molecular Biology
M Boyer, M Sowa, I Di Meo, S Eftekharian, M R Steenari, V Tiranti, J E Abdenur
Ethylmalonic encephalopathy (EE) is a devastating neurodegenerative disease caused by mutations in the ETHE1 gene critical for hydrogen sulfide (H2 S) detoxification. Patients present in infancy with hypotonia, developmental delay, diarrhea, orthostatic acrocyanosis and petechiae. Biochemical findings include elevated C4, C5 acylcarnitines and lactic and ethylmalonic acid (EMA) in body fluids. Current treatment modalities include metronidazole and N-acetylcysteine (NAC) to lower the production and promote detoxification of toxic H2 S...
February 14, 2018: Molecular Genetics and Metabolism
Garrett T Dow, James B Thoden, Hazel M Holden
The aminoglycoside antibiotics, discovered as natural products in the 1940s, demonstrate a broad anti-microbial spectrum. Due to their nephrotoxic and ototoxic side effects, however, their widespread clinical usage has typically been limited to the treatment of serious infections. Neomycin B, first isolated from strains of Streptomyces in 1948, is one such drug that was approved for human use by the U. S. Food and Drug Administration in 1964. Only within the last 11 years has the biochemical pathway for its production been elaborated, however...
March 8, 2018: Protein Science: a Publication of the Protein Society
Kristin M Whitworth, Raissa Cecil, Joshua A Benne, Bethany K Redel, Lee D Spate, Melissa S Samuel, Randall S Prather, Kevin D Wells
Genetically engineered pigs are often created with a targeting vector that contains a loxP flanked selectable marker like neomycin. The Cre-loxP recombinase system can be used to remove the selectable marker gene from the resulting offspring or cell line. Here is described a new method to remove a loxP flanked neomycin cassette by direct zygote injection of an mRNA encoding Cre recombinase. The optimal concentration of mRNA was determined to be 10 ng/μL when compared to 2 and 100 ng/μL (P < 0.0001)...
March 7, 2018: Transgenic Research
Aafke W F Janssen, Saeed Katiraei, Barbara Bartosinska, Daniel Eberhard, Ko Willems van Dijk, Sander Kersten
AIMS/HYPOTHESIS: Angiopoietin-like 4 (ANGPTL4) is an important regulator of triacylglycerol metabolism, carrying out this role by inhibiting the enzymes lipoprotein lipase and pancreatic lipase. ANGPTL4 is a potential target for ameliorating cardiometabolic diseases. Although ANGPTL4 has been implicated in obesity, the study of the direct role of ANGPTL4 in diet-induced obesity and related metabolic dysfunction is hampered by the massive acute-phase response and development of lethal chylous ascites and peritonitis in Angptl4-/- mice fed a standard high-fat diet...
March 3, 2018: Diabetologia
Kyosuke Kanai, Ah-Mee Park, Akiko Watanabe, Tomohiro Arikawa, Teruhito Yasui, Hiroki Yoshida, Ikuo Tsunoda, Osamu Yoshie
IL-27 is an immunoregulatory cytokine consisting of p28 and EBI3. Its receptor also has two subunits, WSX1 and gp130. Although IL-27 promotes Th1 differentiation in naive T cells, it also induces IL-10 expression in effector Th1 cells to curtail excessive immune responses. By using p28-deficient mice and WSX1-deficient mice (collectively called IL-27-deficient mice), we examined the role of IL-27 in primary infection by murine γ-herpesvirus 68 (MHV68), a murine model of EBV. Upon airway infection with MHV68, IL-27-deficient mice had more aggravated lung inflammation than wild-type mice, although MHV68 infection per se was better controlled in IL-27-deficient mice...
March 2, 2018: Journal of Immunology: Official Journal of the American Association of Immunologists
Jonathan G Bonchak, Megan E Prouty, Salma F de la Feld
BACKGROUND: Personal care products marketed for babies and children are often regarded as "safe" or "gentle." However, little is known about the prevalence of contact allergens in these types of products. OBJECTIVE: This study assessed the prevalence of important sensitizers in personal care products marketed for babies and children. A secondary objective of this study was to determine whether a product's cost correlates with content of sensitizing ingredients...
February 28, 2018: Dermatitis
Victor A Amadi, Harry Hariharan, Gitanjali Arya, Vanessa Matthew-Belmar, Roxanne Nicholas-Thomas, Rhonda Pinckney, Ravindra Sharma, Roger Johnson
Non-typhoidal salmonellosis remains an important public health problem worldwide. Dogs may harbour Salmonella in their intestines and can easily shed Salmonella in the environment with the possibility of transmission to humans. Thus, monitoring is essential to understand the role of dogs in zoonotic transmission. The objectives of this study were to determine the shedding of Salmonella by owned, apparently healthy dogs in Grenada, West Indies, to identify the serovars, and to examine their antimicrobial susceptibility profiles...
February 2018: Veterinary Medicine and Science
Mireia Uribe-Herranz, Kyle Bittinger, Stavros Rafail, Sonia Guedan, Stefano Pierini, Ceylan Tanes, Alex Ganetsky, Mark A Morgan, Saar Gill, Janos L Tanyi, Frederic D Bushman, Carl H June, Andrea Facciabene
Adoptive T cell therapy (ACT) is a promising new modality for malignancies. Here, we report that adoptive T cell efficacy in tumor-bearing mice is significantly affected by differences in the native composition of the gut microbiome or treatment with antibiotics, or by heterologous fecal transfer. Depletion of bacteria with vancomycin decreased the rate of tumor growth in mice from The Jackson Laboratory receiving ACT, whereas treatment with neomycin and metronidazole had no effect, indicating the role of specific bacteria in host response...
February 22, 2018: JCI Insight
Carolyn O Dirain, Brendan Kosko, Patrick J Antonelli
Objective Ciprofloxacin + dexamethasone ear drops have been associated with higher rates of tympanic membrane perforations than ofloxacin. This was thought to be an effect of the steroid; however, ciprofloxacin (sans steroid) has been found to be more toxic to tympanic membrane fibroblasts than ofloxacin in vitro. This study aimed to compare the effect of these agents on tympanic membrane healing in vivo. Study Design Controlled animal study. Setting Academic research laboratory. Methods Perforations were created in 54 rats with a carbon dioxide laser...
February 1, 2018: Otolaryngology—Head and Neck Surgery
Camille Silva Florencio, Fabiana Alves Silva Brandão, Marcus de Mello Teixeira, Anamélia Lorenzetti Bocca, Maria Sueli S Felipe, Vânia Aparecida Vicente, Larissa Fernandes
Fonsecaea pedrosoi, a melanized fungal pathogen that causes Chromoblastomycosis, a human disease with a worldwide distribution. Biolistic is a widely used technique for direct delivery of genetic material into intact cells by particles bombardment. Another well-established transformation method is Agrobacterium-mediated transformation (ATMT), which involves the transfer of a T-DNA from the bacterium to the target cells. In F. pedrosoi there are no reports of established protocols for genetic transformation, which require optimization of physical and biological parameters...
March 2018: Microbiological Research
Tran Huu Dung, Le Thu Huong, Hoon Yoo
The morphological features of pluronic F127 at various concentrations were investigated and 20% aqueous pluronic F127 solution was chosen for the preparation of biomembrane. The pluronic solution was mixed with appropriate concentration of excipients such as PVA, Povidon S630, PG, Nipagin, Neomycin and Panthenol. The prepared pluronic F127 was thermosensitive, a liquid phase at low temperatures (≤10 °C) but converting into gel at temperatures above 20 °C, and formed stable biomembrane at 37 °C. The neomycin impregnated pluronic membrane suppressed the bacterial growth on agar plate by the sustained release of neomycin...
February 1, 2018: Journal of Nanoscience and Nanotechnology
Yingwang Ye, Na Ling, Jina Gao, Maofeng Zhang, Xiyan Zhang, Liaowang Tong, Dexin Ou, Yaping Wang, Jumei Zhang, Qingping Wu
Cronobacter sakazakii is associated with severe infections including sepsis, neonatal meningitis, and necrotizing enterocolitis. Antibiotic resistance in Cronobacter species has been documented in recent years, but the genes involved in resistance in Cronobacter strains are poorly understood. In this study, we determined the role of outer membrane protein W (OmpW) on survival rates, morphologic changes, and biofilm formation between wild type (WT) and an OmpW mutant strain (ΔOmpW) under neomycin sulfate stress...
February 7, 2018: Journal of Dairy Science
Kazuo Takayama, Yasuko Hagihara, Yukiko Toba, Kiyotoshi Sekiguchi, Fuminori Sakurai, Hiroyuki Mizuguchi
Human iPS cell-derived hepatocyte-like cells are expected to be utilized in pharmaceutical research. However, the purity of high-functioning hepatocyte-like cells is not high enough. In particular, the purity of cytochrome P450 3A4 (CYP3A4), which is a representative hepatic drug-metabolizing enzyme, positive cells is still quite low (approximately 20%). To address this problem, we established the CYP3A4-NeoR-EGFP transgenic reporter human iPS cell line (CYP3A4-NeoR-EGFP iPS cells) by using genome editing technology...
January 18, 2018: Biomaterials
Shasha Liu, Hein Min Tun, Frederick C Leung, Darin C Bennett, Hongfu Zhang, Kimberly M Cheng
Our previous study has shown that genetic selection for susceptibility/resistance to diet-induced atherosclerosis has affected the Japanese quail's cecal environment to accommodate distinctly different cecal microbiota. In this study, we fed the Atherosclerosis-resistant (RES) and -susceptable (SUS) quail a regular and a cholesterol enriched diet to examine the interaction of host genotype and diet on the diversity, composition, and metabolic functions of the duodenal and ileal microbiota with relations to atherosclerosis development...
February 5, 2018: Scientific Reports
Qingxia Xu, Liang Li, Cuihong Han, Li Wei, Lingling Kong, Fanzhong Lin
Sigma-1 receptor (σ1R), an important transmembrane structural protein, has been demonstrated to be overexpressed in various types of human cancer, and has been confirmed to be involved in many biological behaviors during tumorigenesis and tumor progression. The aim of the present study was to explore the essential role of σ1R in hepatic malignant tumors (HMTs), which, to the best of our knowledge, has not been reported to date. We assessed σ1R expression in hepatocellular carcinoma (HCC) tissues and found that σ1R was significantly decreased in HCC when compared with that in benign liver tissues (P<0...
January 22, 2018: Oncology Reports
M Stevens, S Piepers, K Supré, S De Vliegher
The main objectives of this study were to quantify the consumption of antimicrobials on a convenience sample of dairy herds and to determine the association between herd-level antimicrobial consumption and inhibition zone diameters (IZD) of non-aureus staphylococci and Staphylococcus aureus isolates from subclinical mastitis cases. Also, the association between the IZD of non-aureus staphylococci and Staph. aureus isolates within a herd was studied. Antimicrobial consumption data on 56 Flemish dairy farms were obtained between 2013 and 2014 by so-called garbage can audits and expressed as antimicrobial treatment incidence (ATI), with the unit of ATI being the number of defined daily doses animal (DDDA) used per 1,000 cow-days...
February 2, 2018: Journal of Dairy Science
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