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Zhaohui Qiu, Jie Peng, Lingli Mou, Xiao Li, Fanqi Meng, Peng Yu
Multidrug resistance (MDR) of tumors occurs when tumor cells exhibit reduced sensitivity to a large number of unrelated drugs. The molecular mechanism of MDR commonly involves overexpression of the plasma membrane drug efflux pump P-glycoprotein (P-gp). Overexpression of P-gp may be induced by the selection and/or adaptation of cells during exposure to chemotherapeutic drugs, referred to as acquired P-gp-mediated MDR. This study aimed to establish a P-gp quantification method by Ultra Performance Liquid Chromatography and Tandem Mass Spectrometry (UPLC-MS/MS) to better understand the regulation of P-gp expression and its relationship with the level of drug resistance...
March 8, 2018: Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences
Li Bao, Zhaoyang Qian, Maria B Lyng, Ling Wang, Yuan Yu, Ting Wang, Xiuqing Zhang, Huanming Yang, Nils Brünner, Jun Wang, Henrik J Ditzel
Single cancer cell sequencing studies currently use randomly-selected cells, limiting correlations between genomic aberrations, morphology and spatial localization. We laser-captured microdissected single cells from morphologically-distinct areas of primary breast cancer and corresponding lymph node metastasis and performed whole-exome or deep-target sequencing of greater than 100 such cells. Two major subclones co-existed in different areas of the primary tumor, and the lymph node metastasis originated from a minor subclone in the invasive front of the primary tumor with additional copy number changes including 8q gain, but no additional point mutations in driver genes...
March 15, 2018: Journal of Clinical Investigation
Caroline L Furness, Marcela B Mansur, Victoria J Weston, Luca Ermini, Frederik W van Delft, Sarah Jenkinson, Rosemary Gale, Christine J Harrison, Maria S Pombo-de-Oliveira, Marta Sanchez-Martin, Adolfo A Ferrando, Pamela Kearns, Ian Titley, Anthony M Ford, Nicola E Potter, Mel Greaves
Single-cell genetics were used to interrogate clonal complexity and the sequence of mutational events in STIL-TAL1+ T-ALL. Single-cell multicolour FISH was used to demonstrate that the earliest detectable leukaemia subclone contained the STIL-TAL1 fusion and copy number loss of 9p21.3 (CDKN2A/CDKN2B locus), with other copy number alterations including loss of PTEN occurring as secondary subclonal events. In three cases, multiplex qPCR and phylogenetic analysis were used to produce branching evolutionary trees recapitulating the snapshot history of T-ALL evolution in this leukaemia subtype, which confirmed that mutations in key T-ALL drivers, including NOTCH1 and PTEN, were subclonal and reiterative in distinct subclones...
March 20, 2018: Leukemia: Official Journal of the Leukemia Society of America, Leukemia Research Fund, U.K
Liang-Qing Dong, Yang Shi, Li-Jie Ma, Liu-Xiao Yang, Xiao-Ying Wang, Shu Zhang, Zhi-Chao Wang, Meng Duan, Zhao Zhang, Long-Zi Liu, Bo-Hao Zheng, Zhen-Bin Ding, Ai-Wu Ke, Da-Ming Gao, Ke Yuan, Jian Zhou, Jia Fan, Ruibin Xi, Qiang Gao
BACKGROUND & AIMS: Intrahepatic cholangiocarcinoma (ICC) is the second-most lethal primary liver cancer. Little is known about intratumoral heterogeneity (ITH) and its impact on ICC progression. We aim to investigate its ITH in hope of helping develop new therapeutic strategies. METHODS: We obtained 69 spatially distinct regions from 6 operable ICCs. Patient-derived primary cancer cells (PDPCs) were established for each region, followed by whole-exome sequencing(WES) and multi-level validation...
March 15, 2018: Journal of Hepatology
Azucena Mora, Francisco Javier García-Peña, María Pilar Alonso, Susana Pedraza-Diaz, Luis Miguel Ortega-Mora, Daniel Garcia-Parraga, Cecilia López, Susana Viso, Ghizlane Dahbi, Juan Marzoa, Martin J Sergeant, Vanesa García, Jorge Blanco
There is growing concern about the spreading of human microorganisms in relatively untouched ecosystems such as the Antarctic region. For this reason, three pinniped species (Leptonychotes weddellii, Mirounga leonina and Arctocephalus gazella) from the west coast of the Antartic Peninsula were analysed for the presence of Escherichia spp. with the recovery of 158 E. coli and three E. albertii isolates. From those, 23 harboured different eae variants (α1, β1, β2, ε1, θ1, κ, ο), including a bfpA-positive isolate (O49:H10-A-ST206, eae-k) classified as typical enteropathogenic E...
March 16, 2018: Scientific Reports
Anita Ferraretto, Michela Bottani, Paola De Luca, Laura Cornaghi, Francesca Arnaboldi, Margherita Maggioni, Amelia Fiorilli, Elena Donetti
An intestinal 70/30 Caco2/HT-29 co-culture was set up starting from the parental populations of differentiated cells to mimic the human intestinal epithelium. Co-culture was harvested at confluence 0 (T0) and at 3, 6, 10 and 14 days of post-confluence after plating (T3, T6, T10 and T14, respectively) for morphological and functional analysis. Transmission electron microscopy revealed different features from T0 to T14: microvilli and a complete junctional apparatus from T6, mucus granules from T3, as also confirmed by PAS/Alcian Blue staining...
March 14, 2018: Bioscience Reports
Sveva Grande, Alessandra Palma, Lucia Ricci-Vitiani, Anna Maria Luciani, Mariachiara Buccarelli, Mauro Biffoni, Agnese Molinari, Annarica Calcabrini, Emanuela D'Amore, Laura Guidoni, Roberto Pallini, Vincenza Viti, Antonella Rosi
Clustering of patient-derived glioma stem-like cells (GSCs) through unsupervised analysis of metabolites detected by magnetic resonance spectroscopy (MRS) evidenced three subgroups, namely clusters 1a and 1b, with high intergroup similarity and neural fingerprints, and cluster 2, with a metabolism typical of commercial tumor lines. In addition, subclones generated by the same GSC line showed different metabolic phenotypes. Aerobic glycolysis prevailed in cluster 2 cells as demonstrated by higher lactate production compared to cluster 1 cells...
2018: Stem Cells International
Ornella Franzese, Fiorenzo Battaini, Grazia Graziani, Lucio Tentori, Maria Luisa Barbaccia, Angelo Aquino, Mario Roselli, Maria Pia Fuggetta, Enzo Bonmassar, Francesco Torino
In recent years, immune checkpoint inhibitors (ICpI) have provided the ground to bring tumor immunity back to life thanks to their capacity to afford a real clinical benefit in terms of patient's survival. Essential to ICpI success is the presence of tumor-associated neoantigens generated by non-synonymous mutations, since a direct relationship between mutation load of malignant cells and susceptibility to ICpI has been confidently established. However, it has been also suggested that high intratumor heterogeneity (ITH) associated with subclonal neoantigens could not elicit adequate immune responses...
March 9, 2018: Pharmacological Research: the Official Journal of the Italian Pharmacological Society
Zhaolai Guo, Xudong Sun, Huini Xu
The inducible vectors pER8-Gateway-3Flag and pER8-Gateway-3Flag-SRDX have been subjected to considerable research in terms of the function of transcription factors (TFs) via transcription activity and repression, respectively. Approximately 1500 TFs have been identified in Arabidopsis thaliana genome. To identify their functions, loss-of-function and gain-of-function mutants were generated to analyze the phenotype. However, many loss-of-function mutants did not show any evident phenotype because of the functional redundancy within the TF family...
March 8, 2018: Planta
Yoshihiko Maeyama, Yui Taniguchi, Wataru Hayashi, Yusuke Ohsaki, Shunsuke Osaka, Shota Koide, Kiyoko Tamai, Yukiko Nagano, Yoshichika Arakawa, Noriyuki Nagano
In recent years, besides the widespread occurrence of extended-spectrum β-lactamase (ESBL)- and/or plasmid-mediated AmpC (pAmpC)-producing Enterobacteriaceae in both healthcare and community settings of humans, the third-generation cephalosporin (3GC)-resistant microbes have also been reported from companion animals worldwide. Here, we characterized ESBL- and/or pAmpC-producing Enterobacteriaceae clinical isolates from companion animals. Among the 487 clinical isolates mainly from urine of dogs and cats between May and September 2016, 104 non-repetitive isolates were resistant to the 3GC, and they consisted of 81 of 381 (21...
March 2018: Veterinary Microbiology
Meagan A Jacoby, Eric J Duncavage, Gue Su Chang, Christopher A Miller, Jin Shao, Kevin Elliott, Joshua Robinson, Robert S Fulton, Catrina C Fronick, Michelle O'Laughlin, Sharon E Heath, Iskra Pusic, John S Welch, Daniel C Link, John F DiPersio, Peter Westervelt, Timothy J Ley, Timothy A Graubert, Matthew J Walter
Allogeneic hematopoietic cell transplantation (alloHCT) is a potentially curative treatment for myelodysplastic syndromes (MDS), but patients who relapse after transplant have poor outcomes. In order to understand the contribution of tumor clonal evolution to disease progression,we applied exome and error-corrected targeted sequencing coupled with copy number analysis to comprehensively define changes in the clonal architecture of MDS in response to therapy using 51 serially acquired tumor samples from 9 patients who progressed after an alloHCT...
March 8, 2018: JCI Insight
Guihua Duan, Mengyue Shi, Lijuan Xie, Mingcui Xu, Yun Wang, Hongli Yan, Yuzheng Zhuge, Xiaoping Zou
The emergence of drug-resistant subclones remains the primary reason for tumor treatment failure. Some theories suggest that drug-resistant cell growth can be suppressed by drug-sensitive cells because resistant cells are less fit than sensitive cells in the absence of drug. We investigated fitness differences and their underlying mechanisms in cisplatin (ddp)-resistant cells and parental cells. We found that glutamine (Gln) consumption was substantially higher in ddp-resistant cells than that in sensitive cells, indicating that significantly fewer ddp-resistant cells than sensitive cells could be generated under the same Gln conditions...
March 6, 2018: Scientific Reports
Ana Carolina C Campos, Nathália L Andrade, Mithila Ferdous, Monika A Chlebowicz, Carla C Santos, Julio C D Correal, Jerome R Lo Ten Foe, Ana Cláudia P Rosa, Paulo V Damasco, Alex W Friedrich, John W A Rossen
Urinary tract infections (UTIs) are often caused by Escherichia coli . Their increasing resistance to broad-spectrum antibiotics challenges the treatment of UTIs. Whereas, E. coli ST131 is often multidrug resistant (MDR), ST69 remains susceptible to antibiotics such as cephalosporins. Both STs are commonly linked to community and nosocomial infections. E. coli phylogenetic groups B2 and D are associated with virulence and resistance profiles making them more pathogenic. Little is known about the population structure of E...
2018: Frontiers in Microbiology
L Xie, W J Hu, B R Liu
The accumulation of genomic and epigenetic changes gives rise to the tumorigenesis and progression. Currently, clonal evolution model and cancer stem cell model, two leading theories of caner origin, are becoming complementary to one another to explain the nature of tumor heterogeneity. Precision medicine that is based on the next generation sequencing and big data describes the phenomena of tumor heterogeneity more precisely. The future cancer therapy may need more comprehensive and dynamical understandings of the distinct subclones of tumor and follow the trends of cancer evolution...
February 23, 2018: Zhonghua Zhong Liu za Zhi [Chinese Journal of Oncology]
Matteo Forloni, Alex Y Liu, Narendra Wajapeyee
In this protocol, two oligonucleotides are used to prime DNA synthesis by a high-fidelity polymerase on a denatured plasmid template. The two oligonucleotides both contain the desired mutation and have the same starting and ending positions on opposite strands of the plasmid DNA. The entire lengths of both strands of the plasmid DNA are amplified in a linear fashion during several rounds of thermal cycling, generating a mutated plasmid containing staggered nicks on opposite strands. Because of the amount of template DNA used in the amplification reaction, the background of transformed colonies containing wild-type plasmid DNA can be quite high unless steps are taken to enrich for mutant molecules...
March 1, 2018: Cold Spring Harbor Protocols
Julien Levy, Cecilia Tamborindeguy, Giridhar Athrey, Douglas C Scheuring, Jeffrey W Koym, J Creighton Miller
OBJECTIVES: Potato has a large genetic diversity. This diversity is in part due to somaclonal variability that appears within potato selections for which tubers are used as seeds. However, the potato tetraploid genome, as well as the use of tubers for crop propagation, does not allow for easy genetic studies. The objective is to gain knowledge at the genomic level from standard Russet Norkotah and a subclonal Russet Norkotah selection TXNS278. DATA DESCRIPTION: In this report, we used RNA-seq, which allows genome-wide gene expression analysis to sequence the transcriptomes of the subclonal Russet Norkotah selection TXNS278 with standard Russet Norkotah grown in commercial fields...
March 1, 2018: BMC Research Notes
Samira Sabzi, Rahil Mashhadi, Mohammad Reza Pourmand
Background and Objectives: Enterococcus faecalis is the leading cause of several human infections. This opportunist pathogen expresses surface components that have various functions in the infection process including bacterial adhesion, lytic activity, and induction of host immune responses. EF0737, a novel cell wall associated protein, may play an important role in pathogenesis of E. faecalis , based on our experiments. This study was conducted to clone and express EF0737 and demonstrate its interaction with biotinylated plasma proteins and patients' sera...
December 2017: Iranian Journal of Microbiology
Deqiang Chen, Dongwei Wang, Chunling Xu, Chun Chen, Junyi Li, Wenjia Wu, Xin Huang, Hui Xie
Controlling Radopholus similis, an important phytopathogenic nematode, is a challenge worldwide. Herein, we constructed a metagenomic fosmid library from the rhizosphere soil of banana plants, and six clones with protease activity were obtained by functionally screening the library. Furthermore, subclones were constructed using the six clones, and three protease genes with nematicidal activity were identified: pase1, pase4, and pase6. The pase4 gene was successfully cloned and expressed, demonstrating that the protease PASE4 could effectively degrade R...
February 27, 2018: Applied Microbiology and Biotechnology
Kiran Tawana, Michael W Drazer, Jane E Churpek
Comprehensive genomic profiling of myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) cases have enabled the detection and differentiation of driver and subclonal mutations, informed risk prognostication, and defined targeted therapies. These insights into disease biology, and management have made multigene-acquired mutation testing a critical part of the diagnostic assessment of patients with sporadic MDS and AML. More recently, our understanding of the role of an increasing number of inherited genetic factors on MDS/AML risk and management has rapidly progressed...
February 27, 2018: Leukemia: Official Journal of the Leukemia Society of America, Leukemia Research Fund, U.K
Ikram Ullah, Govindasamy-Muralidharan Karthik, Amjad Alkodsi, Una Kjällquist, Gustav Stålhammar, John Lövrot, Nelson-Fuentes Martinez, Jens Lagergren, Sampsa Hautaniemi, Johan Hartman, Jonas Bergh
Metastatic breast cancers are still incurable. Characterizing the evolutionary landscape of these cancers, including the role of metastatic axillary lymph nodes (ALNs) in seeding distant organ metastasis, can provide a rational basis for effective treatments. Here, we have described the genomic analyses of the primary tumors and metastatic lesions from 99 samples obtained from 20 patients with breast cancer. Our evolutionary analyses revealed diverse spreading and seeding patterns that govern tumor progression...
February 26, 2018: Journal of Clinical Investigation
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