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Lentiviral

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https://www.readbyqxmd.com/read/28810880/nlrp7-contributes-to-in-vitro-decidualization-of-endometrial-stromal-cells
#1
Jyun-Yuan Huang, Pei-Hsiu Yu, Yueh-Chun Li, Pao-Lin Kuo
BACKGROUND: Nucleotide-binding oligomerization domain (NACHT), leucine rich repeat (LRR) and pyrin domain (PYD) 7 containing protein, NLRP7, is a member of the NLR family which serves as innate immune sensors. Mutations and genetic variants of NLRP7 have been found in women with infertility associated conditions, such as recurrent hydatidiform mole, recurrent miscarriage, and preeclampsia. Decidualization of endometrial stromal cells is a hallmark of tissue remodeling to support embryo implantation and proper placental development...
August 15, 2017: Reproductive Biology and Endocrinology: RB&E
https://www.readbyqxmd.com/read/28809019/lentiviral-mediated-transgenesis-in-songbirds
#2
Wan-Chun Liu, Marian Hruska-Plochan, Atsushi Miyanohara
Transgenesis involves the insertion of an exogenous gene into an animal's genome, which allows the identification of the expressed phenotypes in brain function or behavior. Lentiviral-mediated transgenesis offers unique transduction potency making it possible to deliver and stably integrate transgenes into a wide variety of dividing and nondividing cells. The ability to establish long-term expression of such transgenes allows their use for transgenesis which is especially useful in organisms lacking quality pluripotent stem cell lines and which is otherwise difficult to produce via traditional pronuclear microinjection, such as songbirds...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28809018/fluorescent-quail-a-transgenic-model-system-for-the-dynamic-study-of-avian-development
#3
David Huss, Rusty Lansford
Real-time four-dimensional (4D, xyzt) imaging of cultured avian embryos is an ideal method for investigating the complex movements of cells and tissues during early morphogenesis. While methods that transiently label cells, such as electroporation, are highly useful for dynamic imaging, they can also be limiting due to the number and type of cells that can be effectively targeted. In contrast, the heritable, stable, and long-term expression of a fluorescent protein driven by the exogenous promoter of a transgene overcomes these challenges...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28808970/genome-wide-crispr-cas9-screening-for-high-throughput-functional-genomics-in-human-cells
#4
Shiyou Zhu, Yuexin Zhou, Wensheng Wei
It is highly desirable to identify gene's function in a high-throughput fashion, and the CRISPR/Cas9 system has been harnessed to meet such a need. Here, we describe a general method to generate genome-scale lentiviral single-guide RNA (sgRNA) library and conduct a pooled function-based screening in human cells. This protocol would be of interest to researchers to rapidly identify genes in a variety of biological processes.
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28808928/regulation-of-survival-motor-neuron-protein-by-the-nuclear-factor-kappa-b-pathway-in-mouse-spinal-cord-motoneurons
#5
Saravanan Arumugam, Stefka Mincheva-Tasheva, Ambika Periyakaruppiah, Sandra de la Fuente, Rosa M Soler, Ana Garcera
Survival motor neuron (SMN) protein deficiency causes the genetic neuromuscular disorder spinal muscular atrophy (SMA), characterized by spinal cord motoneuron degeneration. Since SMN protein level is critical to disease onset and severity, analysis of the mechanisms involved in SMN stability is one of the central goals of SMA research. Here, we describe the role of several members of the NF-κB pathway in regulating SMN in motoneurons. NF-κB is one of the main regulators of motoneuron survival and pharmacological inhibition of NF-κB pathway activity also induces mouse survival motor neuron (Smn) protein decrease...
August 14, 2017: Molecular Neurobiology
https://www.readbyqxmd.com/read/28807730/development-of-a-dual-reporter-screening-assay-for-distinguishing-the-inhibition-of-hiv-tat-mediated-transcription-from-off-target-effects
#6
YoungHyun Shin, Byeong-Sun Choi, Kyung-Chang Kim, Chun Kang, Kisoon Kim, Cheol-Hee Yoon
Human immunodeficiency virus (HIV) encodes a transcription trans-activator (Tat) with an essential role in the transcriptional elongation of viral RNA based on the viral promoter long terminal repeat (LTR). Tat-mediated transcription is conserved and can be distinguished from host transcription, so it is a therapeutic target for combating HIV replication. Traditional screening assays for Tat-mediated transcriptional inhibitors are based on the biochemical properties of Tat and transactivation-responsive RNA...
August 11, 2017: Journal of Virological Methods
https://www.readbyqxmd.com/read/28807012/protein-deubiquitinase-usp7-is-required-for-osteogenic-differentiation-of-human-adipose-derived-stem-cells
#7
Yiman Tang, Longwei Lv, Wenyue Li, Xiao Zhang, Yong Jiang, Wenshu Ge, Yongsheng Zhou
BACKGROUND: Human adipose-derived stem cells (hASCs) are multipotent progenitor cells with self-renewal capabilities and multilineage differentiation potential, including osteogenesis. Although protein deubiquitinases have been linked to stem cell fate determination, whether protein deubiquitination contributes to lineage commitment during osteogenic differentiation of hASCs remains to be investigated. The objective of this study was to evaluate the effects of the ubiquitin specific protease 7 (USP7) on osteogenic differentiation of hASCs...
August 14, 2017: Stem Cell Research & Therapy
https://www.readbyqxmd.com/read/28807003/overexpression-of-mir-21-in-stem-cells-improves-ovarian-structure-and-function-in-rats-with-chemotherapy-induced-ovarian-damage-by-targeting-pdcd4-and-pten-to-inhibit-granulosa-cell-apoptosis
#8
Xiafei Fu, Yuanli He, Xuefeng Wang, Dongxian Peng, Xiaoying Chen, Xinran Li, Qing Wang
BACKGROUND: Chemotherapy-induced premature ovarian failure (POF) is a severe complication affecting tumor patients at a childbearing age. Mesenchymal stem cells (MSCs) can partially restore the ovarian structure and function damaged by chemotherapy. miR-21 is a microRNA that can regulate cell apoptosis. This study discusses the repair effect and mechanism of MSCs overexpressing miR-21 on chemotherapy-induced POF. METHODS: Rat MSCs and granulosa cells (GCs) were isolated in vitro...
August 14, 2017: Stem Cell Research & Therapy
https://www.readbyqxmd.com/read/28805067/the-lncrna-nespas-is-associated-with-osteoarthritis-progression-and-serves-as-a-potential-new-prognostic-biomarker
#9
Sujung Park, Myeungsoo Lee, Churl-Hong Chun, Eun-Jung Jin
INTRODUCTION: In this article, we explored the hypothesis that the long noncoding RNA, Nespas, promotes osteoarthritis (OA) by supporting abnormal lipid metabolism in human chondrocytes. MATERIALS AND METHODS: Human articular chondrocytes from osteoarthritis patients were used and expression level of Nespas were determined by real-time polymerase chain reaction. Introduction of Nespas and Nespas-associated genes/miRNAs were performed by using a lentiviral system...
August 1, 2017: Cartilage
https://www.readbyqxmd.com/read/28803141/the-poly-proline-tail-of-sivmac-vpx-provides-gain-of-function-for-resistance-to-a-cryptic-proteasome-dependent-degradation-pathway
#10
Nannan Zhang, Haoran Guo, Jiaxin Yang, Guanchen Liu, Shuang Li, Siying Li, Dongyin Wang, Rui Li, Chang Shu, Hongmei Xu, Zhentong Wei, Honglan Huang, Songling Zhang, Pujun Gao, Shan Cen, Richard Markham, Yongsheng Wang, Xiao-Fang Yu, Wei Wei
The lentiviral accessory protein Vpx is critical for viral infection of myeloid cells and acts by hijacking CRL4(DCAF1) E3 ubiquitin ligase to induce the degradation of the host restriction factor SAMHD1. It has been observed that the sequences from HIV-2 and SIVsmm/SIVmac Vpx contain a poly-proline tail which is distinct from other SIV Vpx proteins. However, the role of this region in Vpx function is controversial. Herein, we found proteasome-dependent degradation of a Vpx mutant lacking the poly-proline tail in the nucleus in a CRL4(DCAF1) E3 ligase-independent fashion...
August 10, 2017: Virology
https://www.readbyqxmd.com/read/28801899/bioluminescence-monitoring-of-promoter-activity-in-vitro-and-in-vivo
#11
Juliette M K M Delhove, Rajvinder Karda, Kate E Hawkins, Lorna M FitzPatrick, Simon N Waddington, Tristan R McKay
The application of luciferase reporter genes to provide quantitative outputs for the activation of promoters is a well-established technique in molecular biology. Luciferase catalyzes an enzymatic reaction, which in the presence of the substrate luciferin produces photons of light relative to its molar concentration. The luciferase transgene can be genetically inserted at the first intron of a target gene to act as a surrogate for the gene's endogenous expression in cells and transgenic mice. Alternatively, promoter sequences can be excised and/or amplified from genomic sources or constructed de novo and cloned upstream of luciferase in an expression cassette transfected into cells...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28801671/the-cpg-sites-of-the-cbx3-ubiquitous-chromatin-opening-element-are-critical-structural-determinants-for-the-anti-silencing-function
#12
Jessica Kunkiel, Natascha Gödecke, Mania Ackermann, Dirk Hoffmann, Axel Schambach, Nico Lachmann, Dagmar Wirth, Thomas Moritz
Suppression of therapeutic transgene expression from retroviral gene therapy vectors by epigenetic defence mechanisms represents a problem that is particularly encountered in pluripotent stem cells (PSCs) and their differentiated progeny. Transgene expression in these cells, however, can be stabilised by CpG-rich ubiquitous chromatin opening elements (UCOEs). In this context we recently demonstrated profound anti-silencing properties for the small (679 bp) CBX3-UCO element and we now confirmed this observation in the context of the defined murine chromosomal loci ROSA26 and TIGRE...
August 11, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28801449/engraftment-and-in-vivo-proliferation-advantage-of-gene-corrected-mobilized-cd34-cells-from-fanconi-anemia-patients
#13
Paula Río, Susana Navarro, Guillermo Guenechea, Rebeca Sánchez-Domínguez, Maria Luisa Lamana, Rosa Yañez, Jose A Casado, Parinda A Mehta, Maria Roser Pujol, Jordi Surrallés, Sabine Charrier, Anne Galy, José C Segovia, Cristina Díaz de Heredia, Julián Sevilla, Juan Bueren
Previous Fanconi anemia (FA) gene therapy studies have failed to demonstrate engraftment of gene corrected hematopoietic stem and progenitor cells (HSPC) from FA patients, either after autologous transplantation or infusion into immunodeficient mice. In this study we demonstrate that a validated short transduction protocol of G-CSF plus plerixafor-mobilized CD34(+) cells from FA-A patients with a therapeutic FANCA-lentiviral vector corrects the phenotype of in vitro cultured hematopoietic progenitor cells. Transplantation of transduced FA CD34(+) cells into immunodeficient mice resulted in reproducible engraftment of myeloid, lymphoid and CD34(+) cells...
August 11, 2017: Blood
https://www.readbyqxmd.com/read/28801289/-lkb1-regulates-epithelial-mesenchymal-transition-in-peutz-jeghers-hamartoma-and-intestinal-epithelial-cells
#14
Chao Zhong, Liang Peng, Ran Li, Jing Chen, Xin-Qi Chen, Di Zeng, Xiao-Ping Xu, Zhi-Qing Wang, Chu-di Chen, Ya-Dong Wang, Ai-Min Li, Si-de Liu, Bao-Ping Wu
OBJECTIVE: To investigate the molecular mechanism by which LKB1 regulates epithelial-mesenchymal transition (EMT) in Peutz-Jeghers hamartoma and intestinal epithelial cells. METHODS: Immunohistochemistry was used to detect gene expression of LKB1, E-cadherin, and vimentin in 20 hamartoma tissues and 10 normal intestinal tissues, and collagen fiber deposition was analyzed using Masson trichrome staining. Normal intestinal epithelial NCM460 cells were transfected with LKB1 shRNA plasmid or negative control via lentiviral vectors, and the role of LKB1 in cell polarization and migration were determined using CCK8 and Transwell assays...
August 20, 2017: Nan Fang Yi Ke da Xue Xue Bao, Journal of Southern Medical University
https://www.readbyqxmd.com/read/28800762/the-novel-cannabinoid-receptor-gpr55-mediates-anxiolytic-like-effects-in-the-medial-orbital-cortex-of-mice-with-acute-stress
#15
Qi-Xin Shi, Liu-Kun Yang, Wen-Long Shi, Lu Wang, Shi-Meng Zhou, Shao-Yu Guan, Ming-Gao Zhao, Qi Yang
The G protein-coupled receptor 55 (GPR55) is a novel cannabinoid receptor, whose exact role in anxiety remains unknown. The present study was conducted to explore the possible mechanisms by which GPR55 regulates anxiety and to evaluate the effectiveness of O-1602 in the treatment of anxiety-like symptoms. Mice were exposed to two types of acute stressors: restraint and forced swimming. Anxiety behavior was evaluated using the elevated plus maze and the open field test. We found that O-1602 alleviated anxiety-like behavior in acutely stressed mice...
August 11, 2017: Molecular Brain
https://www.readbyqxmd.com/read/28800590/assessment-and-optimization-of-theileria-parva-sporozoite-full-length-p67-antigen-expression-in-mammalian-cells
#16
Giulia Tebaldi, Laura B Williams, Andrea E Verna, Francesca Macchi, Valentina Franceschi, Lindsay M Fry, Donald P Knowles, Gaetano Donofrio
Delivery of various forms of recombinant Theileria parva sporozoite antigen (p67) has been shown to elicit antibody responses in cattle capable of providing protection against East Coast fever, the clinical disease caused by T. parva. Previous formulations of full-length and shorter recombinant versions of p67 derived from bacteria, insect, and mammalian cell systems are expressed in non-native and highly unstable forms. The stable expression of full-length recombinant p67 in mammalian cells has never been described and has remained especially elusive...
August 11, 2017: PLoS Neglected Tropical Diseases
https://www.readbyqxmd.com/read/28799202/immune-tolerance-induced-by%C3%A2-platelet-targeted-factor-viii-gene-therapy-in-hemophilia-a-mice-is-cd4%C3%A2-t-cell-mediated
#17
Y Chen, X Luo, J A Schroeder, J Chen, C K Baumgartner, J Hu, Q Shi
BACKGROUND: Immune responses are a major concern in gene therapy. Our previous studies demonstrated that platelet-targeted FVIII (2bF8) gene therapy together with in vivo drug-selection of transduced cells can rescue the bleeding diathesis and induce immune tolerance in FVIII(null) mice. OBJECTIVE: To investigate whether non-selectable 2bF8 lentiviral vector (LV) for the induction of platelet-FVIII expression is sufficient to induce immune tolerance and how immune tolerance is induced after 2bF8LV gene therapy...
August 11, 2017: Journal of Thrombosis and Haemostasis: JTH
https://www.readbyqxmd.com/read/28796255/pdi-is-an-essential-redox-sensitive-activator-of-perk-during-the-unfolded-protein-response-upr
#18
Philip Kranz, Fabian Neumann, Alexandra Wolf, Fabian Classen, Mosche Pompsch, Tobias Ocklenburg, Jennifer Baumann, Kirsten Janke, Melanie Baumann, Kirsten Goepelt, Helena Riffkin, Eric Metzen, Ulf Brockmeier
Endoplasmic reticulum (ER) stress leads to activation of the unfolded protein response (UPR) that results in transient suppression of protein translation to allow recovery but leads to cell death when stress cannot be resolved. Central to initiation of the UPR is the activation of the ER transmembrane kinase protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK). Here we report that the thiol oxidoreductase ERp57 and protein disulfide isomerase-A1 (PDI), which belong to the same family of luminal ER oxidoreductases, have strikingly opposing roles in the regulation of PERK function...
August 10, 2017: Cell Death & Disease
https://www.readbyqxmd.com/read/28796105/overexpression-of-glypican-3-promotes-proliferation-regulates-cell-cycle-progression-and-inhibits-apoptosis-of-human-fetal-osteoblastic-cell-line-1-19
#19
Tianyi Cai, Yingzhi Wu, Ronghu Ke, Junyi Yang, Abdulsamad Ghanem, Xiongzheng Mu
Craniosynostosis is a complex disease condition, which involves premature fusion of cranial vault sutures and lacks desirable treatment. Previous studies have demonstrated decreased proliferation rate of osteoblasts and downregulated expression of glypican 3 (GPC3) in syndromic craniosynostosis patients. In this study, quantitative and qualitative analysis were utilized to assess the effect of GPC3 in human fetal osteoblastic cell line, hFOB 1.19. Lentiviral transfection efficiency with green fluorescent protein images was obtained after 72 hours...
August 8, 2017: Journal of Craniofacial Surgery
https://www.readbyqxmd.com/read/28795673/-under-expression-of-lats1-promotes-the-differentiation-proliferation-and-migration-of-mesenchymal-stem-cells-by-inhibition-the-hippo-signaling-pathway-in-vitro
#20
Lang Li, Liang Dong, Jiaojie Hui, Fei Gao, Qiuhui Wang, Lan Yang, Jiangqian Zhang, Jie Yan
OBJECTIVE: To explore the effects of under-expression of large tumor suppressor 1 (LATS1) on activation of Hippo signaling pathway and differentiation, proliferation, migration of bone marrow mesenchymal stem cells (mMSCs) of mice in vitro. METHODS: mMSCs of C57BL/6 mice were divided into normal control (MSC) group, empty vector control (MSC-GFP) group, LATS1-over-expressing (MSC-LATS1) group, empty vector without LATS1 shRNA control (MSC-shControl) group and LATS1-under-expressing (MSC-shLATS1) group...
August 2017: Zhonghua Wei Zhong Bing Ji Jiu Yi Xue
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