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Mammalian cell culture

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https://www.readbyqxmd.com/read/29141004/lack-of-mitochondrial-muts-homolog-1-in-toxoplasma-gondii-disrupts-maintenance-and-fidelity-of-mitochondrial-dna-and-reveals-metabolic-plasticity
#1
Tamila Garbuz, Gustavo Arrizabalaga
The importance of maintaining the fidelity of the mitochondrial genome is underscored by the presence of various repair pathways within this organelle. Presumably, the repair of mitochondrial DNA would be of particular importance in organisms that possess only a single mitochondrion, like the human pathogens Plasmodium falciparum and Toxoplasma gondii. Understanding the machinery that maintains mitochondrial DNA in these parasites is of particular relevance, as mitochondrial function is a validated and effective target for anti-parasitic drugs...
2017: PloS One
https://www.readbyqxmd.com/read/29134714/identification-of-perivascular-and-stromal-mesenchymal-stem-progenitor-cells-in-porcine-endometrium
#2
J Wiater, M Niedziela, A Posmysz, K Wartalski, B Gajda, Z Smorąg, Z Rajfur, J Karasiński
Mammalian uterus contains a population of mesenchymal stem/progenitor cells that likely contribute to endometrial regeneration during each reproductive cycle. In human and mouse, they reside in perivascular, epithelial and stromal compartments of the endometrial functionalis and basalis. Here, we aimed to identify tissue resident cells expressing mesenchymal stem cell markers CD29, CD44, CD90, CD105, CD140b and CD146 in the porcine endometrium. We used single immunofluorescence and Western blotting. Each of these markers was detected in small cells surrounding endometrial blood vessels...
November 14, 2017: Reproduction in Domestic Animals, Zuchthygiene
https://www.readbyqxmd.com/read/29134459/risk-mitigation-in-preventing-adventitious-agent-contamination-of-mammalian-cell-cultures
#3
Masaru Shiratori, Robert Kiss
Industrial-scale mammalian cell culture processes have been contaminated by viruses during the culturing phase. Although the historical frequency of such events has been quite low, the impact of contamination can be significant for the manufacturing company and for the supply of the product to patients. This chapter discusses sources of adventitious agent contamination risk in a cell culture process, provides a semiquantitative assessment of such risks, and describes potential process barriers that can be used to reduce contamination risk...
November 14, 2017: Advances in Biochemical Engineering/biotechnology
https://www.readbyqxmd.com/read/29134442/endothelial-tnf-%C3%AE-induction-by-hsp60-secreted-from-thp-1-monocytes-exposed-to-hyperglycaemic-conditions
#4
Ryan Dennis Martinus, Julie Goldsbury
A non-resolving inflammation of the endothelium is recognised to be an important process leading to atherosclerosis. In diabetes, this process is thought to account for a significant number of cardiovascular disease-associated death and disability. However, the molecular mechanisms by which diabetes contributes to endothelial inflammation remain to be established. Whilst there is some evidence linking hyperglycaemia-induced reactive oxygen species (ROS) formation by the mitochondrial electron-transport chain to oxidative stress, cellular injury and apoptosis in the endothelium, a clear link to endothelium inflammation has not yet been established...
November 13, 2017: Cell Stress & Chaperones
https://www.readbyqxmd.com/read/29133401/multiple-legionella-pneumophila-effector-virulence-phenotypes-revealed-through-high-throughput-analysis-of-targeted-mutant-libraries
#5
Stephanie R Shames, Luying Liu, James C Havey, Whitman B Schofield, Andrew L Goodman, Craig R Roy
Legionella pneumophila is the causative agent of a severe pneumonia called Legionnaires' disease. A single strain of L. pneumophila encodes a repertoire of over 300 different effector proteins that are delivered into host cells by the Dot/Icm type IV secretion system during infection. The large number of L. pneumophila effectors has been a limiting factor in assessing the importance of individual effectors for virulence. Here, a transposon insertion sequencing technology called INSeq was used to analyze replication of a pool of effector mutants in parallel both in a mouse model of infection and in cultured host cells...
November 13, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/29128351/drosophila-models-of-fop-provide-mechanistic-insight
#6
REVIEW
Viet Le, Edward Anderson, Takuya Akiyama, Kristi A Wharton
Fibrodysplasia ossificans progressiva (FOP) is a rare bone disease characterized by episodic events of heterotopic ossification (HO). All cases of FOP have been attributed to mutations in the ACVR1 gene that render the encoded BMP type I ALK2 receptor hypersensitive, resulting in the activation of BMP signaling, at inappropriate times in inappropriate locations. The episodic or sporadic nature of HO associated with FOP rests with the occurrence of specific 'triggers' that push the hypersensitive ALK2-FOP receptor into full signaling mode...
November 8, 2017: Bone
https://www.readbyqxmd.com/read/29126263/notch-signaling-regulates-differentiation-and-steroidogenesis-in-female-mouse-ovarian-granulosa-cells
#7
Rexxi D Prasasya, Kelly E Mayo
The Notch pathway is a highly conserved juxtacrine signaling mechanism that is important for many cellular processes during development, including differentiation and proliferation. While Notch is important during ovarian follicle formation and early development, its functions during the gonadotropin-dependent stages of follicle development are largely unexplored. We observed positive regulation of Notch activity and expression of Notch ligands and receptors following activation of the LH-receptor in prepubertal mouse ovary...
November 8, 2017: Endocrinology
https://www.readbyqxmd.com/read/29126233/endothelin-1-promotes-human-germinal-vesicle-stage-oocyte-maturation-by-downregulating-connexin-26-expression-in-cumulus-cells
#8
Long Cui, Jiajie Shen, Li Fang, Xiaodan Mao, Hanzhi Wang, Yinghui Ye
STUDY QUESTION: Does endothelin-1 (ET-1) promote human oocyte maturation and by what mechanism? SUMMARY ANSWER: Addition of ET-1 to the medium in which human germinal vesicle (GV)-stage immature oocytes are cultured enhances the GV breakdown (GVBD) rate; the resumption of meiosis may be initiated by ET-1 downregulating the expression of connexin-26 (Cx26) in cumulus cells via endothelin receptor type B (ETRB), leading to decreased cAMP levels in the oocyte. WHAT IS KNOWN ALREADY: The paracrine factor ET-1 is secreted by ovarian somatic cells in pre-ovulatory follicles and regulates oocyte maturation in mice...
November 8, 2017: Molecular Human Reproduction
https://www.readbyqxmd.com/read/29123210/plasticity-of-the-inner-cell-mass-in-mouse-blastocyst-is-restricted-by-the-activity-of-fgf-mapk-pathway
#9
M Wigger, K Kisielewska, K Filimonow, B Plusa, M Maleszewski, A Suwińska
In order to ensure successful development, cells of the early mammalian embryo must differentiate to either trophectoderm (TE) or inner cell mass (ICM), followed by epiblast (EPI) or primitive endoderm (PE) specification within the ICM. Here, we deciphered the mechanism that assures the correct order of these sequential cell fate decisions. We revealed that TE-deprived ICMs derived from 32-cell blastocysts are still able to reconstruct TE during in vitro culture, confirming totipotency of ICM cells at this stage...
November 9, 2017: Scientific Reports
https://www.readbyqxmd.com/read/29121962/direct-interaction-with-14-3-3%C3%AE-promotes-surface-expression-of-best1-channel-in-astrocyte
#10
Soo-Jin Oh, Junsung Woo, Young-Sun Lee, Minhee Cho, Eunju Kim, Nam-Chul Cho, Jae-Yong Park, Ae Nim Pae, C Justin Lee, Eun Mi Hwang
BACKGROUND: Bestrophin-1 (Best1) is a calcium-activated anion channel (CAAC) that is expressed broadly in mammalian tissues including the brain. We have previously reported that Best1 is expressed in hippocampal astrocytes at the distal peri-synaptic regions, called microdomains, right next to synaptic junctions, and that it disappears from the microdomains in Alzheimer's disease mouse model. Although Best1 appears to be dynamically regulated, the mechanism of its regulation and modulation is poorly understood...
November 9, 2017: Molecular Brain
https://www.readbyqxmd.com/read/29119608/synthetic-aav-crispr-vectors-for-blocking-hiv-1-expression-in-persistently-infected-astrocytes
#11
Christine Kunze, Kathleen Börner, Eike Kienle, Tanja Orschmann, Ejona Rusha, Martha Schneider, Milena Radivojkov-Blagojevic, Micha Drukker, Sabrina Desbordes, Dirk Grimm, Ruth Brack-Werner
Astrocytes, the most abundant cells in the mammalian brain, perform key functions and are involved in several neurodegenerative diseases. The human immunodeficiency virus (HIV) can persist in astrocytes, contributing to the HIV burden and neurological dysfunctions in infected individuals. While a comprehensive approach to HIV cure must include the targeting of HIV-1 in astrocytes, dedicated tools for this purpose are still lacking. Here we report a novel Adeno-associated virus-based vector (AAV9P1) with a synthetic surface peptide for transduction of astrocytes...
November 9, 2017: Glia
https://www.readbyqxmd.com/read/29118915/a-novel-culture-system-robustly-maintained-pluripotency-of-embryonic-stem-cells-and-accelerated-somatic-reprogramming-by-activating-wnt-signaling
#12
Shaojie Li, Xiao Lu, Haipeng He, Rong Cui, Xianxin Wang, Xiaoyun Wang, Xia Wu
Wnt signaling is intrinsic to embryonic stem cell self-renewal and mammalian development. However, the effects of wnts on ES cells self-renewal and iPS cells transduction was not clearly understood. In this study, L-Wnt3a cells that secreted activated Wnt3a protein into medium were used to produce Wnt3a condition medium (Wnt3a-CM) or feeder layer for ES cells cultivation and iPS cells transduction. The results showed that L-Wnt3a cells as feeder layer significantly promoted establishment of ES cell lines and generation of iPS cells...
2017: American Journal of Translational Research
https://www.readbyqxmd.com/read/29118117/a-y161f-hemagglutinin-substitution-increases-thermostability-and-improves-yields-of-2009-h1n1-influenza-a-virus-in-cells
#13
Feng Wen, Lei Li, Nan Zhao, Meng-Jung Chiang, Hang Xie, Jim Cooley, Richard Webby, Peng George Wang, Xiu-Feng Wan
Vaccination is the primary strategy for influenza prevention and control. Yet egg-based vaccines, the predominant production platform, have several disadvantages including the emergence of viral antigenic variants that can be induced during egg passage. These limitations have prompted development of cell-based vaccines which themselves are not without issue. Most importantly, vaccine seed viruses often do not grow efficiently in mammalian cell lines. Here we aimed to identify novel high-yield signatures for influenza viruses in continuous Madin-Darby canine kidney (MDCK) and Vero cells...
November 8, 2017: Journal of Virology
https://www.readbyqxmd.com/read/29116521/high-throughput-igg-reformatting-and-expression
#14
Chao-Guang Chen, Georgina Sansome, Michael J Wilson, Con Panousis
We have recently described a one-step zero-background IgG reformatting method that enables the rapid reformatting of phage-displayed antibody fragments into a single-mammalian cell expression vector for full IgG expression (Chen et al. Nucleic Acids Res 42:e26, 2014). The strategy utilizes our unique positive selection method, referred to as insert-tagged (InTag) positive selection, where a positive selection marker (e.g. chloramphenicol-resistance gene) is cloned together with the antibody inserts into the expression vector...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29113465/porcine-neural-progenitor-cells-derived-from-tissue-at-different-gestational-ages-can-be-distinguished-by-global-transcriptome
#15
Jing Yang, Steven Menges, Ping Gu, Ronald Tongbai, Melissa Samuel, Randall S Prather, Henry Klassen
The impact of gestational age on mammalian neural progenitor cells is potentially important for both an understanding of neural development and the selection of donor cells for novel cell-based treatment strategies. In terms of the latter, it can be problematic to rely entirely on rodent models in which the gestational period is significantly shorter and the brain much smaller than is the case in humans. Here, we analyzed pig brain progenitor cells (pBPCs) harvested at 2 different gestational ages (E45 and E60) using gene expression profiles, obtained by microarray analysis and quantitative polymerase chain reaction (qPCR), across time in culture...
September 2017: Cell Transplantation
https://www.readbyqxmd.com/read/29112296/live-faecalibacterium-prausnitzii-induces-greater-tlr2-and-tlr2-6-activation-than-the-dead-bacterium-in-an-apical-anaerobic-co-culture-system
#16
Eva Maier, Rachel C Anderson, Eric Altermann, Nicole C Roy
Inappropriate activation of intestinal innate immune receptors, such as Toll-like receptors (TLRs), by pathogenic bacteria is linked to chronic inflammation. In contrast, a 'tonic' level of TLR activation by commensal bacteria is required for intestinal homeostasis. A technical challenge when studying this activation in vitro is the co-culturing of oxygen-requiring mammalian cells with obligate anaerobic commensal bacteria. To overcome this, we used a novel apical anaerobic co-culture system to successfully adapt a TLR activation assay to be conducted in conditions optimised for both cell types...
November 7, 2017: Cellular Microbiology
https://www.readbyqxmd.com/read/29110761/the-cell-biology-of-apol1
#17
REVIEW
John F O'Toole, Leslie A Bruggeman, Sethu Madhavan, John R Sedor
The association of variants in the APOL1 gene, which encodes apolipoprotein L1 (APOL1), with progressive nondiabetic kidney diseases in African Americans has prompted intense investigation into the function(s) of APOL1. APOL1 is an innate immune effector that protects human beings from infection by some trypanosomal parasites. We review the data characterizing APOL1 trypanolytic function, which has been a basis for studies of APOL1 function in mammalian cells. Subsequently, we discuss the studies that use animal models, mammalian cell culture models, and kidney biopsy tissue to discover the mechanisms of variant APOL1-associated kidney diseases...
November 2017: Seminars in Nephrology
https://www.readbyqxmd.com/read/29109483/safety-of-nonporous-silica-nanoparticles-in-human-corneal-endothelial-cells
#18
Ja-Yeon Kim, Joo-Hee Park, Martha Kim, Hyejoong Jeong, Jinkee Hong, Roy S Chuck, Choul Yong Park
Nonporous silica nanoparticles (SiNPs) are promising drug carrier platforms for intraocular drug delivery. In this study, we investigated the safety of three different sizes of SiNPs (50, 100, and 150 nm) in a human corneal endothelial cell (HCEC) line, B4G12. The HCECs were exposed to different concentrations (0, 25, 50, and 100 µg/ml) of three sizes of SiNPs for up to 48 h. Cellular viability, autophagy, lactate dehydrogenase (LDH) assay, and mammalian target of rapamycin (mTOR) pathway activation were evaluated...
November 6, 2017: Scientific Reports
https://www.readbyqxmd.com/read/29107996/epigenetic-memory-via-concordant-dna-methylation-is-inversely-correlated-to-developmental-potential-of-mammalian-cells
#19
Minseung Choi, Diane P Genereux, Jamie Goodson, Haneen Al-Azzawi, Shannon Q Allain, Noah Simon, Stan Palasek, Carol B Ware, Chris Cavanaugh, Daniel G Miller, Winslow C Johnson, Kevin D Sinclair, Reinhard Stöger, Charles D Laird
In storing and transmitting epigenetic information, organisms must balance the need to maintain information about past conditions with the capacity to respond to information in their current and future environments. Some of this information is encoded by DNA methylation, which can be transmitted with variable fidelity from parent to daughter strand. High fidelity confers strong pattern matching between the strands of individual DNA molecules and thus pattern stability over rounds of DNA replication; lower fidelity confers reduced pattern matching, and thus greater flexibility...
November 6, 2017: PLoS Genetics
https://www.readbyqxmd.com/read/29107535/genetically-encoded-tools-for-optical-dissection-of-the-mammalian-cell-cycle
#20
Asako Sakaue-Sawano, Masahiro Yo, Naoki Komatsu, Toru Hiratsuka, Takako Kogure, Tetsushi Hoshida, Naoki Goshima, Michiyuki Matsuda, Hiroyuki Miyoshi, Atsushi Miyawaki
Eukaryotic cells spend most of their life in interphase of the cell cycle. Understanding the rich diversity of metabolic and genomic regulation that occurs in interphase requires the demarcation of precise phase boundaries in situ. Here, we report the properties of two genetically encoded fluorescence sensors, Fucci(CA) and Fucci(SCA), which enable real-time monitoring of interphase and cell-cycle biology. We re-engineered the Cdt1-based sensor from the original Fucci system to respond to S phase-specific CUL4(Ddb1)-mediated ubiquitylation alone or in combination with SCF(Skp2)-mediated ubiquitylation...
October 25, 2017: Molecular Cell
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