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https://www.readbyqxmd.com/read/29165030/in-mammalian-skeletal-muscle-phosphorylation-of-tomm22-by-protein-kinase-csnk2-ck2-controls-mitophagy
#1
Bojana Kravic, Angelika B Harbauer, Vanina Romanello, Luca Simeone, F-Nora Vögtle, Tobias Kaiser, Marion Straubinger, Danyil Huraskin, Martin Böttcher, Cristina Cerqua, Eva Denise Martin, Daniel Poveda-Huertes, Andreas Buttgereit, Adam J Rabalski, Dieter Heuss, Rüdiger Rudolf, Oliver Friedrich, David Litchfield, Michael Marber, Leonardo Salviati, Dimitrios Mougiakakos, Winfried Neuhuber, Marco Sandri, Chris Meisinger, Said Hashemolhosseini
In yeast, Tom22 the central component of the TOMM (translocase of outer mitochondrial membrane) receptor complex is responsible for the recognition and translocation of synthesized mitochondrial precursor proteins, and its protein kinase CK2-dependent phosphorylation is mandatory for TOMM complex biogenesis and proper mitochondrial protein import. In mammals, the biological function of protein kinase CSNK2/CK2 remains vastly elusive and it is unknown whether CSNK2-dependent phosphorylation of TOMM protein subunits has a similar role as that in yeast...
November 22, 2017: Autophagy
https://www.readbyqxmd.com/read/29163026/generation-of-urine-cell-derived-non-integrative-human-ipscs-and-inscs-a-step-by-step-optimized-protocol
#2
Lin Cheng, Qiannan Lei, Chen Yin, Hui-Yun Wang, Kangxin Jin, Mengqing Xiang
Objective: Establishing a practical procedure to generate induced pluripotent stem cells (iPSCs) and induced neural stem cells (iNSCs) from human urine cells (UCs). In this report, we optimized a non-integrative protocol to generate patient-specific iPSC and iNSC lines with high reprogramming efficiency. Methods: UCs were electroporated with the pEP4-EO2S-ET2K and pEP4-M2L plasmids containing the OCT4, SOX2, KLF4, SV40LT, c-MYC, and LIN28 genes, and then cultured with N2B27 medium plus four small molecule compounds (A83-01, PD0325901, Thiazovivin, and CHIR99021)...
2017: Frontiers in Molecular Neuroscience
https://www.readbyqxmd.com/read/29155724/generation-of-a-gene-disrupted-streptococcus-mutans-strain-without-gene-cloning
#3
Takatoshi Murata, Ayako Okada, Khairul Matin, Nobuhiro Hanada
Typical methods for the elucidation of the function of a particular gene involve comparative phenotypic analyses of the wild-type strain and a strain in which the gene of interest has been disrupted. A gene-disruption DNA construct containing a suitable antibiotic resistance marker gene is useful for the generation of gene-disrupted strains in bacteria. However, conventional construction methods, which require gene cloning steps, involve complex and time-consuming protocols. Here, a relatively facile, rapid, and cost-effective method for targeted gene disruption in Streptococcus mutans is described...
October 23, 2017: Journal of Visualized Experiments: JoVE
https://www.readbyqxmd.com/read/29153290/locally-advanced-pancreatic-cancer-an-emerging-entity
#4
REVIEW
Grainne M O Kane, Jennifer J Knox
Pancreatic adenocarcinoma (PDAC) remains a highly fatal disease that is increasing in incidence. PDAC can be classified according to resectability status with 3 nonmetastatic groups defined: resectable, borderline resectable, and locally advanced PDAC (LAPC). Delineating these subtypes is important with the optimal treatment approach dictated by high-quality CT imaging and multidisciplinary team discussion. Patients with LAPC are thought unresectable and are therefore rarely cured. In these patients, chemotherapy remains the mainstay of treatment...
November 16, 2017: Current Problems in Cancer
https://www.readbyqxmd.com/read/29151221/quantitative-determination-of-interacting-protein-surfaces-in-prokaryotes-and-eukaryotes-by-using-in-cell-nmr-spectroscopy
#5
David S Burz, Christopher M DeMott, Asma Aldousary, Stephen Dansereau, Alexander Shekhtman
This paper describes three protocols for identifying interacting surfaces on (15)N-labeled target proteins of known structure by using in-cell NMR spectroscopy. The first protocol describes how to identify protein quinary structure interaction surfaces in prokaryotes by using cross-relaxation-induced polarization transfer, CRIPT, based in-cell NMR. The second protocol describes how to introduce labeled protein into eukaryotic (HeLa) cells via electroporation for CRIPT-based in-cell studies. The third protocol describes how to quantitatively map protein interacting surfaces by utilizing singular value decomposition, SVD, analysis of STructural INTeractions by in-cell NMR, STINT-NMR, data...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29150011/optimization-of-crispr-cas9-genome-editing-for-loss-of-function-in-the-early-chick-embryo
#6
Shashank Gandhi, Michael L Piacentino, Felipe M Vieceli, Marianne E Bronner
The advent of CRISPR/Cas9 has made genome editing possible in virtually any organism, including those not previously amenable to genetic manipulations. Here, we present an optimization of CRISPR/Cas9 for application to early avian embryos with improved efficiency via a three-fold strategy. First, we employed Cas9 protein flanked with two nuclear localization signal sequences for improved nuclear localization. Second, we used a modified guide RNA (gRNA) scaffold that obviates premature termination of transcription and unstable Cas9-gRNA interactions...
December 1, 2017: Developmental Biology
https://www.readbyqxmd.com/read/29150004/dynamics-of-indel-profiles-induced-by-various-crispr-cas9-delivery-methods
#7
Michael Kosicki, Sandeep S Rajan, Flaminia C Lorenzetti, Hans H Wandall, Yoshiki Narimatsu, Emmanouil Metzakopian, Eric P Bennett
The introduction of CRISPR/Cas9 gene editing in mammalian cells is a scientific breakthrough, which has greatly affected basic research and gene therapy. The simplicity and general access to CRISPR/Cas9 reagents has in an unprecedented manner "democratized" gene targeting in biomedical research, enabling genetic engineering of any gene in any cell, tissue, organ, and organism. The ability for fast, precise, and efficient profiling of the double-stranded break induced insertions and deletions (indels), mediated by any of the available programmable nucleases, is paramount to any given gene targeting approach...
2017: Progress in Molecular Biology and Translational Science
https://www.readbyqxmd.com/read/29146422/numerical-study-of-the-effect-of-soft-layer-properties-on-bacterial-electroporation
#8
Jeffrey L Moran, Naga Neehar Dingari, Paulo A Garcia, Cullen R Buie
We present a numerical model of electroporation in a gram-positive bacterium, which accounts for the presence of a negatively charged soft polyelectrolyte layer (which may include a periplasmic space, peptidoglycan layer, cilia, flagella, and other surface appendages) surrounding its plasma membrane. We model the ion transport within and outside the soft layer using the soft layer electrokinetics-based Poisson-Nernst-Planck formalism. Additionally, we model the electroporation dynamics on the plasma membrane using the pore nucleation-based electroporation formalism developed by Krassowska and Filev...
September 18, 2017: Bioelectrochemistry
https://www.readbyqxmd.com/read/29143247/using-circulating-tumor-cells-to-evaluate-the-efficacy-of-irreversible-electroporation-for-unresectasble-pancreatic-cancer
#9
Mao Lin, Mohammed Alnaggar, Shu-Zhen Liang, Jian Shi, Li-Zhi Niu, Ji-Bing Chen, Ke-Cheng Xu
We used circulating tumor cells (CTCs) as biomarkers to evaluate the efficacy of pre-irreversible electroporation (IRE) and post-IRE for unresectable pancreatic cancer (PC). Real-time qPCR was used to detect potential biomarker genes in CTCs, and magnetic-activated cell sorting (MACS) and fluorescence-activated cell sorting (FACS) were performed on 43 patients with PC who underwent IRE. Some patients experienced adverse reactions within 30 days of the operation, including arrhythmia (6.9%), intraoperative transient change of blood pressure (25...
November 15, 2017: Immunologic Research
https://www.readbyqxmd.com/read/29143077/calcein-release-from-cells-in-vitro-via-reversible-and-irreversible-electroporation
#10
Violeta Rajeckaitė, Baltramiejus Jakštys, Arnas Rafanavičius, Martynas Maciulevičius, Milda Jakutavičiūtė, Saulius Šatkauskas
The aim of this study was to investigate the dependence of calcein extraction and cell viability on the parameters of pulsed electric field (PEF). Two different approaches concerning PEF parameters were investigated: (1) extraction efficiency and cell viability dependence on pulse number, exploiting 1200 V/cm 100 µs duration high voltage (HV) electric pulses and (2) extraction efficiency and cell viability dependence on the pulses with different duration (44-400 µs) and electric field strength (600-1800 V/cm) that result in the same amount of electric field energy delivered to Chinese hamster ovary cells...
November 15, 2017: Journal of Membrane Biology
https://www.readbyqxmd.com/read/29141562/a-review-of-electroporation-based-antitumor-skin-therapies-and-investigation-of-betulinic-acid-loaded-ointment
#11
Monika Bakonyi, Szilvia Berko, Gabor Eros, Gabor Varju, Cristina Dehelean, Maria Budai Szucs, Erzsebet Csanyi
BACKGROUND: Electrochemotherapy is a novel treatment for cutaneous and subcutaneous tumors utilizing the combination of electroporation and chemotherapeutic agents. Since tumors have an increasing incidence nowadays as a result of environmental and genetic factors, electrochemotherapy could be a promising treatment for cancer patients. OBJECTIVE: The aim of this article is to summarize the novel knowledge about the use of electroporation for antitumor treatments and to present a new application of electrochemotherapy with a well-known plant derived antitumor drug betulinic acid...
November 13, 2017: Anti-cancer Agents in Medicinal Chemistry
https://www.readbyqxmd.com/read/29140566/mesenchymal-stem-cell-preparation-and-transfection-free-ferumoxytol-labeling-for-mri-cell-tracking
#12
Li Liu, Chien Ho
Mesenchymal stem cells (MSCs) are multipotent cells and are the most widely studied cell type for stem cell therapies. In vivo cell tracking of MSCs labeled with an FDA-approved superparamagnetic iron-oxide (SPIO) particle by magnetic resonance imaging (MRI) provides essential information, e.g., MSC engraftment, survival, and fate, thus improving cell therapy accuracy. However, current methodology for labeling MSCs with Ferumoxytol (Feraheme(®) ), the only FDA-approved SPIO particle, needs transfection agents...
November 15, 2017: Current Protocols in Stem Cell Biology
https://www.readbyqxmd.com/read/29138988/irreversible-electroporation-of-the-uterine-cervix-in-a-rabbit-model
#13
Wei Chai, Wenlong Zhang, Zhentong Wei, Ying Xu, Jian Shi, Xiaomei Luo, Jianying Zeng, Manhua Cui, Jiannan Li, Lizhi Niu
The aim of this study was to evaluate the effects of irreversible electroporation (IRE) on the uterine cervix in a rabbit model. IRE ablation was performed in the cervices of 48 New Zealand rabbits, with one ablation lesion in each animal. Gross pathology, transmission electron microscopy, hematoxylin and eosin (H&E), Masson's trichrome (MT) stain, and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays were performed at pre-set time points (0 h, 12 h, 1 d, 2 d, 4 d, 7 d, 14 d, and 28 d post-IRE)...
November 14, 2017: Biomedical Microdevices
https://www.readbyqxmd.com/read/29138199/electrosclerotherapy-for-capillary-malformations-study-protocol-for-a-randomised-within-patient-controlled-pilot-trial
#14
Sophie E R Horbach, Albert Wolkerstorfer, Daniel Martijn de Bruin, Chantal M A M van der Horst
INTRODUCTION: The current state-of-the-art treatment modality for hypertrophic capillary malformations (CMs), laser therapy, has a considerable rate of non-responders and recurrence. Intralesional bleomycin injections (or 'sclerotherapy') are commonly used to treat venous and lymphatic malformations with an excellent effect, but these intravascular injections are not possible in CMs due to the small diameter of the vessels. Electroporation-an electric field applied to the tissue-could increase the permeability of endothelial cells, which could theoretically facilitate targeted localised bleomycin delivery...
November 14, 2017: BMJ Open
https://www.readbyqxmd.com/read/29137924/nasal-and-otic-placode-specific-regulation-of-sox2-involves-both-activation-by-sox-sall4-synergism-and-multiple-repression-mechanisms
#15
Satoko Sugahara, Tooru Fujimoto, Hisato Kondoh, Masanori Uchikawa
Transcription factor gene Sox2 is expressed throughout sensory development, but the enhancers that regulate the gene vary depending on the developmental stages and tissues. To gain new insights into the gene regulatory network in sensory placode specification, regulation of the nasal-otic bispecific NOP1 enhancer of Sox2 was investigated in chicken embryos. Deletion and mutational analyses using electroporation showed that transcriptional repression mechanisms in combination with activation mechanisms determine placodal specificity...
November 11, 2017: Developmental Biology
https://www.readbyqxmd.com/read/29135337/dna-vaccines-elicit-durable-protective-immunity-against-individual-or-simultaneous-infections-with-lassa-and-ebola-viruses-in-guinea-pigs
#16
Kathleen A Cashman, Eric R Wilkinson, Suzanne E Wollen, Joshua D Shamblin, Justine M Zelko, Jeremy J Bearss, Xiankun Zeng, Kate E Broderick, Connie S Schmaljohn
We previously developed optimized DNA vaccines against both Lassa fever and Ebola hemorrhagic fever viruses and demonstrated that they were protective individually in guinea pig and nonhuman primate models. In this study, we vaccinated groups of strain 13 guinea pigs two times, four weeks apart with 50 µg of each DNA vaccine or a mock vaccine at discrete sites by intradermal electroporation. Five weeks following the second vaccinations, guinea pigs were exposed to lethal doses of Lassa virus, Ebola virus, or a combination of both viruses simultaneously...
November 14, 2017: Human Vaccines & Immunotherapeutics
https://www.readbyqxmd.com/read/29135047/effects-of-voltage-strength-during-electroporation-on-the-development-and-quality-of-in-vitro-produced-porcine-embryos
#17
K Nishio, F Tanihara, T-V Nguyen, T Kunihara, M Nii, M Hirata, T Takemoto, T Otoi
This study was conducted to determine suitable conditions for an experimental method in which the CRISPR/Cas9 system is introduced into in vitro-produced porcine zygotes by electroporation. In the first experiment, when putative zygotes derived from in vitro fertilization (IVF) were electroporated by either unipolar or bipolar pulses, keeping the voltage, pulse duration and pulse number fixed at 30 V/mm, 1 msec and five repeats, respectively, the rate of blastocyst formation from zygotes electroporated by bipolar pulses decreased compared to zygotes electroporated by unipolar pulses...
November 14, 2017: Reproduction in Domestic Animals, Zuchthygiene
https://www.readbyqxmd.com/read/29134602/applications-of-the-bacteriophage-mu-in-vitro-transposition-reaction-and-genome-manipulation-via-electroporation-of-dna-transposition-complexes
#18
Saija Haapa-Paananen, Harri Savilahti
The capacity of transposable elements to insert into the genomes has been harnessed during the past decades to various in vitro and in vivo applications. This chapter describes in detail the general protocols and principles applicable for the Mu in vitro transposition reaction as well as the assembly of DNA transposition complexes that can be electroporated into bacterial cells to accomplish efficient gene delivery. These techniques with their modifications potentiate various gene and genome modification applications, which are discussed briefly here, and the reader is referred to the original publications for further details...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29132995/comparative-functional-potency-of-dna-vaccines-encoding-plasmodium-falciparum-transmission-blocking-target-antigens-pfs48-45-and-pfs25-administered-alone-or-in-combination-by-in-vivo-electroporation-in-rhesus-macaques
#19
Dibyadyuti Datta, Geetha P Bansal, Brooke Grasperge, Dale S Martin, Mario Philipp, Dietlind Gerloff, Barry Ellefsen, Drew Hannaman, Nirbhay Kumar
Antibodies recognizing conformational epitopes in Pfs48/45, an antigen expressed on the surface of Plasmodium falciparum gametes and zygotes, have firmly established Pfs48/45 as a promising transmission blocking vaccine (TBV) candidate. However, it has been difficult to reproducibly express Pfs48/45 in a variety of recombinant expression systems. The goal of our studies was to evaluate functional immunogenicity of Pfs48/45 using DNA vaccine format in rhesus macaques. An additional goal was to ensure that when used in combination with another malarial antigen, specific immunity to both antigens was not compromised...
November 10, 2017: Vaccine
https://www.readbyqxmd.com/read/29132503/gyrification-of-the-cerebral-cortex-requires-fgf-signaling-in-the-mammalian-brain
#20
Naoyuki Matsumoto, Yohei Shinmyo, Yoshie Ichikawa, Hiroshi Kawasaki
Although it has been believed that the evolution of cortical folds was a milestone, allowing for an increase in the number of neurons in the cerebral cortex, the mechanisms underlying the formation of cortical folds are largely unknown. Here we show regional differences in the expression of fibroblast growth factor receptors (FGFRs) in the developing cerebral cortex of ferrets even before cortical folds are formed. By taking the advantage of our in utero electroporation technique for ferrets, we found that cortical folding was impaired in the ferret cerebral cortex when FGF signaling was inhibited...
November 14, 2017: ELife
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