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Electroporation

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https://www.readbyqxmd.com/read/29352001/keap1-editing-using-crispr-cas9-for-therapeutic-nrf2-activation-in-primary-human-t-lymphocytes
#1
Sanjeev Noel, Sul A Lee, Mohanraj Sadasivam, Abdel R A Hamad, Hamid Rabb
Oxidant stress modifies T lymphocyte activation and function. Previous work demonstrated that murine T cell-specific kelch like-ECH-associated protein 1 (Keap1) deletion enhances antioxidant capacity and protects from experimental acute kidney injury. In this study, we used CRISPR technology to develop clinically translatable human T cell-specific KEAP1 deletion. Delivery of KEAP1 exon 2 specific Cas9:guide RNA in Jurkat T cells led to significant (∼70%) editing and upregulation of NRF2-regulated antioxidant genes NADPH dehydrogenase quinone 1 (NQO1) (up to 11-fold), heme oxygenase 1 (HO1) (up to 11-fold), and GCLM (up to 2-fold)...
January 19, 2018: Journal of Immunology: Official Journal of the American Association of Immunologists
https://www.readbyqxmd.com/read/29349911/identification-of-fish-source-vibrio-alginolyticus-and-evaluation-of-its-bacterial-ghosts-vaccine-immune-effects
#2
Ji Cao, Jiajun Zhang, Lin Ma, Lin Li, Wenchang Zhang, Jinnian Li
Vibrio alginolyticus (V. alginolyticus) is a common pathogen for humans and marine aquatic animals. Vibriosis of marine aquatic animals, caused by V. alginolyticus, has become more prevalent globally in recent years. Hence, a safe and effective vaccine is urgently needed for the control of this disease. Here, the strain 16-3 isolated from the large yellow croaker (Larimichthys crocea) suffered from canker was identified as V. alginolyticus based on morphological, biochemical, and 16S rDNA sequencing analysis...
January 19, 2018: MicrobiologyOpen
https://www.readbyqxmd.com/read/29346108/modeling-of-electroporation-induced-by-pulsed-electric-fields-in-irregularly-shaped-cells
#3
Luciano Mescia, Michele A Chiapperino, Pietro Bia, Johan Gielis, Diego Caratelli
During the past decades, the poration of cell membrane induced by pulsed electric fields has been widely investigated. Since the basic mechanisms of this process have not yet been fully clarified, many research activities are focused on the development of suitable theoretical and numerical models. To this end, a nonlinear, nonlocal, dispersive, and space-time numerical algorithm has been developed and adopted to evaluate the transmembrane voltage and pore density along the perimeter of realistic irregularly shaped cells...
February 2018: IEEE Transactions on Bio-medical Engineering
https://www.readbyqxmd.com/read/29342244/gli2-rescues-delays-in-brain-development-induced-by-kif3a-dysfunction
#4
Jia-Long Chen, Chia-Hsiang Chang, Jin-Wu Tsai
The primary cilium in neural stem cells plays distinct roles in different stages during cortical development. Ciliary dysfunctions in human (i.e., ciliopathy) cause developmental defects in multiple organs, including brain developmental delays, which lead to intellectual disabilities and cognitive deficits. However, effective treatment to this devastating developmental disorder is still lacking. Here, we first investigated the effects of ciliopathy on neural stem cells by knocking down Kif3a, a kinesin II motor required for ciliogenesis, in the neurogenic stage of cortical development by in utero electroporation of mouse embryos...
January 12, 2018: Cerebral Cortex
https://www.readbyqxmd.com/read/29337375/transfection-by-electroporation
#5
Huntington Potter, Richard Heller
Electroporation-the use of high-voltage electric shocks to introduce DNA into cells-can be used with most cell types, yields a high frequency of both stable transformation and transient gene expression, and, because it requires fewer steps, can be easier than alternate techniques. This unit describes electroporation of mammalian cells, including ES cells for the preparation of knock-out, knock-in, and transgenic mice. Protocols are described for the use of electroporation in vivo to perform gene therapy for cancer therapy and DNA vaccination...
January 16, 2018: Current Protocols in Molecular Biology
https://www.readbyqxmd.com/read/29337302/critical-roles-of-%C3%AE-ii-spectrin-in-brain-development-and-epileptic-encephalopathy
#6
Yu Wang, Tuo Ji, Andrew D Nelson, Katarzyna Glanowska, Geoffrey G Murphy, Paul M Jenkins, Jack M Parent
The nonerythrocytic α-spectrin-1 (SPTAN1) gene encodes the cytoskeletal protein αII spectrin. Mutations in SPTAN1 cause early infantile epileptic encephalopathy type 5 (EIEE5); however, the role of αII spectrin in neurodevelopment and EIEE5 pathogenesis is unknown. Prior work suggests that αII spectrin is absent in the axon initial segment (AIS) and contributes to a diffusion barrier in the distal axon. Here, we have shown that αII spectrin is expressed ubiquitously in rodent and human somatodendritic and axonal domains...
January 16, 2018: Journal of Clinical Investigation
https://www.readbyqxmd.com/read/29335228/irreversible-electroporation-for-locally-advanced-pancreatic-cancer-through-a-minimally-invasive-surgery-supported-by-laparoscopic-ultrasound
#7
Ernesto Tartaglia, Massimiliano Fabozzi, Antonia Rizzuto, Anna Settembre, Roberta Abete, Ludovica Guerriero, Pasqualino Favoriti, Diego Cuccurullo, Francesco Corcione
INTRODUCTION: Pancreatic cancer is one of the most lethal cancers worldwide, with 5-years survival rate as low as 6%. The majority of pancreatic cancer patients present locally advanced or metastatic disease at diagnosis. Typically, patients affected by locally advanced pancreatic cancer (LAPC) do not undergo radical surgery but are treated with focal ablative therapies. However, a high rate of morbidity due to the heat sink effect has limited the application of ablative techniques on a routine basis in LAPC patients...
December 28, 2017: International Journal of Surgery Case Reports
https://www.readbyqxmd.com/read/29334773/macf1-overexpression-by-transfecting-the-21-kbp-large-plasmid-pegfp-c1a-acf7-promotes-osteoblast-differentiation-and-bone-formation
#8
Yan Zhang, Chong Yin, Lifang Hu, Zhihao Chen, Fan Zhao, Dijie Li, Jianhua Ma, Xiaoli Ma, Peihong Su, Wuxia Qiu, Chaofei Yang, Pai Wang, Siyu Li, Ge Zhang, Liping Wang, Airong Qian, Cory Xian
Microtubule actin crosslinking factor 1 (MACF1) is a large spectraplakin protein known to have crucial roles in regulating cytoskeletal dynamics, cell migration, growth and differentiation. However, its role and action mechanism in bone remain unclear. This study investigated the optimal conditions for effective transfection of the large plasmid PEGFP-C1A-ACF7 (~21 kbp) containing the full-length human MACF1 cDNA and the potential role of MACF1 in bone formation. To enhance MACF1 expression, the large plasmid was transfected into osteogenic cells by electroporation in vitro and mouse calvaria with nanoparticles...
January 15, 2018: Human Gene Therapy
https://www.readbyqxmd.com/read/29333120/focused-transhepatic-electroporation-mediated-by-hypersaline-infusion-through-the-portal-vein-in-rat-model-preliminary-results-on-differential-conductivity
#9
Clara Pañella, Quim Castellví, Xavier Moll, Rita Quesada, Alberto Villanueva, Mar Iglesias, Dolores Naranjo, Patricia Sánchez-Velázquez, Anna Andaluz, Luís Grande, Antoni Ivorra, Fernando Burdío
Background: Spread hepatic tumours are not suitable for treatment either by surgery or conventional ablation methods. The aim of this study was to evaluate feasibility and safety of selectively increasing the healthy hepatic conductivity by the hypersaline infusion (HI) through the portal vein. We hypothesize this will allow simultaneous safe treatment of all nodules by irreversible electroporation (IRE) when applied in a transhepatic fashion. Material and methods: Sprague Dawley (Group A, n = 10) and Athymic rats with implanted hepatic tumour (Group B, n = 8) were employed...
December 2017: Radiology and Oncology
https://www.readbyqxmd.com/read/29330458/architecture-of-a-mammalian-glomerular-domain-revealed-by-novel-volume-electroporation-using-nanoengineered-microelectrodes
#10
D Schwarz, M Kollo, C Bosch, C Feinauer, I Whiteley, T W Margrie, T Cutforth, A T Schaefer
Dense microcircuit reconstruction techniques have begun to provide ultrafine insight into the architecture of small-scale networks. However, identifying the totality of cells belonging to such neuronal modules, the "inputs" and "outputs," remains a major challenge. Here, we present the development of nanoengineered electroporation microelectrodes (NEMs) for comprehensive manipulation of a substantial volume of neuronal tissue. Combining finite element modeling and focused ion beam milling, NEMs permit substantially higher stimulation intensities compared to conventional glass capillaries, allowing for larger volumes configurable to the geometry of the target circuit...
January 12, 2018: Nature Communications
https://www.readbyqxmd.com/read/29323173/electroporation-of-mice-zygotes-with-dual-guide-rna-cas9-complexes-for-simple-and-efficient-cloning-free-genome-editing
#11
Marie Teixeira, Bénédicte F Py, Christophe Bosc, Daphné Laubreton, Marie-Jo Moutin, Jacqueline Marvel, Frédéric Flamant, Suzy Markossian
In this report, we present an improved protocol for CRISPR/Cas9 genome editing in mice. The procedure consists in the electroporation of intact mouse zygotes with ribonucleoprotein complexes prepared in vitro from recombinant Cas9 nuclease and synthetic dual guide RNA. This simple cloning-free method proves to be extremely efficient for the generation of indels and small deletions by non-homologous end joining, and for the generation of specific point mutations by homology-directed repair. The procedure, which avoids DNA construction, in vitro transcription and oocyte microinjection, greatly simplifies genome editing in mice...
January 11, 2018: Scientific Reports
https://www.readbyqxmd.com/read/29322238/development-of-inhibitory-synaptic-inputs-on-layer-2-3-pyramidal-neurons-in-the-rat-medial-prefrontal-cortex
#12
Mari A Virtanen, Claudia Marvine Lacoh, Hubert Fiumelli, Markus Kosel, Shiva Tyagarajan, Mathias de Roo, Laszlo Vutskits
Inhibitory control of pyramidal neurons plays a major role in governing the excitability in the brain. While spatial mapping of inhibitory inputs onto pyramidal neurons would provide important structural data on neuronal signaling, studying their distribution at the single cell level is difficult due to the lack of easily identifiable anatomical proxies. Here, we describe an approach where in utero electroporation of a plasmid encoding for fluorescently tagged gephyrin into the precursors of pyramidal cells along with ionotophoretic injection of Lucifer Yellow can reliably and specifically detect GABAergic synapses on the dendritic arbour of single pyramidal neurons...
January 10, 2018: Brain Structure & Function
https://www.readbyqxmd.com/read/29318727/transformation-of-diplonema-papillatum-the-type-species-of-the-highly-diverse-and-abundant-marine-micro-eukaryotes-diplonemida-euglenozoa
#13
Binnypreet Kaur, Matus Valach, Priscila Peña-Diaz, Sandrine Moreira, Patrick J Keeling, Gertraud Burger, Julius Lukeš, Drahomíra Faktorová
Diplonema papillatum is the type species of diplonemids, which are among the most abundant and diverse heterotrophic micro-eukaryotes in the world's oceans. Diplonemids are also known for a unique form of post-transcriptional processing in mitochondria. However, the lack of reverse genetics methodologies in these protists has hampered elucidation of their cell and molecular biology. Here we report a protocol for D. papillatum transformation. We have identified several antibiotics to which D. papillatum is sensitive and thus are suitable selectable markers, and focus in particular on puromycin...
January 10, 2018: Environmental Microbiology
https://www.readbyqxmd.com/read/29317485/meis1-coordinates-cerebellar-granule-cell-development-by-regulating-pax6-transcription-bmp-signaling-and-atoh1-degradation
#14
Tomoo Owa, Shinichiro Taya, Satoshi Miyashita, Mariko Yamashita, Toma Adachi, Koyo Yamada, Miwa Yokoyama, Shogo Aida, Tomoki Nishioka, Yukiko U Inoue, Ryo Goitsuka, Takuro Nakamura, Takayoshi Inoue, Kozo Kaibuchi, Mikio Hoshino
Cerebellar granule cell precursors (GCPs) and granule cells (GCs) represent good models to study neuronal development. Here we report that the transcription factor, Meis1, plays pivotal roles to regulate mouse GC development. We found that Meis1 is expressed in granule cell lineage cells and astrocytes in the cerebellum during development. Targeted disruption of the Meis1 gene specifically in the GC lineage resulted in smaller cerebella with disorganized lobules. Knockdown/knockout experiments for Meis1 as well as in vitro assays show that Meis1 binds to an upstream sequence of Pax6 to enhance its transcription in GCPs/GCs and further suggested that the Meis1-Pax6 cascade regulates morphology of GCPs/GCs during development...
January 9, 2018: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
https://www.readbyqxmd.com/read/29307242/technology-of-irreversible-electroporation-and-review-of-its-clinical-data-on-liver-cancers
#15
Emil I Cohen, David Field, George Emmett Lynskey, Alexander Y Kim
Irreversible electroporation (IRE) has developed as a novel percutaneous ablative technique over the past decade and its utility in the treatment of primary and metastatic liver disease has progressed rapidly. Areas Covered: After discussing the principles behind the technology and the practical steps in its use, this article offers a detailed analysis of the recent published work that evaluates its safety and efficacy. The strengths and weaknesses of other ablative techniques, including radiofrequency ablation, microwave ablation and cryoablation, are discussed in detail...
January 8, 2018: Expert Review of Medical Devices
https://www.readbyqxmd.com/read/29305201/initial-assessment-of-the-efficacy-of-irreversible-electroporation-ire-in-the-focal-treatment-of-localised-renal-cell-carcinoma-rcc-with-delayed-interval-kidney-tumour-resection-irene-trial-an-ablate-and-resect-pilot-study
#16
Johann J Wendler, Maciej Pech, Frank Fischbach, Julian Jürgens, Björn Friebe, Daniel Baumunk, Markus Porsch, Simon Blaschke, Daniel Schindele, Sandra Siedentopf, Jens Ricke, Martin Schostak, Jens Köllermann, Uwe B Liehr
OBJECTIVES: To assess the efficacy of irreversible electroporation (IRE) ablation of pT1a renal-cell carcinoma (RCC) in the first prospective, monocentric Phase 2a pilot ablate-and-resect study (IRENE trial). It has been postulated that focal IRE can bring about complete ablation of soft-tissue tumours with protection of healthy peritumoral tissue and anatomical structures. METHODS: The first seven study patients, with biopsy-proven pT1a RCC (15-39mm) underwent IRE...
January 2, 2018: Urology
https://www.readbyqxmd.com/read/29305086/the-candidate-schizophrenia-risk-gene-dgcr2-regulates-early-steps-of-corticogenesis
#17
Aude Molinard-Chenu, Alexandre Dayer
BACKGROUND: Alterations in early steps of cortical circuit assembly are thought to play a critical role in vulnerability to schizophrenia (SZ), but the pathogenic impact of SZ-risk mutations on corticogenesis remains to be determined. DiGeorge syndrome critical region 2 (DGCR2) is located in the 22q11.2 locus, whose deletion is a major risk factor for SZ. Moreover, exome sequencing of individuals with idiopathic SZ identified a rare missense mutation in DGCR2, further suggesting that DGCR2 is involved in SZ...
November 21, 2017: Biological Psychiatry
https://www.readbyqxmd.com/read/29303914/current-opinion-in-urology-2017-focal-therapy-of-small-renal-lesions
#18
Brett A Johnson, Jeffrey A Cadeddu
PURPOSE OF REVIEW: With the increasing incidence of small renal masses (SRMs), ablative technologies are becoming more commonly utilized. With any nascent treatment modality, outcomes literature needs to be constantly re-evaluated. The purpose of this review is to revisit the most updated literature regarding the safety and efficacy of ablative treatments of renal lesions. RECENT FINDINGS: Recent literature demonstrates that small renal tumor ablation is safe and effective...
January 4, 2018: Current Opinion in Urology
https://www.readbyqxmd.com/read/29298437/absence-of-tangentially-migrating-glutamatergic-neurons-in-the-developing-avian-brain
#19
Fernando García-Moreno, Edward Anderton, Marta Jankowska, Jo Begbie, Juan Manuel Encinas, Manuel Irimia, Zoltán Molnár
Several neuronal populations orchestrate neocortical development during mammalian embryogenesis. These include the glutamatergic subplate-, Cajal-Retzius-, and ventral pallium-derived populations, which coordinate cortical wiring, migration, and proliferation, respectively. These transient populations are primarily derived from other non-cortical pallial sources that migrate to the dorsal pallium. Are these migrations to the dorsal pallium conserved in amniotes or are they specific to mammals? Using in ovo electroporation, we traced the entire lineage of defined chick telencephalic progenitors...
January 2, 2018: Cell Reports
https://www.readbyqxmd.com/read/29297692/comparison-of-the-expression-kinetics-and-immunostimulatory-activity-of-replicating-mrna-non-replicating-mrna-and-pdna-after-intradermal-electroporation-in-pigs
#20
Bregje Leyman, Hanne Huysmans, Séan Mc Cafferty, Francis Combes, Eric Cox, Bert Devriendt, Niek N Sanders
Synthetic mRNA is becoming increasingly popular as an alternative to pDNA-based gene therapy. Currently, multiple synthetic mRNA platforms have been developed. In this study we investigated the expression kinetics and the changes in mRNA encoding cytokine and chemokine levels following intradermal electroporation in pigs of pDNA, self-replicating mRNA, modified- and unmodified mRNA. The self-replicating mRNA tended to induce the highest protein expression, followed by pDNA, modified mRNA and unmodified mRNA...
January 3, 2018: Molecular Pharmaceutics
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