scholarly journals Japanese Macaque Rhadinovirus Encodes a Viral MicroRNA Mimic of the miR-17 Family

2016 ◽  
Vol 90 (20) ◽  
pp. 9350-9363 ◽  
Author(s):  
Rebecca L. Skalsky ◽  
Sarah A. Barr ◽  
Andrew J. Jeffery ◽  
Tiffany Blair ◽  
Ryan Estep ◽  
...  
Keyword(s):  
Multiple Sclerosis ◽  
Demyelinating Disease ◽  
Japanese Macaque ◽  
Large Animal ◽  
Primate Model ◽  
Sequence Comparisons ◽  
Necrosis Factor Alpha ◽  
Viral Mirnas ◽  
Large Animal Models

ABSTRACTJapanese macaque (JM) rhadinovirus (JMRV) is a novel, gamma-2 herpesvirus that was recently isolated from JM with inflammatory demyelinating encephalomyelitis (JME). JME is a spontaneous and chronic disease with clinical characteristics and immunohistopathology comparable to those of multiple sclerosis in humans. Little is known about the molecular biology of JMRV. Here, we sought to identify and characterize the small RNAs expressed during lytic JMRV infection using deep sequencing. Fifteen novel viral microRNAs (miRNAs) were identified in JMRV-infected fibroblasts, all of which were readily detectable by 24 h postinfection and accumulated to high levels by 72 h. Sequence comparisons to human Kaposi's sarcoma-associated herpesvirus (KSHV) miRNAs revealed several viral miRNA homologs. To functionally characterize JMRV miRNAs, we screened for their effects on nuclear factor kappa B (NF-κB) signaling in the presence of two proinflammatory cytokines, tumor necrosis factor alpha (TNF-α) and interleukin-1β (IL-1β). Multiple JMRV miRNAs suppressed cytokine-induced NF-κB activation. One of these miRNAs, miR-J8, has seed sequence homology to members of the cellular miR-17/20/106 and miR-373 families, which are key players in cell cycle regulation as well as inflammation. Using reporters, we show that miR-J8 can target 3′ untranslated regions (UTRs) with miR-17-5p or miR-20a cognate sites. Our studies implicate JMRV miRNAs in the suppression of innate antiviral immune responses, which is an emerging feature of many viral miRNAs.IMPORTANCEGammaherpesviruses are associated with multiple diseases linked to immunosuppression and inflammation, including AIDS-related cancers and autoimmune diseases. JMRV is a recently identified herpesvirus that has been linked to JME, an inflammatory demyelinating disease in Japanese macaques that mimics multiple sclerosis. There are few large-animal models for gammaherpesvirus-associated pathogenesis. Here, we provide the first experimental evidence of JMRV miRNAsin vitroand demonstrate that one of these viral miRNAs can mimic the activity of the cellular miR-17/20/106 family. Our work provides unique insight into the roles of viral miRNAs during rhadinovirus infection and provides an important step toward understanding viral miRNA function in a nonhuman primate model system.

scholarly journals Review: Tissue Engineering of Small-Diameter Vascular Grafts and Their In Vivo Evaluation in Large Animals and Humans

Cells ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 713
Author(s):  
Shu Fang ◽  
Ditte Gry Ellman ◽  
Ditte Caroline Andersen
Keyword(s):  
Animal Models ◽  
Vascular Grafts ◽  
Small Diameter ◽  
Large Animal ◽  
Large Animal Models ◽  
Graft Outcome ◽  
Limited Success ◽  
Large Animals

To date, a wide range of materials, from synthetic to natural or a mixture of these, has been explored, modified, and examined as small-diameter tissue-engineered vascular grafts (SD-TEVGs) for tissue regeneration either in vitro or in vivo. However, very limited success has been achieved due to mechanical failure, thrombogenicity or intimal hyperplasia, and improvements of the SD-TEVG design are thus required. Here, in vivo studies investigating novel and relative long (10 times of the inner diameter) SD-TEVGs in large animal models and humans are identified and discussed, with emphasis on graft outcome based on model- and graft-related conditions. Only a few types of synthetic polymer-based SD-TEVGs have been evaluated in large-animal models and reflect limited success. However, some polymers, such as polycaprolactone (PCL), show favorable biocompatibility and potential to be further modified and improved in the form of hybrid grafts. Natural polymer- and cell-secreted extracellular matrix (ECM)-based SD-TEVGs tested in large animals still fail due to a weak strength or thrombogenicity. Similarly, native ECM-based SD-TEVGs and in-vitro-developed hybrid SD-TEVGs that contain xenogeneic molecules or matrix seem related to a harmful graft outcome. In contrast, allogeneic native ECM-based SD-TEVGs, in-vitro-developed hybrid SD-TEVGs with allogeneic banked human cells or isolated autologous stem cells, and in-body tissue architecture (IBTA)-based SD-TEVGs seem to be promising for the future, since they are suitable in dimension, mechanical strength, biocompatibility, and availability.

scholarly journals Mode of bacterial killing affects the inflammatory response and associated organ dysfunctions in a porcine E. coli intensive care sepsis model

Critical Care ◽  
2020 ◽  
Vol 24 (1) ◽  
Author(s):  
Paul Skorup ◽  
Lisa Maudsdotter ◽  
Miklós Lipcsey ◽  
Anders Larsson ◽  
Jan Sjölin
Keyword(s):  
Inflammatory Response ◽  
Structural Changes ◽  
Interleukin 10 ◽  
Induced Response ◽  
Large Animal ◽  
Necrosis Factor Alpha ◽  
Bacterial Killing ◽  
Treatment Groups ◽  
Sepsis Model

Abstract Background Sepsis is often treated with penicillin-binding protein 3 (PBP-3) acting β-lactam antibiotics, such as piperacillin-tazobactam, cefotaxime, and meropenem. They cause considerable bacterial structural changes and have in vitro been associated with an increased inflammatory response. In a clinically relevant large animal sepsis model, our primary aim was to investigate whether bacteria killed by a PBP-3-active antibiotic has a greater effect on the early inflammatory response and organ dysfunction compared with corresponding amounts of live or heat-killed bacteria. A secondary aim was to determine whether the addition of an aminoglycoside could mitigate the cefuroxime-induced response. Method Killed or live Escherichia coli were administrated as a 3-h infusion to 16 healthy pigs in a prospective, randomized controlled interventional experimental study. Cefuroxime was chosen as the PBP-3-active antibiotic and tobramycin represented the aminoglycosides. The animals were randomized to receive (I) bacteria killed by cefuroxime, (II) live bacteria, (III) bacteria killed by heat, or (IV) bacteria killed by the combination of cefuroxime and tobramycin. Plasma endotoxin, tumor necrosis factor alpha, interleukin-6, interleukin-10, leukocytes, and organ function were recorded at the start of the experiment and then hourly for 6 h. Results Differences in dynamics of concentration over time between the four treatment groups were found for the three cytokines (p < 0.001). Animals receiving cefuroxime-killed bacteria demonstrated higher responses than those receiving live (p < 0.05) or heat-killed bacteria (p < 0.01). The addition of tobramycin reduced the cefuroxime-induced responses (p < 0.001). The cytokine responses were associated with leucocyte activation that was further associated with pulmonary dysfunction and increases in lactate (p < 0.01). Conclusions In comparison with live or heat-killed bacteria, bacteria killed by a PBP-3-active antibiotic induced an increased inflammatory response that appears to be associated with deteriorated organ and cellular function. The addition of an aminoglycoside to the PBP-3-active antibiotic reduced that response.

scholarly journals A Novel Approach to Drug Delivery for Hepatities C Virus (HCV) for High Immune Responses

2008 ◽  
Vol 2 (2) ◽  
Author(s):  
W. T. Chen ◽  
C. Zhang
Keyword(s):  
Immune Responses ◽  
High Strength ◽  
Azomethine Ylides ◽  
Enzyme Linked Immunosorbent Assay ◽  
Large Animal ◽  
Testing Environment ◽  
Large Animal Models ◽  
Novel Approach ◽  
Large Animals

Hepatities C Virus (HCV) is a significant health problem worldwide due to the lack of effective vaccines. HCV plasmid DNA (pDNA) vaccine represents a promising means to induce a Th1-biased cell-mediated response which tends to be associated with HCV clearance. However, the immune responses induced by naked pDNA vaccine in large animals as well as in humans are usually too weak to show sufficient protection against new infections. Therefore, it is interesting to look for new ways to deliver HCV pDNA vaccine. In this research, carbon nanotube (CNT) is used as a carrier to deliver the pDNA vaccine of HCV to induce high immune responses, because CNT has some excellent properties such as high strength and good biocompatibility. One of the key approaches to make this idea work is to treat CNT so that it can bind with HCV pDNA with good stability. An approach called 1, 3-dipolar cycloaddition of azomethine ylides was modified. We analyzed the complex of f-CNTs combined with pDNA vaccines expressing HCV E2 protein by using Enzyme-linked immunospot (ELISPOT) or Enzyme-linked immunosorbent assay (ELISA) assay in vitro. The result showed that the CNT approach can induce stronger protective immune responses than the needle delivery of naked pDNA vaccine. We have also found an optimal way to treat CNT in light of the highest immune response in the same testing environment. The success of this research will warrant testing HCV vaccine in large animal models and human clinical trials.

scholarly journals Adult Canine Intestinal Derived Organoids: A Novel In Vitro System for Translational Research in Comparative Gastroenterology

2018 ◽  
Author(s):  
Lawrance Chandra ◽  
Dana C Borcherding ◽  
Dawn Kingsbury ◽  
Todd Atherly ◽  
Yoko M Ambrosini ◽  
...  
Keyword(s):  
Animal Models ◽  
Translational Research ◽  
Three Dimensional ◽  
Metabolic Imaging ◽  
Large Animal ◽  
Large Animal Models ◽  
Molecular Features ◽  
Naturally Occurring

AbstractBackgroundLarge animal models, such as the dog, are increasingly being used over rodent models for studying naturally occurring diseases including gastrointestinal (GI) disorders. Dogs share similar environmental, genomic, anatomical, and intestinal physiologic features with humans. To bridge the gap between currently used animal models (e.g. mouse) and humans, and expand the translational potential of the dog model, we developed a three dimensional (3D) canine GI organoid (enteroid and colonoid) system. Organoids have recently gained interest in translational research as this model system better recapitulates the physiological and molecular features of the tissue environment in comparison with two-dimensional cultures.ResultsOrganoids were propagated from isolation of adult intestinal stem cells (ISC) from whole jejunal tissue as well as endoscopically obtained duodenal, ileal and colonic biopsy samples of healthy dogs and GI cases, including inflammatory bowel disease (IBD) and intestinal carcinomas. Intestinal organoids were comprehensively characterized using histology, immunohistochemistry, RNA in situ hybridization and transmission electron microscopy, and organoids mimicked the in vivo tissue environment. Physiological relevance of the enteroid system was defined using functional assays such as Optical Metabolic Imaging (OMI), the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) function assay, and Exosome-Like Vesicles (EV) uptake assay, as a basis for wider applications of this technology in basic, preclinical and translational GI research.ConclusionsIn summary, our findings establish the canine GI organoid systems as a novel model to study naturally occurring intestinal diseases in dogs and humans. Furthermore, canine organoid systems will help to elucidate host-pathogen interactions contributing to GI disease pathogenesis.

276 CHARACTERIZATION OF BOVINE EPIBLAST OUTGROWTH COLONIES DERIVED FROM DAY 12 BLASTOCYSTS

2008 ◽  
Vol 20 (1) ◽  
pp. 218
Author(s):  
E. Østrup ◽  
K. Schauser ◽  
J. O. Gjørret ◽  
P. Maddox-Hyttel
Keyword(s):  
Es Cells ◽  
Cell Sheet ◽  
Calf Serum ◽  
Embryonic Stem ◽  
Large Animal ◽  
Santa Cruz ◽  
Cuboidal Cell ◽  
The Core ◽  
Large Animal Models

Isolation and culture of mouse embryonic stem (ES) cells has been performed for many years, and the improvements achieved throughout the last decade in the human field has evoked great hopes for future cell replacement therapies. However, despite certain similarities in the molecular regulation of pluripotency between man and mouse, there is a need for developing large animal models. The aim of our study was to isolate, culture, and characterize bovine ES-like cell colonies derived from the epiblast. Embryos were produced by in vitro maturation, fertilization, and culture. After 6 days of in vitro culture, blastocysts were transferred to synchronized heifers and allowed to develop for an additional 6 days in vivo. At Day 12 after insemination, embryos were collected by nonsurgical flushing. Embryonic discs were isolated from 15 blastocysts by microsurgery and cultured on mitomycin-inactivated mouse embryonic fibroblasts (SLN cells) in DMEM/F12 medium supplemented with 15% fetal calf serum (FCS), 5% knockout serum replacement (KSR), 106UmL–1 leukemia inhibitory factor (LIF), basic fibroblast growth factor (bFGF), nonessential amino acids (NEAA), and nucleosides. After 4 (n = 6), 6 (n = 4), and 8 days (n = 5) of culture, the primary outgrowth colonies were fixed in 4% paraformaldehyde, embedded in paraffin, sectioned, and exposed to antisera recognizing Oct-4 (pluripotency marker; Santa Cruz Biotechnology, Santa Cruz, CA, USA), Vimentin (mesenchyme marker; Zymed Laboratories, South San Francisco, CA, USA), Cytokeratin-8 (trophectoderm marker; Becton, Dickinson and Co., Franklin Lakes, NJ, USA), and α-1-Fetoprotein (hypoblast marker; DakoCytomation, Glostrup, Denmark). The site of antigen-antibody reaction was revealed using the ABC-AEC-method and counterstained with hematoxylin. At Day 4, all colonies had developed a compact central core of cells with a low cytoplasm-to-nucleus ratio, surrounded by a monolayer of squamous cells. At Days 6 and 8, 3 out of 4 and 3 out of 5 colonies, respectively, still presented the compact core which occasionally was encapsulated by a squamous or cuboidal cell sheet. In the remaining colonies, a compact core was less defined. Oct-3/4 staining was observed in the nuclei of the compact core in 5 out of 6 colonies on Day 4, and in all colonies presenting a compact core on Days 6 and 8. However, whereas all nuclei in the core were stained on Days 4 and 6, only scattered nuclei were stained on Day 8. Vimentin staining was observed in the cytoplasm of cells in the compact core in 3 out of 6 Day 4 colonies, in all Day 6 colonies presenting a compact core, but not in any Day 8 colonies. In contrast, α-1-Fetoprotein staining intensity increased with culture period and was mostly observed in squamous monolayer portions. Cytokeratin-8 staining was weak and restricted to the cytoplasm of the cells encapsulating and surrounding the core in 2 Day 6 colonies and a single Day 8 colony. In conclusion, epiblasts isolated from Day 12 bovine blastocysts efficiently attach to feeder cells and develop outgrowth colonies with cores containing presumptive pluripotent cells (Oct-4). However, these cells to some degree lost Oct-4 expression toward Day 8 and were, in parallel, to some degree overgrown by cells of hypoblast (α-1-Fetoprotein) and trophectoderm (Cytokeratin-8) origin.

Skeletal tissue engineering—from in vitro studies to large animal models

Biomaterials ◽  
2004 ◽  
Vol 25 (9) ◽  
pp. 1487-1495 ◽  
Author(s):  
Pieter Buma ◽  
Willem Schreurs ◽  
Nico Verdonschot
Keyword(s):  
Tissue Engineering ◽  
Animal Models ◽  
In Vitro Studies ◽  
Large Animal ◽  
Skeletal Tissue ◽  
Large Animal Models

scholarly journals The adult oligodendrocyte can participate in remyelination

2018 ◽  
Vol 115 (50) ◽  
pp. E11807-E11816 ◽  
Author(s):  
Ian D. Duncan ◽  
Abigail B. Radcliff ◽  
Moones Heidari ◽  
Grahame Kidd ◽  
Benjamin K. August ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Multiple Sclerosis ◽  
Nonhuman Primates ◽  
Light And Electron Microscopy ◽  
Vitamin B ◽  
Oligodendrocyte Progenitor Cells ◽  
Large Animal ◽  
Myelin Sheaths ◽  
Large Animal Models ◽  
2D And 3D

Endogenous remyelination of the CNS can be robust and restore function, yet in multiple sclerosis it becomes less complete with time. Promoting remyelination is a major therapeutic goal, both to restore function and to protect axons from degeneration. Remyelination is thought to depend on oligodendrocyte progenitor cells, giving rise to nascent remyelinating oligodendrocytes. Surviving, mature oligodendrocytes are largely regarded as being uninvolved. We have examined this question using two large animal models. In the first model, there is extensive demyelination and remyelination of the CNS, yet oligodendrocytes survive, and in recovered animals there is a mix of remyelinated axons interspersed between mature, thick myelin sheaths. Using 2D and 3D light and electron microscopy, we show that many oligodendrocytes are connected to mature and remyelinated myelin sheaths, which we conclude are cells that have reextended processes to contact demyelinated axons while maintaining mature myelin internodes. In the second model in vitamin B12-deficient nonhuman primates, we demonstrate that surviving mature oligodendrocytes extend processes and ensheath demyelinated axons. These data indicate that mature oligodendrocytes can participate in remyelination.

scholarly journals Mode of Bacterial Killing Affects the Inflammatory Response and Associated Organ Dysfunctions in a Porcine E. coli Intensive Care Sepsis Model 

2020 ◽  
Author(s):  
Paul Skorup ◽  
Lisa Maudsdotter ◽  
Miklós Lipcsey ◽  
Anders Larsson ◽  
Jan Sjölin
Keyword(s):  
Inflammatory Response ◽  
Structural Changes ◽  
Interleukin 10 ◽  
Induced Response ◽  
Large Animal ◽  
Necrosis Factor Alpha ◽  
Bacterial Killing ◽  
Treatment Groups ◽  
Sepsis Model

Abstract Background: Sepsis is often treated with penicillin-binding protein 3 (PBP-3) acting b-lactam antibiotics, such as piperacillin-tazobactam, cefotaxime and meropenem. They cause considerable bacterial structural changes and have in vitro been associated with an increased inflammatory response. In a clinically relevant large animal sepsis model our primary aim was to investigate whether bacteria killed by a PBP-3-active antibiotic has a greater effect on the early inflammatory response and organ dysfunction compared with corresponding amounts of live or heat-killed bacteria. A secondary aim was to determine whether the addition of an aminoglycoside could mitigate the cefuroxime-induced response.Method: Killed or live Escherichia coli were administrated as a 3-hour (h) infusion to 16 healthy pigs in a prospective, randomized controlled interventional experimental study. Cefuroxime was chosen as the PBP-3-active antibiotic and tobramycin represented the aminoglycosides. The animals were randomized to receive I) bacteria killed by cefuroxime, II) live bacteria, III) bacteria killed by heat or IV) bacteria killed by the combination of cefuroxime and tobramycin. Plasma endotoxin, tumor necrosis factor alpha, interleukin-6, interleukin-10, leukocytes and organ function were recorded at the start of the experiment and then hourly for 6 h. Results: Differences in dynamics of concentration over time between the four treatment groups were found for the three cytokines (p<0.001). Animals receiving cefuroxime-killed bacteria demonstrated higher responses than those receiving live (p<0.05) or heat-killed bacteria (p<0.01). The addition of tobramycin reduced the cefuroxime-induced responses (p<0.001). The cytokine responses were associated with leucocyte activation that was further associated with pulmonary dysfunction and increases in lactate (p<0.01).Conclusions: In comparison with live or heat-killed bacteria, bacteria killed by a PBP-3-active antibiotic induced an increased inflammatory response that appears to be associated with deteriorated organ and cellular function. The addition of an aminoglycoside to the PBP-3-active antibiotic reduced that response.

Embryos, DOHaD and David Barker

2015 ◽  
Vol 6 (5) ◽  
pp. 377-383 ◽  
Author(s):  
T. P. Fleming ◽  
M. A. Velazquez ◽  
J. J. Eckert
Keyword(s):  
Short Review ◽  
Early Embryo ◽  
Embryonic Cell ◽  
Large Animal ◽  
Permanent Change ◽  
Large Animal Models ◽  
Low Protein ◽  
Periconceptional Period

The early embryo and periconceptional period is a window during which environmental factors may cause permanent change in the pattern and characteristics of development leading to risk of adult onset disease. This has now been demonstrated across small and large animal models and also in the human. Most evidence of periconceptional ‘programming’ has emerged from maternal nutritional models but also other in vivo and in vitro conditions including assisted reproductive treatments, show consistent outcomes. This short review first reports on the range of environmental in vivo and in vitro periconceptional models and resulting long-term outcomes. Second, it uses the rodent maternal low protein diet model restricted to the preimplantation period and considers the stepwise maternal-embryonic dialogue that comprises the induction of programming. This dialogue leads to cellular and epigenetic responses by the embryo, mainly identified in the extra-embryonic cell lineages, and underpins an apparently permanent change in the growth trajectory during pregnancy and associates with increased cardiometabolic and behavioural disease in adulthood. We recognize the important advice of David Barker some years ago to investigate the sensitivity of the early embryo to developmental programming, an insight for which we are grateful.

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