Isolation and characterization of multipotent mesenchymal stromal cells from the gingiva and the periodontal ligament of the horse

<p>Abstract</p> <p>Background</p> <p>The equine periodontium provides tooth support and lifelong tooth eruption on a remarkable scale. These functions require continuous tissue remodeling. It is assumed that multipotent mesenchymal stromal cells (MSC) reside in the periodontal ligament (PDL) and play a crucial role in regulating physiological periodontal tissue regeneration. The aim of this study was to isolate and characterize equine periodontal MSC.</p> <p>Tissue samples were obtained from four healthy horses. Primary cell populations were har-vested and cultured from the gingiva, from three horizontal levels of the PDL (apical, midtooth and subgingival) and for comparison purposes from the subcutis (masseteric region). Colony-forming cells were grown on uncoated culture dishes and typical <it>in vitro </it>characteristics of non-human MSC, i.e. self-renewal capacity, population doubling time, expression of stemness markers and trilineage differentiation were analyzed.</p> <p>Results</p> <p>Colony-forming cell populations from all locations showed expression of the stemness markers CD90 and CD105. In vitro self-renewal capacity was demonstrated by colony-forming unit fibroblast (CFU-F) assays. CFU-efficiency was highest in cell populations from the apical and from the mid-tooth PDL. Population doubling time was highest in subcutaneous cells. All investigated cell populations possessed trilineage differentiation potential into osteogenic, adipogenic and chondrogenic lineages.</p> <p>Conclusions</p> <p>Due to the demonstrated in vitro characteristics cells were referred to as equine subcutaneous MSC (eSc-MSC), equine gingival MSC (eG-MSC) and equine periodontal MSC (eP-MSC). According to different PDL levels, eP-MSC were further specified as eP-MSC from the apical PDL (eP-MSCap), eP-MSC from the mid-tooth PDL (eP-MSCm) and eP-MSC from the subgingival PDL (eP-MSCsg). Considering current concepts of cell-based regenerative therapies in horses, eP-MSC might be promising candidates for future clinical applications in equine orthopedic and periodontal diseases.</p

Task force veterinary anatomy: Joint efforts of the five German veterinary schools to ensure education during the COVID-19 pandemic

At the start of the COVID-pandemic in March 2020, the Institutes of Veterinary Anatomy of the five German educational institutions were confronted with the challenge of digitalising all lectures for the second and fourth semesters of veterinary students. After an online kick-off event and a preliminary status quo meeting, all available digital teaching material was exchanged for students to stream from learning platforms. Lectures were either synchronized or made available as audio recordings and connotated slides on the learning platforms. Fortunately, digital microscopic slides had already been in use, which made it easy for students to access them. Dissection exercises mostly consisted of self- study, using instructive videos and interactive exercises. In the second half of the semester, four of the educational institutions were able offer a restricted number of in-person gross anatomy classes under reinforced conditions. Success monitoring took place online through different formats, and partially on a voluntary basis, via the learning platforms. Although the past two semesters had to almost exclusively take place online due to the unprecedented circumstances, and joint efforts of the five veterinary institutions, there is a general consensus that the practical education in anatomy, histology and embryology is essential to veterinary students. In fact, it is the only way they can obtain the necessary skills to successfully complete the rest of their degree.Die Veterinäranatomischen Institute der fünf deutschen Bildungsstätten standen zu Beginn der Coronapandemie im März 2020 vor der Herausforderung, die gesamte Lehre für die Studierenden des zweiten und vierten Fachsemesters in digitaler Form durchzuführen. Nach einem Kickoff-Onlinemeeting und einer ersten Bestandsaufnahme wurden vorhandene digitale Lehrmedien zum Streamen auf den Lernplattformen ausgetauscht. Vorlesungen wurden synchron gehalten oder asynchron als vertonte bzw. annotierte Dateien auf den Lernplattformen hochgeladen. Die mikroskopischen Präparate konnten den Studierenden dank schon bestehender Anwendungen zur virtuellen Mikroskopie komplikationslos zur Verfügung gestellt werden. Die Präparierübungen wurden überwiegend zum Selbststudium anhand von Lehrvideos und interaktiven Übungsaufgaben angeboten. In der zweiten Semesterhälfte wurden in der makroskopischen Anatomie an vier Standorten einige wenige Präsenztermine für die Studierenden unter verschärften Hygienebedingungen angeboten. Erfolgskontrollen wurden ausschließlich online über die Lernplattformen in verschiedenen Formaten und zum Teil fakultativ durchgeführt. Auch wenn die vergangenen beiden Semester in Zusammenarbeit aller Standorte den Umständen geschuldet fast ausschließlich digital stattfinden musste, besteht der allgemeine Konsens, dass die praktische Ausbildung in den Fächern Anatomie, Histologie und Embryologie essentiell für die Studierenden der Veterinärmedizin ist. Nur so können sie die notwendigen Kenntnisse und Fertigkeiten für ihr weiteres erfolgreiches Studium erlangen

Bone marrow-derived multipotent mesenchymal stromal cells from horses after euthanasia

Abstract Allogeneic equine multipotent mesenchymal stromal cells (eMSCs) have been proposed for use in regenerative therapies in veterinary medicine. A source of allogeneic eMSCs might be the bone marrow from euthanized horses. The purpose of this study was to compare in vitro characteristics of equine bone marrow derived eMSC (eBM-MSCs) from euthanized horses (eut-MSCs) and from narcotized horses (nar-MSCs). Eut-MSCs and nar-MSCs showed typical eMSC marker profiles (positive: CD44, CD90; negative: CD11a/CD18 and MHCII) and possessed tri-lineage differentiation characteristics. Although CD105 and MHCI expression varied, no differences were detected between eut-MSCs and nar-MSCs. Proliferation characteristics did not differ between eut-MSCs and nar-MSCs, but age dependent decrease in proliferation and increase in MHCI expression was detected. These results suggest the possible use of eut-MSCs for therapeutic applications and production of commercial available eBM-MSC products

Crosstalk between Neospora caninum and the bovine host at the maternal-foetal interface determines the outcome of infection

© The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creat iveco mmons .org/licen ses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creat iveco mmons .org/publi cdoma in/ zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.[EN]Neospora caninum is an apicomplexan cyst-forming parasite that is considered one of the main causes of abortion. The pathogenic mechanisms associated with parasite virulence at the maternal-foetal interface that are responsible for the outcome of infection are largely unknown. Here, utilizing placentomes from cattle experimentally infected with high-virulence (Nc-Spain7) and low-virulence (Nc-Spain1H) isolates, we studied key elements of the innate and adaptive immune responses, as well as components of the extracellular matrix (ECM), at 10 and 20 days post-infection (dpi). The low-virulence isolate elicited a robust immune response characterized by upregulation of genes involved in pathogen recognition, chemokines and pro-inflammatory cytokines, crucial for its adequate control. In addition, Nc-Spain1H triggered the expression of anti-inflammatory cytokines and other mechanisms implicated in the maintenance of ECM integrity to ensure foetal survival. In contrast, local immune responses were initially (10 dpi) impaired by Nc-Spain7, allowing parasite multiplication. Subsequently (20 dpi), a predominantly pro-inflammatory Th1-based response and an increase in leucocyte infiltration were observed. Moreover, Nc-Spain7-infected placentomes from animals carrying non-viable foetuses exhibited higher expression of the IL-8, TNF-α, iNOS and SERP-1 genes and lower expression of the metalloproteases and their inhibitors than Nc-Spain7-infected placentomes from animals carrying viable foetuses. In addition, profound placental damage characterized by an alteration in the ECM organization in necrotic foci, which could contribute to foetal death, was found. Two different host-parasite interaction patterns were observed at the bovine placenta as representative examples of different evolutionary strategies used by this parasite for transmission to offspring.SIThis work was supported by the Spanish Ministry of Economy and Competitiveness (AGL2013-44694-R and AGL2016-75935-C2-1-R) and the Community of Madrid (PLATESA2-CM P2018/BAA-4370). Laura Jiménez-Pelayo was financially supported by a fellowship from the University Complutense of Madrid (including two research stays in 2017 and 2018) and Marta García-Sánchez was financially supported through a grant from the Spanish Ministry of Economy and Competitiveness (BES-2014-070723). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Placentation in the anteaters Myrmecophaga tridactyla and Tamandua tetradactyla (Eutheria, Xenarthra)

Background: Since Xenarthra are serious candidates for being basal to Eutheria, their characteristics, e.g. the placental system, influence perceptions of evolution. However, in the subgroup containing the anteaters, data are very limited. The present study aims to elucidate the nature of the feto-maternal interface in the anteater placenta and to interpret these data within an evolutionary context. Methods: Placentas of two species were investigated with histology, immunohistochemistry and transmission electron microscopy. Results: Remnants of the maternal vessel endothelium were absent, resulting in a fully haemochorial barrier throughout the placenta. Two structurally different parts, the villous and trabecular areas were complex and intermingled. In particular, the trabeculae which consisted of cellular, proliferative trophoblast, associated with connective tissue, were attached to the decidua. The villi contained fetal capillaries and hypertrophied mesenchymal cells that occured near the surface near the end of gestation. The surface of the villi consisted of flat, syncytial trophoblast, interspersed with proliferative trophoblast cells. Conclusions: Based on fundamental differences between anteaters and armadillos, we inferred that placental evolution was more complex than previously thought. The haemochorial pattern of anteaters was likely an ancient condition of xenarthrans. Consequently, villous placentation may be attributed, at least in part, by convergent evolution, but was also characterized by some features that were widespread among xenarthrans.This project was supported by CNPq and FAPESP

Immune response of healthy horses to DNA constructs formulated with a cationic lipid transfection reagent

Background Deoxyribonucleic acid (DNA) vaccines are used for experimental immunotherapy of equine melanoma. The injection of complexed linear DNA encoding interleukin (IL)-12/IL-18 induced partial tumour remission in a clinical study including 27 grey horses. To date, the detailed mechanism of the anti-tumour effect of this treatment is unknown. Results In the present study, the clinical and cellular responses of 24 healthy horses were monitored over 72 h after simultaneous intradermal and intramuscular application of equine IL-12/IL-18 DNA (complexed with a transfection reagent) or comparative substances (transfection reagent only, nonsense DNA, nonsense DNA depleted of CG). Although the strongest effect was observed in horses treated with expressing DNA, horses in all groups treated with DNA showed systemic responses. In these horses treated with DNA, rectal temperatures were elevated after treatment and serum amyloid A increased. Total leukocyte and neutrophil counts increased, while lymphocyte numbers decreased. The secretion of tumour necrosis factor alpha (TNFα) and interferon gamma (IFNγ) from peripheral mononuclear blood cells ex vivo increased after treatments with DNA, while IL-10 secretion decreased. Horses treated with DNA had significantly higher myeloid cell numbers and chemokine (C-X-C motif) ligand (CXCL)-10 expression in skin samples at the intradermal injection sites compared to horses treated with transfection reagent only, suggesting an inflammatory response to DNA treatment. In horses treated with expressing DNA, however, local CXCL-10 expression was highest and immunohistochemistry revealed more intradermal IL-12-positive cells when compared to the other treatment groups. In contrast to non-grey horses, grey horses showed fewer effects of DNA treatments on blood lymphocyte counts, TNFα secretion and myeloid cell infiltration in the dermis. Conclusion Treatment with complexed linear DNA constructs induced an inflammatory response independent of the coding sequence and of CG motif content. Expressing IL-12/IL-18 DNA locally induces expression of the downstream mediator CXCL-10. The grey horses included appeared to display an attenuated immune response to DNA treatment, although grey horses bearing melanoma responded to this treatment with moderate tumour remission in a preceding study. Whether the different immunological reactivity compared to other horses may contributes to the melanoma susceptibility of grey horses remains to be elucidated

Correlations between ERG, OCT, and Anatomical Findings in the rd10 Mouse

Background. To evaluate the correlation between ERG, OCT, and microscopic findings in the rd10 mouse. Methods. C57BL/6J wild type mice and rd10 mice were compared at the age of 2, 3, 5, 7, 9, 12, 24, and 48 weeks (each age group n=3) using full-field electroretinography (ERG), spectral domain Optical Coherence Tomography (sd-OCT), fluorescein angiography (FA), Hematoxylin & Eosin histology (HE), and immunohistology (IH). Results. While in wild type mice, the amplitude of a- and b-wave increased with light intensity and with the age of the animals, the rd10 mice showed extinction of the ERG beginning with the age of 5 weeks. In OCT recordings, the thickness of the retina decreased up to 9 weeks of age, mainly based on the degradation of the outer nuclear layer (ONL). Afterwards, the ONL was no longer visible in the OCT. HE staining and immunohistological findings confirmed the in vivo data. Conclusion. ERG and OCT are useful methods to evaluate the retinal function and structure in vivo. The retinal changes seen in the OCT closely match those observed in histological staining