IntroductionStudies with mesenchymal stem cells (MSCs) are increasing due to their immunomodulatory, anti-inflammatory and tissue regenerative properties. However, there is still no agreement about the best source of equine MSCs for a bank for allogeneic therapy. The aim of this study was to evaluate the cell culture and immunophenotypic characteristics and differentiation potential of equine MSCs from bone marrow (BM-MSCs), adipose tissue (AT-MSCs) and umbilical cord (UC-MSCs) under identical in vitro conditions, to compare these sources for research or an allogeneic therapy cell bank.MethodsThe BM-MSCs, AT-MSCs and UC-MSCs were cultured and evaluated in vitro for their osteogenic, adipogenic and chondrogenic differentiation potential. Additionally, MSCs were assessed for CD105, CD44, CD34, CD90 and MHC-II markers by flow cytometry, and MHC-II was also assessed by immunocytochemistry. To interpret the flow cytometry results, statistical analysis was performed using ANOVA.ResultsThe harvesting and culturing procedures of BM-MSCs, AT-MSCs and UC-MSCs were feasible, with an average cell growth until the third passage of 25 days for BM-MSCs, 15 days for AT-MSCs and 26 days for UC-MSCs. MSCs from all sources were able to differentiate into osteogenic (after 10 days for BM-MSCs and AT-MSCs and 15 days for UC-MSCs), adipogenic (after 8 days for BM-MSCs and AT-MSCs and 15 days for UC-MSCs) and chondrogenic (after 21 days for BM-MSCs, AT-MSCs and UC-MSCs) lineages. MSCs showed high expression of CD105, CD44 and CD90 and low or negative expression of CD34 and MHC-II. The MHC-II was not detected by immunocytochemistry techniques in any of the MSCs studied.ConclusionsThe BM, AT and UC are feasible sources for harvesting equine MSCs, and their immunophenotypic and multipotency characteristics attained minimal criteria for defining MSCs. Due to the low expression of MHC-II by MSCs, all of the sources could be used in clinical trials involving allogeneic therapy in horses. However, the BM-MSCs and AT-MSCs showed fastest ‘‘in vitro’’ differentiation and AT-MSCs showed highest cell growth until third passage. These findings suggest that BM and AT may be preferable for cell banking purposes.

Pesq. Vet. Bras. 34(8) The incidence of obstructive urolithiasis in sheep is high, especially in feedlot males, both for meat production, or the breeder of high genetic value. The urinary acidification is one way to prevent this disease and can be performed effectively supplementation with ammonium chloride in the diet, which may facilitate the installation of metabolic acidosis. The blood gas analysis evaluates the acid-base balance of blood in a practical and easy way. In this study, it was evaluated the effect of ammonium chloride on acid-base and electrolyte in feedlot sheep blood gas analysis to determine the occurrence of metabolic acidosis. It was used 100 male lambs, in a feedlot, aged approximately three months. It was constituted three groups: Group I (n=40) that received 400mg/kg/PV of ammonium chloride/animal/ day for 21 consecutive days, the time of discontinuation of the urinary acidifiers (M3) and continued clinical follow until the end of the experiment (M6); Group II (n=40), that received 400mg/kg/PV of ammonium chloride/animal/day for 42 consecutive days, Group III (n=20), that did not receive ammonium chloride throughout the experimental period. The moments (M) of samples and clinical assessment were established on seven days of interval, M0 (immediately before the beginning of the treatment with ammonium chloride), M1 (seven days after), M2, M3, M4, M5 and M6, totalizing 56 days of feedlot. The feed consisted of a total mixed ration consisting of 15% of ground hay and 85 % of concentrate, water and mineral salt ad libitum. After 15 days of adaptation to the diet of feedlot, urine samples for measurement of pH, and venous blood for blood gas analysis were collected from all animals at different moments. The urinary acidification was maintained as was the administration of ammonium chloride in GI and GII. The values of Na + and K + remained within the normal range for the species. Ammonium chloride caused metabolic acidosis compensated change in GI and GII, confirmed by values of HCO 3 -and EB below the reference values, with a normal pH, and high levels of Cl -, and decreased SID. It was concluded that although ammonium chloride to cause decrease of alkalinity in the body, caused no loss in animal development and can be used as a preventive agent obstructive urolithiasis in sheep.INDEX TERMS: Ammonium chloride, electrolyte balance, acid-base balance, metabolic acidosis, urinary acidifiers, hyperchloremia, small ruminants, urolithiasis.

The aim of this study was to isolate, culture, and characterize mesenchymal stem cells (MSCs) from horse bone marrow (BM) using the techniques of flow cytometry, immunocytochemistry, cytogenetics, and electron microscopy. Immunophenotypic analysis revealed the presence of MSCs with high expression of the CD90 marker, lower expression of the CD44 marker, and absent expression of the CD34 marker. In assays of differentiation, the positive response to osteogenic (OST), chondrogenic (CDG), and adipogenic (ADP) differentiation signals was observed and characterized by deposition of calcium-rich extracellular matrix (OST), proteoglycans and collagen II (CDG) and intracellular deposition of fat drops (ADP). In immunocytochemical characterization, MSCs were immunopositive for CD44, vimentin, and PCNA, and they were negative for CD13. In the ultrastructural analysis of MSCs, the most outstanding characteristic was the presence of rough endoplasmic reticulum with very dilated cisterns filled with a low electrodensity material. Additionally, MSCs had normal karyotypes (2n = 64) as evidenced by cytogenetic analysis, and aneuploidy in metaphase was not observed. The protocols for isolating, culturing, and characterizing equine MSCs used in this study were shown to be appropriate for the production of a cell population with a good potential for differentiation and without aneuploidy that can be used to study future cellular therapies.

BackgroundIt is currently unknown if the intrathecal administration of a high dose of allogeneic mesenchymal stem cells (MSCs) is safe, how MSCs migrate throughout the vertebral canal after intrathecal administration, and whether MSCs are able to home to a site of injury. The aims of the study were: 1) to evaluate the safety of intrathecal injection of 100 million allogeneic adipose-derived MSCs (ASCs); 2) to assess the distribution of ASCs after atlanto-occipital (AO) and lumbosacral (LS) injection in healthy horses; and 3) to determine if ASCs homed to the site of injury in neurologically diseased horses.MethodsSix healthy horses received 100 × 106 allogeneic ASCs via AO (n = 3) or LS injection (n = 3). For two of these horses, ASCs were radiolabeled with technetium and injected AO (n = 1) or LS (n = 1). Neurological examinations were performed daily, and blood and cerebrospinal fluid (CSF) were evaluated prior to and at 30 days after injection. Scintigraphic images were obtained immediately postinjection and at 30 mins, 1 h, 5 h, and 24 h after injection. Three horses with cervical vertebral compressive myelopathy (CVCM) received 100 × 106 allogeneic ASCs labeled with green fluorescent protein (GFP) via AO injection and were euthanized 1–2 weeks after injection for a full nervous system necropsy. CSF parameters were compared using a paired student’s t test.ResultsThere were no significant alterations in blood, CSF, or neurological examinations at any point after either AO or LS ASC injections into healthy horses. The radioactive signal could be identified all the way to the lumbar area after AO ASC injection. After LS injection, the signal extended caudally but only a minimal radioactive signal extended further cranially. GFP-labeled ASCs were not present at the site of disease at either 1 or 2 weeks following intrathecal administration.ConclusionsThe intrathecal injection of allogeneic ASCs was safe and easy to perform in horses. The AO administration of ASCs resulted in better distribution within the entire subarachnoid space in healthy horses. ASCs could not be found after 7 or 15 days of injection at the site of injury in horses with CVCM.Electronic supplementary materialThe online version of this article (10.1186/s13287-018-0849-6) contains supplementary material, which is available to authorized users.

Realizou-se estudo retrospectivo (1987-2002) dos aspectos clínicos das fraturas vertebrais em eqüinos, bovinos, ovinos, caprinos e suínos atendidos no hospital veterinário da FMVZ-Unesp de Botucatu. Todos os casos tiveram confirmação radiográfica ou post-mortem. Segundo a espécie, a ordem de acometimento foi: bovina, eqüina, ovina, caprina e suína. As lesões ocorreram desde os 12 dias de idade até os 10 anos, com maior freqüência até os 12 meses. O segmento torácico foi o mais comprometido seguido dos segmentos lombar, cervical e sacral. As fraturas vertebrais devem fazer parte da lista de diagnósticos diferenciais de animais em decúbito, independente da espécie, sexo ou idade.

BackgroundRecent studies have demonstrated numerous biological properties of mesenchymal stem cells and their potential application in treating complex diseases or injuries to tissues that have difficulty regenerating, such as those affecting the central and peripheral nervous system. Thus, therapies that use mesenchymal stem cells are promising because of their high capacity for self-regeneration, their low immunogenicity, and their paracrine, anti-inflammatory, immunomodulatory, anti-apoptotic and neuroprotective effects. In this context, the purpose of this study was to evaluate the feasibility and safety of intrathecal transplantation of bone marrow-derived mesenchymal stem cells in horses, for future application in the treatment of neurological diseases.ResultsDuring the neurological evaluations, no clinical signs were observed that were related to brain and/or spinal cord injury of the animals from the control group or the treated group. The hematological and cerebrospinal fluid results from day 1 and day 6 showed no significant differences (P > 0.05) between the treated group and the control group. Additionally, analysis of the expression of matrix metalloproteinase (MMP) -2 and −9 in the cerebrospinal fluid revealed only the presence of pro-MMP-2 (latent), with no significant difference (P > 0.05) between the studied groups.ConclusionsThe results of the present study support the hypothesis of the feasibility and safety of intrathecal transplantation of autologous bone marrow-derived mesenchymal stem cells, indicating that it is a promising pathway for cell delivery for the treatment of neurological disorders in horses.

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