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Topic:Stem Cells
Stem cells in horses refer to undifferentiated cells capable of self-renewal and differentiation into specialized cell types. These cells are primarily utilized in regenerative medicine and therapeutic applications to repair or replace damaged tissues in equine patients. Common sources of stem cells in horses include bone marrow, adipose tissue, and umbilical cord blood. Research in equine stem cell therapy focuses on understanding their potential to treat musculoskeletal injuries, such as tendon and ligament damage, and exploring their mechanisms of action. This page compiles peer-reviewed research studies and scholarly articles that examine the isolation, characterization, and therapeutic applications of stem cells in equine medicine.
Further insights into the characterization of equine adipose tissue-derived mesenchymal stem cells. Adipose tissue-derived stem cells (ADSCs) represent a promising subpopulation of adult stem cells for tissue engineering applications in veterinary medicine. In this study we focused on the morphological and molecular biological properties of the ADSCs. The expression of stem cell markers Oct4, Nanog and the surface markers CD90 and CD105 were detected using RT-PCR. ADSCs showed a proliferative potential and were capable of adipogenic and osteogenic differentiation. Expression of Alkaline phosphatase (AP), phosphoprotein (SPP1), Runx2 and osteocalcin (OC) mRNA were positive in osteogenic linea...
Evaluation of senescence in mesenchymal stem cells isolated from equine bone marrow, adipose tissue, and umbilical cord tissue. Mesenchymal stem cells (MSCs) from adult and neonatal tissues are intensively investigated for their use in regenerative medicine. The purpose of this study was to compare the onset of replicative senescence in MSCs isolated from equine bone marrow (BMSC), adipose tissue (ASC), and umbilical cord tissue (UCMSC). MSC proliferation (cell doubling), senescence-associated β-galactosidase staining, telomere length, Sox-2, and lineage-specific marker expression were assessed for MSCs harvested from tissues of 4 different donors. The results show that before senescence ensued, all cell types prolife...
Decreased expression of p63, a regulator of epidermal stem cells, in the chronic laminitic equine hoof. Abnormal epidermal stem cell regulation may contribute to the pathogenesis of equine chronic laminitis. Objective: To analyse the involvement of p63, a regulator of epidermal stem cell proliferative potential, in chronic laminitis. Methods: Epidermal tissues from skin, coronet and lamellae of the dorsal foot were harvested from 5 horses with chronic laminitis and 5 control horses. Tissues were analysed using histopathology, immunofluorescence microscopy and quantitative immunoblotting. Results: Hoof lamellae of laminitic horses had a lower frequency of p63 positive cells than control lamellae,...
Induced pluripotent stem cell lines derived from equine fibroblasts. The domesticated horse represents substantial value for the related sports and recreational fields, and holds enormous potential as a model for a range of medical conditions commonly found in humans. Most notable of these are injuries to muscles, tendons, ligaments and joints. Induced pluripotent stem (iPS) cells have sparked tremendous hopes for future regenerative therapies of conditions that today are not possible to cure. Equine iPS (EiPS) cells, in addition to bringing promises to the veterinary field, open up the opportunity to utilize horses for the validation of stem cell based therapi...
The regenerative medicine laboratory: facilitating stem cell therapy for equine disease. This article focuses on the emerging field of equine regenerative medicine with an emphasis on the use of mesenchymal stem cells (MSCs) for orthopedic diseases. We detail laboratory procedures and protocols for tissue handling and MSC isolation, characterization, expansion, and cryopreservation from bone marrow, fat, and placental tissues. We provide an overview of current clinical uses for equine MSCs and how MSCs function to heal tissues. Current laboratory practices in equine regenerative medicine mirror those in the human field. However, the translational use of autologous and allogeneic M...
Fetal derived embryonic-like stem cells improve healing in a large animal flexor tendonitis model. Tendon injury is a common problem in athletes, with poor tissue regeneration and a high rate of re-injury. Stem cell therapy is an attractive treatment modality as it may induce tissue regeneration rather than tissue repair. Currently, there are no reports on the use of pluripotent cells in a large animal tendon model in vivo. We report the use of intra-lesional injection of male, fetal derived embryonic-like stem cells (fdESC) that express Oct-4, Nanog, SSEA4, Tra 1-60, Tra 1-81 and telomerase. Methods: Tendon injury was induced using a collagenase gel-physical defect model in the mid-metacar...
Biochemical identification and immunolocalizaton of aggrecan, ADAMTS5 and inter-alpha-trypsin-inhibitor in equine degenerative suspensory ligament desmitis. We describe analysis of suspensory ligaments from horses with advanced degenerative suspensory ligament desmitis (DSLD) to identify the major proteoglycans (PGs), ADAMTS-aggrecanases and inter-alpha-trypsin inhibitor (IαI) components associated with ligament degeneration. Specific anatomical regions of suspensory ligaments from two normal horses and four diagnosed with DSLD were analyzed by Western blot and immunohistochemistry for the following: aggrecan, aggrecan fragments, decorin, ADAMTS4, ADAMTS5, and IαI components. When compared to normal, DSLD ligaments showed about a 15-fold increas...
Evaluation of equine peripheral blood apheresis product, bone marrow, and adipose tissue as sources of mesenchymal stem cells and their differentation potential. To evaluate effects of apheresis on mesenchymal stem cells (MSCs) and compare those MSCs with MSCs obtained from adipose tissue or bone marrow (BM). Methods: Samples obtained from 6 adult horses. Methods: Samples of blood from a peripheral vein, adipose tissue, and BM aspirate were obtained from each horse. Samples were processed via apheresis of blood and techniques reported elsewhere for adipose tissue and BM. Cultures were maintained until adherence and subsequently were subjected to differentiation protocols to evaluate adipogenic, osteoblastogenic, and chondrogenic potential. Results: Aph...
Trophoblast stem cell marker gene expression in inner cell mass-derived cells from parthenogenetic equine embryos. Although putative horse embryonic stem (ES)-like cell lines have been obtained recently from in vivo-derived embryos, it is currently not known whether it is possible to obtain ES cell (ESC) lines from somatic cell nuclear transfer (SCNT) and parthenogenetic (PA) embryos. Our aim is to establish culture conditions for the derivation of autologous ESC lines for cell therapy studies in an equine model. Our results indicate that both the use of early-stage blastocysts with a clearly visible inner cell mass (ICM) and the use of pronase to dissect the ICM allow the derivation of a higher proportion...
Markers of stemness in equine mesenchymal stem cells: a plea for uniformity. Mesenchymal stromal cells (MSC) are a very promising subpopulation of adult stem cells for cell-based regenerative therapies in veterinary medicine. Despite major progress in the knowledge on adult stem cells during recent years, a proper identification of MSC remains a challenge. In human medicine, the Mesenchymal and Tissue Stem Cell Committee of the International Society for Cellular Therapy (ISCT) recently proposed three criteria to define MSC. Firstly, cells must be plastic-adherent when maintained under standard culture conditions. Secondly, MSC must express CD73, CD90 and CD105, and lac...
Comparison of equine bone marrow-, umbilical cord matrix and amniotic fluid-derived progenitor cells. The aim of the study was to compare in vitro the stemness features of horse progenitor cells derived from bone marrow (BM-MSCs), amniotic fluid (AF-MSCs) and umbilical cord matrix (EUC-MSCs). It has been suggested that there may be a stem cell population within both umbilical cord matrix and amniotic fluid. However, little knowledge exists about the characteristics of these progenitor cells within these sources in the equine species. This study wanted to investigate an alternative and non-invasive stem cell source for the equine tissue engineering and to learn more about the properties of thes...
Expansion of mesenchymal stem cells on fibrinogen-rich protein surfaces derived from blood plasma. Mesenchymal stem cells (MSCs) are present in low density in bone marrow and culture expansion is necessary to obtain sufficient numbers for many proposed therapies. Researchers have characterized MSC growth on tissue culture plastic (TCP), although few studies have explored proliferation on other growth substrates. Using adult equine MSCs, we evaluated proliferation on fibrinogen-rich precipitate (FRP) surfaces created from blood plasma. When seeded at 1 × 10(4) cells/cm(2) and passaged five times over 10 days, MSCs on FRP in medium containing fibroblast growth factor 2 (FGF2) resulted in a ...
Clinicopathologic findings following intra-articular injection of autologous and allogeneic placentally derived equine mesenchymal stem cells in horses. The development of an allogeneic mesenchymal stem cell (MSC) product to treat equine disorders would be useful; however, there are limited in vivo safety data for horses. We hypothesized that the injection of self (autologous) and non-self (related allogeneic or allogeneic) MSC would not elicit significant alterations in physical examination, gait or synovial fluid parameters when injected into the joints of healthy horses. Methods: Sixteen healthy horses were used in this study. Group 1 consisted of foals (n = 6), group 2 consisted of their dams (n = 5) and group 3 consisted of half-siblings ...
Effects of non-steroidal anti-inflammatory drugs on proliferation, differentiation and migration in equine mesenchymal stem cells. In equine medicine, stem cell therapies for orthopaedic diseases are routinely accompanied by application of NSAIDs (non-steroidal anti-inflammatory drugs). Thus, it has to be analysed how NSAIDs actually affect the growth and differentiation potential of MSCs (mesenchymal stem cells) in vitro in order to predict the influence of NSAIDs such as phenylbutazone, meloxicam, celecoxib and flunixin on MSCs after grafting in vivo. The effects of NSAIDs were evaluated regarding cell viability and proliferation. Additionally, the multilineage differentiation capacity and cell migration was analysed. N...
Adipogenic differentiation of adult equine mesenchymal stromal cells. Equine adipose tissue-derived mesenchymal stem cells (ASCs) have only recently been investigated for their adipogenic, chondrogenic, and osteogenic differentiation potential. This chapter will briefly outline the molecular mechanisms leading to adipogenesis and the methods of equine adipose tissue harvest, ASC isolation, and adipogenic differentiation. The reader is also directed to other reported methods of adipogenesis for ASCs and mesenchymal stem cells (MSCs) from other tissues.
Hydrolyzed fish collagen induced chondrogenic differentiation of equine adipose tissue-derived stromal cells. Adipose-derived stromal cells (ADSCs) are multipotent cells which, in the presence of appropriate stimuli, can differentiate into various lineages such as the osteogenic, adipogenic and chondrogenic. In this study, we investigated the effect of transforming growth factor beta 1 (TGF-β1) in comparison to hydrolyzed fish collagen in terms of the chondrogenic differentiation potential of ADSCs. ADSCs were isolated from subcutaneous fat of horses by liposuction. Chondrogenesis was investigated using a pellet culture system. The differentiation medium was either supplemented with TGF-β1 (5 ng/ml)...
Analysis of CD14 expression levels in putative mesenchymal progenitor cells isolated from equine bone marrow. A long-term goal of mesenchymal progenitor cell (MPC) research is to identify cell-surface markers to facilitate MPC isolation. One reported MPC feature in humans and other species is lack of CD14 (lipopolysaccharide receptor) expression. The aim of this study was to evaluate CD14 as an MPC sorting marker. Our hypothesis was that cells negatively selected by CD14 expression would enrich MPC colony formation compared with unsorted and CD14-positive fractions. After validation of reagents, bone marrow aspirate was obtained from 12 horses. Fresh and cultured cells were analyzed by flow cytometry ...
Comparison of the osteogenic potential of equine mesenchymal stem cells from bone marrow, adipose tissue, umbilical cord blood, and umbilical cord tissue. To determine the optimal osteogenic source of equine mesenchymal stem cells (eMSCs) and optimize collection of and expansion conditions for those cells. Methods: 10 adult Quarter Horses and 8 newborn Thoroughbred foals. Methods: eMSCs were isolated from bone marrow (BM), adipose tissue, and umbilical cord blood and tissue, and the osteogenic potential of each type was assessed. Effects of anatomic site, aspiration volume, and serum type on eMSC yield from BM were investigated. Results: BM-eMSCs had the highest overall expression of the osteogenic genes Cbfa1, Osx, and Omd and staining for ALP ...
Evaluation of the osteogenic and chondrogenic differentiation capacities of equine adipose tissue-derived mesenchymal stem cells. To evaluate the proliferative behavior, telomere length, immunophenotype, and differentiation capacity of equine adipose tissue-derived mesenchymal stem cells (AT-MSCs). Methods: 6 adult racing horses treated for articular Injury but otherwise healthy. Methods: AT-MSCs were Isolated from horses and expanded In Dulbecco modified Eagle medium enriched with fetal bovine serum and antimicrobials. Expression of cell surface antigens and telomere length were Investigated via flow cytometry Differentiation of MSCs Into chondrocytes, osteoblasts, and adipocytes was Induced In vitro by specific stimuli...
Virally and physically transgenized equine adipose-derived stromal cells as a cargo for paracrine secreted factors. Adipose-Derived Stromal Cells have been shown to have multiple lineage differentiation properties and to be suitable for tissues regeneration in many degenerative processes. Their use has been proposed for the therapy of joint diseases and tendon injuries in the horse. In the present report the genetic manipulation of Equine Adipose-Derived Stromal Cells has been investigated. Results: Equine Adipose-Derived Stromal Cells were successfully virally transduced as well as transiently and stably transfected with appropriate parameters, without detrimental effect on their differentiation properties...
Isolation and characterization of equine amniotic fluid-derived multipotent stem cells. Amniotic fluid (AF) is a well-known source of stem cells. However, there have been no reports regarding equine AF stem cells. We have isolated equine AF-derived multipotent stem cells (MSC) (eAF-MSC) and show that these cells exhibit self-renewal ability and multilineage differentiation. Methods: AF was obtained from thoroughbred mares and mononuclear cells (MNC) were isolated by Ficoll-Paque density gradient. We measured the cumulative population doubling level (CPDL) and characterized the immunophenotype by flow cytometry. To investigate differentiation ability, a trilineage differentiation ...
Equine embryonic stem-like cells and mesenchymal stromal cells have different survival rates and migration patterns following their injection into damaged superficial digital flexor tendon. Injury to the superficial digital flexor tendon (SDFT) is common in racing and sport horses and poor tendon regeneration leads to high reinjury rates. Autologous mesenchymal stromal cells (MSCs) are being used clinically to improve tendon regeneration but they have some practical limitations. Embryonic stem cells (ESCs) may overcome these limitations but their fate following injection into the damaged SDFT is unknown. Objective: To inject MSCs and ESCs into distinct areas of damage in the SDFT and monitor their survival over a 3 month period. Methods: MSCs and ESCs expressing different reporte...
Isolation of equine bone marrow-derived mesenchymal stem cells: a comparison between three protocols. There is a need to assess and standardise equine bone marrow (BM) mesenchymal stem cell (MSC) isolation protocols in order to permit valid comparisons between therapeutic trials at different sites. Objective: To compare 3 protocols of equine BM MSC isolation: adherence to a plastic culture dish (Classic) and 2 gradient density separation protocols (Percoll and Ficoll). Methods: BM aspirates were harvested from the sternum of 6 mares and MSCs isolated by all 3 protocols. The cell viability after isolation, MSC yield, number of MSCs attained after 14 days of culture and the functional characteri...
Adult bone marrow stromal cell-based tissue-engineered aggrecan exhibits ultrastructure and nanomechanical properties superior to native cartilage. To quantify the structural characteristics and nanomechanical properties of aggrecan produced by adult bone marrow stromal cells (BMSCs) in peptide hydrogel scaffolds and compare to aggrecan from adult articular cartilage. Methods: Adult equine BMSCs were encapsulated in 3D-peptide hydrogels and cultured for 21 days with TGF-β1 to induce chondrogenic differentiation. BMSC-aggrecan was extracted and compared with aggrecan from age-matched adult equine articular cartilage. Single molecules of aggrecan were visualized by atomic force microscopy-based imaging and aggrecan nanomechanical stiffness...
Coculture of equine mesenchymal stem cells and mature equine articular chondrocytes results in improved chondrogenic differentiation of the stem cells. Bone marrow derived mesenchymal stem cells (MSCs) can be used to repair articular cartilage defects, these cells should be properly stimulated so that they could differentiate morphologically and hold cellular synthetic features closer to maturely differentiated chondrocytes. It is well known that tissue specific environment plays an important role in cell fate determination. Once improved isolation, proliferation and differentiation protocols have been developed, the likelihood of spontaneous differentiation of MSCs into divergent lineages will be reduced, thus increasing their value for cart...
Osteogenic comparison of expanded and uncultured adipose stromal cells. Adipose stromal cells (ASC) are a promising alternative to progenitor cells from other tissue compartments because of their multipotential and capacity to retrieve significantly more progenitor cells. Initial cell samples are heterogeneous, containing a collection of cells that may contribute to tissue repair, but the sample becomes more homogeneous with each passage. Therefore, we hypothesized that the osteogenic potential of culture-expanded ASC would differ from uncultured ASC. Methods: Adipose tissue was collected from a yearling colt, and ASC were isolated and expanded using standard prot...
Histological and immunohistochemical evaluation of autologous cultured bone marrow mesenchymal stem cells and bone marrow mononucleated cells in collagenase-induced tendinitis of equine superficial digital flexor tendon. The aim of this study was to compare treatment with cultured bone marrow stromal cells (cBMSCs), bone marrow Mononucleated Cells (BMMNCs), and placebo to repair collagenase-induced tendinitis in horses. In six adult Standardbred horses, 4000 IU of collagenase were injected in the superficial digital flexor tendon (SDFT). Three weeks after collagenase treatment, an average of either 5.5 x 10(6) cBMSCs or 1.2 x 10(8) BMMNCs, fibrin glue, and saline solution was injected intralesionally in random order. In cBMSC- and BMMNCS-treated tendons, a high expression of cartilage oligomeric matrix protein...
Adult equine bone marrow stromal cells produce a cartilage-like ECM mechanically superior to animal-matched adult chondrocytes. Our objective was to evaluate the age-dependent mechanical phenotype of bone marrow stromal cell- (BMSC-) and chondrocyte-produced cartilage-like neo-tissue and to elucidate the matrix-associated mechanisms which generate this phenotype. Cells from both immature (2-4 month-old foals) and skeletally-mature (2-5 year-old adults) mixed-breed horses were isolated from animal-matched bone marrow and cartilage tissue, encapsulated in self-assembling-peptide hydrogels, and cultured with and without TGF-beta1 supplementation. BMSCs and chondrocytes from both donor ages were encapsulated with high viab...
