Orthopedic Lesions in Horses

Introduction

In horses with limb fractures, delayed mending due to implant failure is a common problem. In addition, when instability is present, the affected horse often develops laminitis in the support limb. Non-mended fractures are more common when they are insufficiently immobilized or as a result of infection. In adult horse fracture repair, particularly in fractures of the upper limb, low-grade instability often causes implant failure and results in lameness in the treated limb. If lameness persists, then overuse of the contralateral limb occurs, resulting in laminitis in the supporting limb. Therefore, an important goal in equine fracture repair is to expedite bone mending to help prevent these fatal complications. The injection of adult-derived MSCs directly into the fracture site represents a promising mechanism to accelerate fracture healing and thereby decrease the risk limb fractures.

Studies

1. Isolation and Differentiation of Horse MSCs

To determine the optimal bone forming source of equine mesenchymal stem cells (eMSCs) and optimize collection of and expansion conditions for those cells we isolated eMSCs from bone marrow (BM), fat tissue, and umbilical cord blood and tissue, and the bone forming potential of each type was assessed. Effects of anatomic site, aspiration volume, and serum type on eMSC yield from BM were investigated. BM-eMSCs had the highest overall expression of the bone forming genes and staining for alkaline phosphatase activity and calcium deposition (measures of bone production). The BM-eMSC expansion rate was significantly higher when cells were cultured in fetal bovine serum instead of autologous serum. In conclusion, eMSCs isolated from BM possessed the highest in vitro bone forming potential; eMSCs from fat tissue also had robust bone forming potential. Expanding BM-eMSCs in autologous serum to avoid potential immunologic reactions decreased the total yield because BM-eMSCs grew significantly slower than in fetal bovine serum. Additional studies are needed to determine optimal ex vivo eMSC culture and expansion conditions, including timing and use of growth factor-supplemented culture medium.

Investigators: Claire Yellowley, Dori Borjesson, Larry Galuppo

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2. Horse MSC Mediated Immunomodulation

Mesenchymal stem cells (MSCs) are used in both human clinical trials and veterinary medicine for the treatment of inflammatory and immune-mediated diseases.  MSCs modulate inflammation by decreasing the cells and products of the inflammatory response. Stimulated equine MSCs from bone marrow (BM), adipose tissue (AT), cord blood (CB), and umbilical cord tissue (CT) inhibit lymphocyte proliferation and decrease inflammatory cytokine production. We hypothesized that equine MSCs inhibit T cell proliferation through secreted mediators and that MSCs from different tissue sources decrease T cell proliferation through different mechanisms. To test our hypotheses, we inhibited interleukin-6 (IL-6), nitric oxide (NO), and prostaglandin E2 (PGE2) to determine their impact on stimulated T cell proliferation. Inhibition of IL-6 or NO did not reverse the immunomodulatory effect of MSCs on activated T cells. In contrast, inhibition of PGE2 restored T cell proliferation, restored the secretion of tumor necrosis factor-a and interferon-g, and increased IL-10 levels. MSCs from solid-tissue-derived sources, AT and CT, inhibited T cell proliferation through induction of lymphocyte apoptosis while blood-derived MSCs, BM and CB, induced lymphocyte cell cycle arrest. Equine MSCs from different tissue sources modulated immune cell function by both overlapping and unique mechanisms. MSC tissue source may determine immunomodulatory properties of MSCs and may have very practical implications for MSC selection in the application of MSC therapy.

Investigators: Dori Borjesson, Larry Galuppo

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