Homing and Engraftment of Intravenously Administered Equine Cord Blood-Derived Multipotent Mesenchymal Stromal Cells to Surgically Created Cutaneous Wound in Horses: A Pilot Project

Abstract

Limb wounds on horses are often slow to heal and are prone to developing exuberant granulation tissue (EGT) and close primarily through epithelialization, which results in a cosmetically inferior and non-durable repair. In contrast, wounds on the body heal rapidly and primarily through contraction and rarely develop EGT. Intravenous (IV) multipotent mesenchymal stromal cells (MSCs) are promising. They home and engraft to cutaneous wounds and promote healing in laboratory animals, but this has not been demonstrated in horses. Furthermore, the clinical safety of administering >1.00 × 108 allogeneic MSCs IV to a horse has not been determined. A proof-of-principle pilot project was performed with two horses that were administered 1.02 × 108 fluorescently labeled allogeneic cord blood-derived MSCs (CB-MSCs) following wound creation on the forelimb and thorax. Wounds and contralateral non-wounded skin were sequentially biopsied on days 0, 1, 2, 7, 14, and 33 and evaluated with confocal microscopy to determine presence of homing and engraftment. Results confirmed preferential homing and engraftment to wounds with persistence of CB-MSCs at 33 days following wound creation, without clinically adverse reactions to the infusion. The absence of overt adverse reactions allows further studies to determine effects of IV CB-MSCs on equine wound healing.

Macroscopic, Histologic, and Immunomodulatory Response of Limb Wounds Following Intravenous Allogeneic Cord Blood-Derived Multipotent Mesenchymal Stromal Cell Therapy in Horses

“…Allogeneic CB-MSCs originating from five unrelated male donor foals (eQcell Inc.; Guelph, ON, Canada) were characterized, prepared, processed, and transported overnight in full concentration HypoThermosol FRS (HTS-FRS) (BioLife Solutions, Bothell, WA, USA) in a cooler approved for transport of mammalian cells (Greenbox 2–8 °C thermal management system, ThermoSafe, Arlington Heights, IL, USA) as previously described [ 26 , 30 , 31 ]. Briefly, CB-MSCs were expanded from cryopreserved sources, and after culture showed a consistent phenotype, had trilineage differentiation capabilities, and had high expression of cluster of differentiation (CD)20, CD44, CD90, and low expression of CD4, CD8, CD11, CD73, major histocompatibility complex (MHC)-I and MHC-II [ 30 , 31 ].…”

“…Cells were passaged 4–5 times and cultured for 35–45 days. CB-MSCs were then harvested using trypsin and ethylenediaminetetraacetic acid, washed with phosphate-buffered saline, and pooled into a mixed population of donor cells [ 21 , 26 , 31 ]. Cells were then resuspended in HTS-FRS prior to being placed in the cooler and shipped to our laboratory.…”

“…The effective cell dose of IV MSCs required to influence wound healing in horses is unknown. Currently, the highest reported dose administered IV to an equid is 1.02 × 10 8 cells per animal [ 26 ], with no clinical adverse responses. Furthermore, the cytokine profile of healing limb wounds in horses has not been reported.…”

Review: Mesenchymal Stem Cell Therapy in Canine Osteoarthritis Research: “Experientia Docet” (Experience Will Teach Us)

“…In contrast, in a study on IV injection of allogeneic adipose tissue-derived MSCs, MSCs labeled with a cell membrane dye were rarely detected in the synovial fluid ( 43 ). In a rabbit and horse study, however, homing of IV injected, respectively, autologous and allogeneic MSCs to places of tissue injury was confirmed ( 70 , 71 ). Olsen et al ( 43 ) reported pulmonary trapping, to early assessment of the synovial fluid, detainment in the synovial membrane and inadequate cell labeling as possible causes of their insufficient MSC homing detection ( 43 ).…”

The Usefulness of Mesenchymal Stem Cells beyond the Musculoskeletal System in Horses

“…Early healing was enhanced, with histology suggesting a pro-healing rather than a pro-inflammatory scenario and smaller sizes in MSC-treated wounds, with no additional advantage of hypoxic preconditioning [ 104 ]. Systemic administration of allogeneic UCB-MSCs resulted in engraftment into induced wounds on the forelimb and thorax at early stages, without clinically adverse reactions [ 105 ]. Autologous PB-SCs injected both locally and systemically in four naturally occurring chronic dermal wounds in the metatarsus, unresponsive to conventional treatments, showed positive outcomes with granulation tissue formed within four weeks, and no adverse effects were noted [ 106 ].…”