Equine Bone Marrow-Derived Mesenchymal Stem Cells -

Evaluation of Muscle Inflammatory Response Following Intramuscular Transplantation

Abstract

Background: Horses hold significant value in sports and the economy, requiring rapid recovery from musculoskeletal injuries. Cell-based therapies, particularly those using mesenchymal stem cells (MSCs), have emerged as promising tools due to their regenerative, anti-inflammatory, and immunomodulatory properties, as well as their capacity for self-renewal. However, their therapeutic efficacy may be influenced by host immune responses, particularly in allogeneic settings. Thus, this study aimed to morphologically evaluate the acute inflammatory response in equine muscle tissue following intramuscular transplantation of autologous and allogeneic bone marrow-derived MSCs (EqBM-MSCs).

Materials, Method & Results: Fifteen clinically healthy adult horses (6-12 years; 300-500 kg) were randomly assigned to 3 groups (n = 5 each): Control (intramuscular injection of phosphate-buffered saline - PBS), Auto-MSCs (intramuscular transplantation of autologous EqBM-MSCs), and Allo-MSCs (intramuscular transplantation of allogeneic EqBM-MSCs). EqBM-MSCs were isolated from bone marrow, expanded under standard culture conditions, and characterized by plastic adherence, fibroblastoid morphology, expression of CD44 and CD90, absence of CD34, and their differentiation potential into adipogenic, osteogenic, and chondrogenic lineages. An average of 2×10⁶ cells in 100 µL of PBS were transplanted into the right gluteus medius muscle under ultrasound guidance. Two muscle biopsies were collected per horse before transplantation (D0) and 24 h post-transplantation (D24). Histological hematoxylin and eosin (H&E); Gomori’s trichrome (GT)) and histochemical (nicotinamide adenine dinucleotide tetrazolium reductase (NADH-tr)) analyses demonstrated preserved muscle architecture and normal fiber organization in both MSC-treated groups. No evidence of inflammation, such as fiber morphological alterations or inflammatory cell infiltration, was observed in any group.

Discussion: MSCs possess self-renewal capacity and secrete bioactive factors that support tissue repair and modulate inflammation. In equines, bone marrow is a commonly used MSC source, and intramuscular delivery has been associated with efficient administration and prolonged local retention. The absence of acute muscle inflammatory changes observed in this study is consistent with previous reports in equine tendon following MSC transplantation. In contrast, inflammatory responses have been described in rat muscle after autologous bone marrow MSC administration, typically characterized by muscle fiber alterations and inflammatory cell infiltration. Such changes were not observed in the present study, and NADH-tr staining confirmed preserved metabolic activity. These findings indicate that intramuscular administration of both autologous and allogeneic EqBM-MSCs is well-tolerated and does not induce acute inflammation. The lack of acute response may be related to the immunomodulatory properties of MSCs, including secretion of factors such as transforming growth factor beta (TGF-β) and prostaglandin E2 (PGE2). The absence of inflammatory changes supports the immunomodulatory properties of MSCs and may reinforces their potential for clinical application in equine muscle injury. Nevertheless, the evaluation performed at 24 h reflects only the acute phase, and longer-term responses remain to be elucidated. Further studies are necessary to investigate the long-term safety and therapeutic potential of intramuscular MSC transplantation in equine musculoskeletal disorders.

Keywords: autologous, cell therapy, gluteus medius, histology, histochemistry.