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Equine veterinary journal2026; doi: 10.1002/evj.70196

Composition of allogeneic equine orthobiologics and their repeated intra-articular administrations.

Abstract: Orthobiologics such as platelet-rich plasma (PRP) and alpha-2-macroglobulin (A2MG) are increasingly used for equine musculoskeletal injuries. However, their composition and safety with repeated allogeneic intra-articular administration remain poorly investigated. Objective: To characterise orthobiological preparations with or without 1 h incubation at 37°C (PRP, PRP + Inc, A2MG, A2MG + Inc) and evaluate immunological safety following repeated intra-articular administration (autologous and allogeneic) of PRP combined with A2MG + Inc in healthy horses. Methods: In vivo experiments. Methods: This was a two-phase study with biochemical characterisation followed by repeated intra-articular injections in four healthy horses. Blood was processed to produce PRP or A2MG, with or without incubation, and characterised using haematology, biochemistry and enzyme-linked immunosorbent assay. For immunogenic assessment, healthy horses received three intra-articular injections of combined PRP and A2MG + Inc at 2-week intervals (Days 0, 14 and 28) (allogeneic n = 3, autologous n = 1). Blood and synovial fluid were collected on Days 0, 14, 28 and 42. Blood was analysed for haematology, biochemistry, CD4/CD8 ratio and SAA, while synovial fluid was analysed for leukocyte counts and immunoglobulins. Results: Biochemical characterisation demonstrated effective removal of blood cells from all orthobiologics. PRP-based formulations were enriched in platelets and growth factors, whereas A2MG preparations had higher alpha-2-macroglobulin and lower growth factor levels. Repeated intra-articular administration of PRP and A2MG + Inc caused no clinically relevant changes in systemic or synovial immune cells, lymphocyte subsets, or acute-phase markers, indicating no immunological activation. Conclusions: Small sample sizes, the limited scope of characterised compositions and the use of combined PRP and A2MG + Inc for which only individual components were characterised. Conclusions: PRP may promote repair via growth factors, and A2MG is proposed to modulate immunity and catabolism. Repeated intra-articular injections of PRP with A2MG + Inc caused no systemic or synovial immune response, with no differences between autologous and allogeneic use.
© 2026 The Author(s). Equine Veterinary Journal published by John Wiley & Sons Ltd on behalf of EVJ Ltd.
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Publication Date: 2026-05-24 PubMed ID: 42178493DOI: 10.1002/evj.70196Google Scholar: Lookup
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Summary

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Overview

  • This study characterizes the composition of equine orthobiologic products, specifically platelet-rich plasma (PRP) and alpha-2-macroglobulin (A2MG), and evaluates the immunological safety of repeated intra-articular injections of these products in healthy horses.

Background and Objective

  • Orthobiologics, such as PRP and A2MG, are biological treatments increasingly used to support healing in equine musculoskeletal injuries.
  • The exact composition of these orthobiologics and their safety, especially with repeated allogeneic (from a donor horse) intra-articular (joint) administrations, are not well understood.
  • The study aimed to:
    • Biochemically characterize PRP and A2MG preparations, including effects of incubation at 37°C.
    • Evaluate immunological safety following repeated intra-articular injections of combined PRP and incubated A2MG both autologously (from the same horse) and allogeneically (from another horse) in healthy horses.

Methods

  • The study was performed in two phases:
    1. Biochemical characterization of orthobiologics.
    2. Repeated intra-articular injections and immunological safety evaluation.
  • Blood was taken from horses and processed to produce:
    • Platelet-rich plasma (PRP).
    • Alpha-2-macroglobulin (A2MG), both with and without 1 hour incubation at 37°C (designated PRP, PRP + Inc, A2MG, A2MG + Inc).
  • Characterization techniques included:
    • Haematology analysis for blood cell counts.
    • Biochemical profiling.
    • Enzyme-linked immunosorbent assay (ELISA) to quantify growth factors and proteins.
  • Four healthy horses were administered three intra-articular injections, two weeks apart (Days 0, 14, and 28):
    • Three horses received allogeneic combined PRP and A2MG + Inc.
    • One horse received autologous combined PRP and A2MG + Inc.
  • Samples collected included blood and synovial fluid on Days 0, 14, 28, and 42.
  • Evaluations included:
    • Haematology and biochemistry of blood for immune cell profiles and systemic markers.
    • Flow cytometry analysis for lymphocyte subsets (CD4/CD8 ratio).
    • Serum amyloid A (SAA) as an acute-phase inflammatory marker.
    • Synovial fluid leukocyte counts and immunoglobulin levels.

Results – Biochemical Characterization

  • All orthobiologic preparations effectively removed blood cells, limiting contamination.
  • PRP-based formulations were enriched with platelets and contained higher levels of growth factors known to promote tissue repair.
  • A2MG preparations had higher concentrations of alpha-2-macroglobulin, a protein playing roles in immune modulation and catabolism, but had lower growth factor levels compared to PRP.
  • Incubation at 37°C affected the composition but specific details on protein changes were not fully outlined.

Results – Immunological Safety

  • Repeated intra-articular injection of the combined PRP and A2MG + Inc preparations generated no clinically significant systemic or local immune responses in healthy horses.
  • There were no notable changes in:
    • Systemic or synovial immune cell counts.
    • Lymphocyte subset ratios (CD4/CD8), suggesting no lymphocyte activation or skewing.
    • Acute-phase markers such as serum amyloid A (SAA), indicating no inflammation.
    • Synovial fluid immunoglobulin concentrations remained stable.
  • No differences were observed between autologous and allogeneic administration routes.
  • This suggests that repeated allogeneic PRP combined with A2MG + Inc is immunologically safe in healthy horses.

Conclusions and Limitations

  • The study supports the concept that PRP may contribute to tissue repair through the delivery of growth factors.
  • A2MG is proposed to modulate immune responses and breakdown of proteins (catabolism), complementing PRP’s effects.
  • Repeated injections of combined PRP and incubated A2MG do not elicit immune activation or adverse inflammation in joints.
  • Allogeneic administration appears as safe as autologous administration from an immunological perspective in this small sample.
  • Limitations include:
    • Small number of horses studied (only four total, three allogeneic, one autologous), limiting statistical power.
    • Composition characterization was limited and focused on individual components rather than the combined product.
    • Study was conducted only in healthy animals and may not reflect responses in injured or diseased joints.
  • Further studies with larger sample sizes and expanded characterization may better elucidate the safety and efficacy of these orthobiologics.

Cite This Article

APA
Van Reusel Y, Broeckx SY, Carolo A, Patruno M, Steenbrugge J, Saunders J, Gugliandolo E, Spaas JH. (2026). Composition of allogeneic equine orthobiologics and their repeated intra-articular administrations. Equine Vet J. https://doi.org/10.1002/evj.70196

Publication

Equine veterinary journal
ISSN: 2042-3306
NlmUniqueID: 0173320
Country: United States
Language: English

Researcher Affiliations

Van Reusel, Yanne
  • INTIBIO, Bree, Belgium.
  • Department of Morphology, Imaging, Orthopedics, Rehabilitation and Nutrition, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium.
  • Laboratory of Biochemistry, Department of Veterinary and Biosciences, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium.
Broeckx, Sarah Y
  • INTIBIO, Bree, Belgium.
Carolo, Anna
  • Department of Comparative Biomedicine and Food Science BCA, University of Padova, Padua, Italy.
Patruno, Marco
  • Department of Comparative Biomedicine and Food Science BCA, University of Padova, Padua, Italy.
Steenbrugge, Jonas
  • Laboratory of Biochemistry, Department of Veterinary and Biosciences, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium.
  • Cancer Research Institute Ghent (CRIG)-Veterinary Oncology Network (VON), Ghent, Belgium.
Saunders, Jimmy
  • Department of Morphology, Imaging, Orthopedics, Rehabilitation and Nutrition, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium.
Gugliandolo, Enrico
  • Department of Veterinary Sciences, University of Messina, Messina, Italy.
Spaas, Jan H
  • INTIBIO, Bree, Belgium.
  • Department of Morphology, Imaging, Orthopedics, Rehabilitation and Nutrition, Faculty of Veterinary Medicine, Ghent University, Merelbeke, Belgium.

Grant Funding

  • HBC.2025.0898 / Agentschap Innoveren en Ondernemen
  • INTIBIO

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