Joint injuries are overwhelmingly common in both human and equine athletes. Chondrocytes, the sole cell type in cartilage, are responsible for producing and maintaining the extra-cellular matrix (ECM), which affords remarkable tensile and compressive strength to the joint surface. Once damaged, cartilage has little to no ability to heal itself. Therefore, posttraumatic osteoarthritis (PTOA) commonly develops following sufficient joint trauma, whether sustained during an acute injury or accumulated overtime.

Unfortunately, PTOA is a progressive, debilitating disease that currently lacks any effective treatment. Pain and decreased mobility are addressed with systemic non-steroidal anti-inflammatory drugs (NSAIDs), intra-articular corticosteroids, rest and rehabilitation. The end stage therapy for OA joints in humans is total joint replacement, while select joints in the horse can be treated with surgical arthrodesis. Both of these procedures are invasive and expensive interventions.

Our lab is focused on understanding the pathogenesis of PTOA and developing regenerative medicine therapies to help regenerate cartilage and prevent the development of PTOA following joint injury. Specifically, we are interested in gene and cell therapies targeted at reducing post-traumatic inflammation and improving repair of the articular surface, thereby preventing the development of PTOA in the first place. Dr. Ortved’s clinical work as an equine orthopedic surgeon facilitates translation of this research from bench to clinic floor.

Research Techniques

• Cell culture of equine mesenchymal stem cells
• Molecular biology: PCR and qRT-PCR, flow cytometry, protein analysis
• Microscopy
• Histology

Stem cell therapy

Cell-based therapies aim to return damaged or injured tissue to a more normal structure and function. An overarching goal in cell-based repair is to restore the articular surface, thereby preventing further joint degradation. Although bone marrow-derived MSCs (BM-MSCs) have been the most frequently used MSC type for cartilage repair to date, full chondrogenic differentiation has been disappointing. Therefore, we are currently investigating the chondrogenic differentiation capability of synovial membrane-derived mesenchymal stem cells (SD-MSCs). Recent experiments using flow cytometry in the Ortved laboratory have demonstrated a similar immunophenotype between BM-MSCs and SD-MSCs, with SD-MSCs expressing the appropriate markers of stemness. We have also shown that SD-MSCs have increased proliferative capacity in vitro, making timely culture expansion of these cells achievable. The ultimate goal of this research is defining a source of adult MSCs suitable for resurfacing cartilage lesions to facilitate healing.

Caption: Bone marrow is harvested from the sternum of a sedated horse and culture-expanded in the lab.  We are using these cells to form cartilage- like pellets through chondrogenic differentiation, in order to optimize a cell-source that could be used to repair cartilage defects in the joint.

Gene Therapy

Gene therapy also has the potential to bolster the weak healing response in articular cartilage. Dr. Ortved has demonstrated improved cartilage repair in large, full-thickness chondral defects created in the lateral trochlear ridge of the equine femur using autologous chondrocytes transduced ex vivo with an adeno-associated virus (AAV) vector overexpressing the anabolic protein IGF-I.

Caption: Arthroscopic images of empty (a), implanted (b) and healing defects 8 weeks postimplantation (c-f). (c) Defects repaired with rAAV5-IGF-I transduced chondrocytes had better defect fill and white “cartilage- like” tissue compared to the defects filled with (d) rAAV5-GFP transduced chondrocytes, (e) untransduced chondrocytes, or (f) fibrin alone.

Dr. Ortved has also demonstrated that AAV vectors can be used to safely transduce equine articular cells in vivo with efficient, sustained expression of a therapeutic transgene. More recently, Dr. Ortved has been investigating AAV-mediated overexpression of interleukin-10 (IL-10), an immunomodulatory cytokine. She showed that L-10 overexpression downregulates expression of proinflammatory mediators in inflamed chondrocytes in vitro. We are now beginning to assess the effects of this therapy in vivo to determine if it can mitigate the post- traumatic inflammatory response that occurs following joint injury.

Caption: Equine chondrocytes have been transduced with an AAV vector expressing red fluorescent protein.

Post-traumatic Osteoarthritis (PTOA)

We are investigating changes in gene expression in cartilage, synovium, fat pad and meniscus in horses with PTOA, in order to understand how different pathways are affected by trauma. We are also developing an ex vivo model of PTOA to help understand the histologic, microscopic and molecular changes that occur post-traumatically.

1. Moss KL, Jiang Z, Linardi RL, Gale AL, Dodson ME, Grzybowski C, Haughan J, Robinson M, Ortved KF. Sustained interleukin-10 transgene expression following intraarticular AAV5-IL-10 administration to horses. Human Gene Therapy, 31:110-118, 2020.
2. Chapman HS,  Gale AL, Dodson ME, Linardi RL, Ortved KF. The effect of autologous platelet lysate on the differentiation potential of equine bone marrow-derived mesenchymal stem cells. Stem Cells Dev, 29:144-155, 2020.
3. Brown KA, Davidson EJ, Johnson A, Wulster KB, Ortved KF. Assessment of intra-synovial inflammatory cytokines in horses with standing cone bean computed tomographic evidence of cervical articular process joint osteoarthritis. EVJ, 2020.
4. Haughan J, Jiang Z, Stefanovski D, Moss KL, Ortved KF, Robinson MA. Detection of intra-articular gene therapy in horses using quantitative real time PCR in synovial fluid and plasma. Drug Testing and Analysis, 2020.
5. Doering AK, Reesink HR, Leudke LK, Moore C, Nixon AJ, Fortier LA, Ducharme NG, Ross MW, Richardson DW, Ortved KF. Return to racing after internal fixation of carpal slab fractures in Thoroughbred and Standardbred racehorses. Veterinary Surgery, 2019.
6. Linardi RL, Dodson ME, Moss KL, King WJ. Ortved KF. The effect of autologous protein solution on the inflammatory cascade in stimulated equine chondrocytes. Frontiers Vet Regen Med, 6:64, 2019.
7. Gale AL, Mammone RM, Linardi RL, McClung G, Ortved KF.  Comparison of the chondrogenic differentiation potential of equine synovial membrane-derived and bone marrow-derived mesenchymal stem cells. Frontiers Vet Regen Med, 6:2019.
8. Gale AL, Linardi RL, Dodson ME, Ortved KF.  The effect of hypoxia on chondrogenesis of equine synovial membrane-derived and bone marrow-derived mesenchymal stem cells. BMC Vet Research, 15:201, 2019.
9. Chapman H, Richardson DW, Ortved KF. Arthrodesis of the metacarpophalangeal and metatarsophalangeal joint to treat osteoarthritis in 17 horses. Veterinary Surgery, 48:850-857, 2019.
10. Ortved KF, Begum L, Stefanovski D, Nixon AJ. AAV-mediated overexpression of IL-10 mitigates the inflammatory cascade in stimulated chondrocyte pellets. Current Gene Therapy, 18:171-179, 2018.
11. Ortved KF, Goodale MB, Ober C, Maylin GA, Fortier LA. Plasma firocoxib concentrations after intra-articular injection of autologous conditioned serum prepared from firocoxib positive horses. Vet J, 2017.
12. Griffin  D, Ortved KF, Nixon AJ, Bonassar L. Mechanical properties and structure- function relationships in articular cartilage repair using IGF-I gene therapy. Journal of Orthopedic Research, 34:149-153, 2016.
13. Ortved KF, Austin BS,  Scimeca MS, Nixon  AJ. RNA interference mediated interleukin-1β silencing in inflamed chondrocytes decreases target and downstream catabolic responses. Arthritis 2016: 3484961, 2016.
14. Ortved KF, Nixon AJ. Cell-based cartilage repair strategies for horses. Vet J 208:1-12, 2016.
15. Ortved KF, Begum L, Mohammed HO, Nixon AJ. Implantation of rAAV5-IGF-I transduced autologous chondrocytes improves cartilage repair in full-thickness defects in the equine model. Molecular Therapy, 23:363-373, 2015.
16. Ortved KF, Wagner B, Calcedo R, Wilson JM, Schaefer DM, Nixon AJ. Humoral and cell-mediated immune response, and growth factor synthesis after direct intra-articular injection of rAAV2-IGF-I or rAAV5-IGF-I in the equine middle carpal joint. Human Gene Therapy, 26:161-171, 2015.
17. Ortved KF, Nixon AJ, Mohammed HO, Fortier LA. Treatment of subchondral bone cysts of the medial femoral condyle in mature horses with growth factor enhanced chondrocyte grafts: A retrospective study of 49 cases. Equine Vet J 44:606-613, 2012.