Development of an equine carpal groove model to study early changes in osteoarthritis - a pilot study

Abstract

Purpose: The cartilage groove model has been shown to be an effective model for investigation of early degenerative joint changes in various animal models, including recently in the equine metacarpophalangeal (MCP) joint. In this pilot study, we investigated: 1) if the groove model induces comparable cartilage degeneration in the equine middle carpal as in the MCP joint and 2) if a modified method for disrupting the collagen network without tissue loss, in combination with loading, also leads to cartilage degeneration. Methods: In two adult Shetland ponies, the dorsal aspect of the condyles of the third metacarpal bone (MCP joint) and the proximal surface of the radial facet of the third carpal bone (middle carpal joint) were grooved as described in the literature, using a hooked arthroscopic probe with a sharpened tip (“blunt grooves”). In the contra-lateral limb, grooves were made with a surgical mini blade that was clamped in a custom-made device (“sharp grooves”). After two weeks of box rest, the ponies were subjected to an eight-week training protocol on a treadmill. Clinical lameness was evaluated weekly. Synovial fluid was collected from the middle carpal joint before surgery and at two and 10 weeks postoperatively for analysis of glycosaminoglycan (GAG) levels and matrix metalloproteinase (MMP) activity. At 10 weeks, ponies were euthanized and osteochondral samples harvested for macroscopic, histologic and gene expression analysis of matrix components. Additionally, “fresh” grooves were made in the MCP and middle carpal joints of cadaver limbs to compare the microscopic appearance of grooves with and without a period of in vivo loading. Results: After 6-8 weeks post-operatively, regular stumbling and mild lameness were observed in both ponies during training. Macroscopically, grooves made by both techniques could be identified and adjacent tissue was affected, mainly in the carpal joints. Safranin-O stained histology sections clearly showed degenerative changes for cartilage with blunt grooves (i.e., loss of staining, fibrillation and cell clustering around the lesions), whereas only very mild changes were observed in cartilage with sharp grooves (Fig.1). Microscopy scores in both blunt and sharp grooved joints were higher at 10 weeks (12.4 and 7.3, respectively) compared with grooved cadaver joints (5.5 and 3.0) according to the OARSI equine microscopic grading system. Synovial fluid GAG concentrations dropped dramatically at two weeks after surgery, whereas MMP activity showed a clear increase. However, at 10 weeks, both markers returned to approximately baseline levels (Fig. 2). Carpal cartilage showed an upregulation of collagen 2, aggrecan and COMP, particularly where blunt grooves were made (Fig. 3). Conclusions: The groove model can be used in the equine carpus for the induction of cartilage degeneration. Contrary to the current paradigm, only disruption of the collagen network seemed not to lead to cartilage degeneration within 10 weeks after surgery. This would imply a crucial role for tissue loss. Synovial fluid GAG concentration and MMP activity do not seem to be sensitive markers once the initial inflammatory reaction has disappeared. A more extensive “omics-approach” could provide a better reflection of subtle changes in joint homeostasis in this early stage of degenerative joint changes. Confirmation in a larger number of animals is necessary, as is research on the long-term fate of the sharp lesions.