Abstract
Autologous conditioned serum (ACS) is a common intra-articular treatment for osteoarthritis in horses. The objective of this study was to investigate the influence of ACS preparation method on product contamination and concentrations of relevant cytokines and the influence of multiple freeze/thaw cycles. Blood was obtained from 10 healthy Thoroughbred horses and processed in parallel using a commercial and a non-commercial method to obtain ACS. Fluorescent microsphere immunoassay (FMIA) analysis was performed to quantify Interleukin 1 receptor antagonist (IL-1Ra), Interleukin-10 (IL-10), Interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α) concentrations in ACS obtained by both production methods. Effect of 3, 4 and 5 freeze/thaw cycles on concentrations of IL-1Ra, IL-10, IL-1β and TNF-α were assessed against baseline samples (2 cycles) in commercial ACS products. Standard aerobic and anaerobic culture methods were applied to both ACS products. Mixed effect one-way analyses of variance (ANOVA) were used to compare the two ACS production method for each cytokine. Repeated measures, mixed effect ANOVA were used to assess the effect of freeze/thaw on cytokine concentrations. Significance was set at P < 0.05. There was no difference in cytokine concentration between production methods (IL-1Ra P = 0.067, IL-1β P = 0.752, IL-10 P = 0.211 and TNF-α P = 0.25). Microbial growth was only observed in two samples obtained using the commercial production method. When compared to baseline, IL-1Ra concentration was decreased following the 5th freeze/thaw cycle (P < 0.001). These results suggest that the concentration of important cytokines are not influenced by ACS production method. When storing ACS samples for future use, freeze/thaw cycles associated with standard clinical practice are unlikely to influence cytokine concentrations. However, the lack of outcome measures associated with 1 or 2 freeze/thaw cycles represents a limitation of this study.
Highlights
Osteoarthritis contributes to ∼60% of all lameness conditions in the horse [1]
Method of Autologous conditioned serum (ACS) production was not associated with a difference in Interleukin 1 receptor antagonist (IL-1Ra); IL-1 or IL-10 concentrations (P = 0.067, P = 0.752, P = 0.211, respectively)
Immunoglobulin binding to monocytes has been shown to activate monocytes and stimulate the production of IL-1Ra alone compared to lipopolysaccharide activation which has been reported to induce both IL-1β and IL-1Ra production; suggesting that monocyte activation is dependent upon external factors [17]
Summary
Osteoarthritis contributes to ∼60% of all lameness conditions in the horse [1] This condition has considerable economic impact on racing and performance horse industries [2].While commonly used for the treatment of osteoarthritis, intra-articular corticosteroid administration is associated with an increased risk of musculoskeletal injury [3]. Tumor necrosis factor α (TNF-α) and Interleukin 1β (IL-1β) are potent activators of intra-articular inflammatory pathways and synovial concentrations correlate with joint disease in humans and horses [5,6,7]. In vivo administration of Interleukin 1 receptor antagonist (IL-1Ra), exerts a dose dependent anti-inflammatory and chondroprotective effect attributable to competitive receptor inhibition in humans and horses [9,10,11,12]. A commercial preparation kit for ACS is widely available to equine practitioners for intra-articular treatment of osteoarthritis and post arthroscopic interventions [11]
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