Factors affecting autologous conditioned serum concentration in the horse

Osteoarthritis is the most common cause of lameness in the horse. Autologous conditioned serum (ACS) is a biological therapeutic product that, when administered into a joint, exerts a dose-dependent anti-inflammatory effect. Autologous conditioned serum exerts its anti-inflammatory effect through competitive inhibition of interleukin-1β (IL-1β) by interleukin-1 receptor antagonist (IL-1Ra). Autologous conditioned serum is produced through the incubation of venous whole blood in the presence of chromium sulfate-etched medical grade glass beads at 37°C for 24hrs. Following centrifugation, the conditioned serum is collected and stored in aliquots for future use. The production of ACS is both costly and time consuming and the biological variability in the product can lead to variable clinical effects. Ways to positively influence the concentration of anti-inflammatory cytokines such as IL-1Ra in an ACS preparation are not well documented.
The aims of this thesis were to: (1) compare the cytokine concentrations and microbial contamination present in a non-commercial method of ACS production and a commercial method of ACS production, (2) investigate the effect of freeze/thaw cycles on cytokine concentrations in a commercial method of ACS production, (3) determine the effect and in-vivo safety of intra-articular administration of a non-commercial method of ACS production, (4) examine the influence of exercise on concentrations of cytokines derived from a non-commercial ACS preparation and (5) understand the influence of electroacupuncture on concentrations of cytokines derived from a non-commercial method of ACS production.
The results of the ACS method study determined that a novel non-commercial method of ACS production is equivalent in cytokine profile and in-vitro microbial safety as a previously investigated commercial method of ACS production. Our results suggest that this non-commercial method of ACS production would be both safe and efficacious for in-vivo use. The findings from this study also confirmed that the thermal stability of cytokines reduces with increasing freeze/thaw iteration. A decrease in cytokine concentration was observed with each increase in freeze/thaw cycle, reaching significance at 5 freeze/thaw cycles. These findings have important implications for future use and storage of ACS aliquots.
The results of the short communication on in-vivo administration of a non-commercial ACS product determined that this method of production is safe, resulting in no adverse events. The clinical benefits observed in objective and subjective lameness grade, effusion grade and flexion grade following administration of the non-commercial method of ACS production were variable and due to the low number of subjects, inference into its effectiveness as an intra-articular treatment for osteoarthritis could not be concluded. Further work, with a larger scale clinical trial is needed to validate these safety assumptions and to investigate the efficacy of this non-commercial method when compared to the commercial method of ACS production.
The results of the study exploring the influence of exercise on ACS cytokine profile determined that exhaustive exercise in standardbred racehorses results in a decrease in the concentration of anti-inflammatory cytokines and an increase in the pro-inflammatory cytokine profile of an ACS product 1 hr following exercise, returning to normal resting levels by 24hrs following exercise. Our results suggest that ACS should not be collected within 24hrs of maximal or exhaustive exercise.
The results of the study exploring the influence of electroacupuncture (EA) on cytokine concentrations in an ACS product determined that EA may decrease pro-inflammatory cytokine concentrations and results in a reduced psychological stress response, which was observed as a decrease in IL-10 concentration. The findings from this study have implications for future research into the influence of electroacupuncture on immune function in horses and how that can be utilized to optimize biological products for osteoarthritis treatment in the future.
This thesis provides contributions to the understanding of ways to influence cytokine concentrations in the pre- and post- collection periods, including container type, storage conditions, exercise and electroacupuncture. These findings have made important contributions to the body of knowledge on ACS that have the potential to impact the practices of the veterinary profession within both clinical and research settings.