Hale Josephine, Hughes Kristopher, Hall Sarah, Labens Raphael
School of Agricultural, Environmental and Veterinary Sciences, Charles Sturt University, Wagga Wagga, NSW, Australia.
Animal & Veterinary Sciences Department, Scotland's Rural College, Edinburgh, United Kingdom.
Front Vet Sci. 2021 Nov 25;8:759828. doi: 10.3389/fvets.2021.759828. eCollection 2021.
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 < 0.05. There was no difference in cytokine concentration between production methods (IL-1Ra = 0.067, IL-1β = 0.752, IL-10 = 0.211 and TNF-α = 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 ( < 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.
自体条件血清（ACS）是马匹骨关节炎常见的关节内治疗方法。本研究的目的是调查ACS制备方法对产品污染及相关细胞因子浓度的影响，以及多次冻融循环的影响。从10匹健康纯种马采集血液，并行采用商业方法和非商业方法进行处理以获得ACS。进行荧光微球免疫分析（FMIA），以量化两种生产方法获得的ACS中白细胞介素1受体拮抗剂（IL-1Ra）、白细胞介素-10（IL-10）、白细胞介素-1β（IL-1β）和肿瘤坏死因子-α（TNF-α）的浓度。针对商业ACS产品中的基线样本（2次循环），评估3次、4次和5次冻融循环对IL-1Ra、IL-10、IL-1β和TNF-α浓度的影响。对两种ACS产品均采用标准需氧和厌氧培养方法。采用混合效应单因素方差分析（ANOVA）比较每种细胞因子的两种ACS生产方法。采用重复测量、混合效应ANOVA评估冻融对细胞因子浓度的影响。显著性设定为<0.05。生产方法之间的细胞因子浓度无差异（IL-1Ra = 0.067，IL-1β = 0.752，IL-10 = 0.211，TNF-α = 0.25）。仅在使用商业生产方法获得的两个样本中观察到微生物生长。与基线相比，第5次冻融循环后IL-1Ra浓度降低（<0.001）。这些结果表明，重要细胞因子的浓度不受ACS生产方法的影响。在储存ACS样本以供未来使用时，与标准临床实践相关的冻融循环不太可能影响细胞因子浓度。然而，缺乏与1次或2次冻融循环相关的结果测量是本研究的一个局限性。
