Assessment of Canine Autologous conditioned PlasmaTM cellular and transforming growth factor-β1 content

Abstract

Objectives: To evaluate (1) the cellular composition of canine ACP™ including using two different preparation protocols with variations on centrifugation time, (2) the effect of different activation protocols on the transforming growth factor (TGF)-β1 content in the ACP, and (3) patient factors that might influence platelet concentration of the ACP. Methods: ACP was made with blood from 15 dogs using a manufacturer-recommended protocol[1]. Each ACP sample was divided into three aliquots that were activated with calcium chloride (CaCl2), human γ-thrombin (HGT), or not activated. TGF-β1 was quantified in each aliquot using an ELISA and comparisons among activation protocols were performed using a Skillings-Mack test. Correlations between platelet and TGF-β1 concentration were assessed with a Pearson correlation coefficient. ACP was subsequently prepared from an additional 17 dogs using a slightly modified centrifugation protocol and cellular composition was assessed. Effects of dog age, body weight, and hematocrit were assessed for their potential impact on ACP platelet concentration using a multiple linear regression analysis. Results: The mean increase in platelet concentration in the ACP above that in the whole blood was 1.2× (±std 0.62) and leukocyte concentration was a mean of 26% (0.37) that in the whole blood using the standard protocol. There was a significant (p < 0.01) effect of activation on TGF-β1 concentrations with mean concentrations of 4,538 (2,317), 14,948 (13,784), and 14,096 (15,210) pg/ml in aliquots that were not activated or were activated with thrombin or CaCl2 respectively. There were significant correlations between the platelet concentration and TGF-β1 concentration in aliquots that were activated with either thrombin (r = 0.66; p < 0.01) or CaCl2 (r = 0.86, p < 0.0001). The mean increase in platelet concentration was 1.4× (0.62) and the leukocyte concentration was 0.28× (0.13) that in whole blood using the modified ACP preparation protocol. Dog age, body weight, and hematocrit were not significant predictors of ACP platelet concentration. Conclusions: These data show that on average this preparation protocol produces a mildly platelet-concentrated, leukoreduced platelet-rich plasma. Intentional activation had a significant effect on TGF-β1 concentrations with use of both CaCl2 and thrombin resulting in higher TGF-β1 concentrations than that obtained in samples that are not activated.