The Journal of ExtraCorporeal Technology No 56-2 | Page 15

42 M . Sancheti et al .: J Extra Corpor Technol 2024 , 56 , 37 – 44

assessment of the efficiency of commercially available biocoated bypass circuits in blocking the CPB-mediated platelet activation in a surgical setting .

The main objective of this study was to establish a standardized method to study platelet activation in vitro for future comparative studies , and to further determine if there are any significant inter-variations across the circuit batches that would potentially impact the accuracy of not only the biomarker measurements across technical and biological replicates , but also the reliability of our statistical analyses . Moreover , the data collected in the current pilot study will allow for appropriate study sample size calculation that provides enough power for more reliable and reproducible comparative future studies across different types of biocoated CPB circuits .

In this study , we measured platelet counts and the secretion and translocation levels of known and well-established biomarkers of platelet activation including PF4 , b-TG , GPIIb / IIIa , and soluble P-selectin at different time points of CBP . Together , these biomarkers provide a comprehensive profile of platelet activation , aggregation , adhesion , and subsequent coagulation cascade that is triggered by the contact of platelets with foreign surfaces such as the lumen of the CPB circuits .

PF4 is a cytokine released from alpha granules of platelets during activation and blood coagulation , and its increased level was found to be consistent with the incidence of cardiac complications during bypass surgeries [ 3 ]. In addition , PF4 was shown to be elevated in peripheral artery disease , acute ischemic stroke , essential thrombocytopenia , and other clinical conditions that were associated with platelet activation and is thus considered a relatively more sensitive marker of platelet activation during pathological events [ 14 ]. Xcoating TM showed a decrease in PF4 total amount throughout the trial , however , when normalized with platelet count , there was an increase then decrease in PF4 . Although there was an increase in PF4 normalized concentration , it remained unchanged at the end of the trial . Trillium Ò Biosurface showed a constant PF4 concentration indicating that no further activation occurred to increase PF4 .

In addition to PF4 , b-TG is another cytokine released early following platelet adhesion and is commonly used as a reliable marker of platelet activation in vitro [ 14 ]. After the initial increase in b-TG concentration , Trillium Ò Biosurface and Xcoating TM did not have significant effects throughout the experiments . Another platelet marker GPIIb / IIIa belongs to the large family of integrin complexes and plays a critical role in platelet aggregation and increased binding to plasma fibrinogen and endothelial von Willebrand factor , therefore , facilitating thrombin generation and blocking hemorrhage [ 15 ]. Furthermore , GPIIb / IIIa surface translocation increases upon platelet activation making it a valuable biomarker in quantifying platelet activation and aggregation [ 16 ]. Both circuit coatings prevented further GPIIb / IIIa activation indicated by a decrease in concentration during the trials . Another platelet activation marker P-selectin is stored in alpha granules of platelets as well as in the Weibel-bodies of endothelial cells that cover the lumen of the blood vessels . Upon activation , P-selectin is translocated to the surface of endothelium and released into the blood circulation and plays an important role during the initial adhesion and rolling of platelets in the site of injury [ 17 , 18 ]. In addition , activated platelets result in an increase in soluble P-selectin translocation and release allowing for quantification of platelet activation [ 19 ]. The inhibition of P-selectin release was displayed by a decrease in concentration over the course of the trial in Trillium Ò Biosurface . When normalized over the platelet counts , soluble P-selectin levels did not show any significant changes throughout the experiment using Xcoating TM circuits .

This was a longitudinal study that measured the platelet activation at different time points during a circuit run for two different types of CPB circuit coatings . The baseline measurement was arbitrarily set at 5 min , so there would be adequate time for the crystalloid to mix with the bovine blood within the circuit . Measurements done at 30 min and 55 min were compared to 5 min and shown to both fall within ± 2 standard deviations of the mean at each time point for each type of coated circuit . ELISA assays were then utilized to measure platelet activation markers due to their high sensitivity and specificity . While these assays are not intended to be used in a clinical setting for patients undergoing CPB , they can provide insight into the activity of platelets within the CPB coating during cardiac surgery .

It is noteworthy , that although this study has shown minimum inter-variations within each type of coating , it cannot provide conclusive comparative data of any potential differences between the coating types . The present study was designed to establish a standardized protocol in a laboratory setting that would allow us to plan future experiments to compare different types of bio-coated circuits in the market and address a very critical question : which type of commercially available CPB circuit would provide better protection against platelet activation during human bypass surgeries ? In addition , we were not able to have a positive control group for this study , as uncoated reservoirs or tubing are not available from the same vendor for clinical use . The use of custom-made uncoated CPB circuit would have added confounding variables to the study . Our established protocol remains accurate shown by the similarities in biomarker activation in two different circuit coatings and the precision , consistency , and duplicability of the ELISA results .