Whole blood, flow-chamber studies in real-time indicate a biphasic role for thymosin β-4 in platelet adhesion

Abstract

Background Thymosin beta 4 (Tβ 4) is a major actin sequestering peptide present in most mammalian cells. It also acts as an anti-inflammatory agent and promotes corneal wound healing. Methods In the present study, we constructed a four channel cylindrical flow chambers out of polydimethylsiloxane (PDMS) on microscope coverslips. The platelet-binding proteins–fibrinogen and collagen–were immobilized onto the middle ~ 25% of the inner cylindrical surface. The flow method introduced here was employed to determine the effect of Tβ 4, on the deposition of ADP-activated platelets onto fibrinogen cross-linked flow chambers. Results The binding data from the flow chambers indicated that the both the rate constant of platelet deposition (average: 0.026 ± 0.0015 s − 1 , corresponding to a half-life of 26.7 s) and the total number of deposited platelets were independent of the platelet binding protein and the activating agent. Our results show that low concentrations of Tβ 4 (0.2 μM to 0.5 μM) increased both the rate constant of platelet deposition by ~ 1.5-fold (i.e. half-life decreased from 26.7 s to 17.6 s) and the total number of deposited platelets by ~ 3-fold. However at higher concentrations (> 1 μM) the Tβ 4-potentiating effect was diminished to near control levels. Tβ 4 did interact with fibrinogen with an estimated K D of ~ 126 ± 18 nM or 66 ± 20 nM under equilibrium or flow, respectively. Conclusion These results suggest that Tβ 4 could potentially increase the affinity of platelet receptors for their ligands thus promoting platelet deposition. Tβ 4 could also bind to fibrinogen and as its concentration increased would prevent platelet–fibrinogen interactions resulting in the attenuation of platelet deposition. General significance This work suggests that Tβ 4 might have a dual role in platelet function.