Thrombolytic activity of recombinant tissue-type plasminogen activator (rtPA) in in-vitro model for subretinal hemorrhages.

Abstract

Purpose: Recombinant tissue-type plasminogen activator (rtPA) has become central to the treatment of subretinal hemorrhages. However, individual results show highly variable and unpredictable efficacy of thrombolytic function. The aim of this study is to investigate the thrombolytic activity of rtPA in an in-vitro model for submacular hemorrhages.

Methods: Using UV/Vis spectroscopy, the thrombolytic activity of rtPA was analyzed in an in-vitro model for subretinal hemorrhages with citrated human venous blood from 24 volunteers. The effects of blood clot volume (20 µL and 100 µL) and age (1d and 4d), rtPA concentration (0, 10, 50, 100, 150, and 200 µg/mL), and incubation time (30 min, 1d, and 2d) on thrombolysis was analyzed on blood clots prepared through different synthesis routes (tube vs. hanging droplet).

Results: For each period of incubation with rtPA, thrombolysis was significantly reduced for large blood clots with a volume of 100 μL compared with 20 μL blood clots. We found significantly more thrombolysis in 4-day-old clots than in 1-day-old clots after 30 min incubation with rtPA for each blood clot size studied. For each blood clot size and age as well as rtPA incubation time, thrombolysis did not increase linearly across the tested concentrations. Thrombolysis was significantly lower after 30 min than after 1d and 2d rtPA incubations.

Conclusion: The effectiveness of thrombolysis with rtPA increases with smaller clot size (20 µL), increasing clot age (4d), and increasing incubation time (1-2d). Within the range of biocompatible rtPA concentrations, there was no concentration effect of rtPA on thrombolysis.