Abstract 13104: Small Molecules Prevent Thrombin Induced Rho-Rock-MLC Mediated Hyperpermeability in Endothelial Cells

Abstract

Introduction: Thrombin, a serine protease plays an important role in a wide array of a normal and pathological process, including infection, inflammation and tissue injury. There are limited numbers of molecules that prevent vascular inflammation and injury during ALI or ARDS. Our novel combination therapy with epigenetic modifiers that could target the genes epigenetically to reduce thrombin induced hyperpermeability and injury. Here, we use 5-Aza 2-deoxycytidine (Aza) and Trichostatin A (TSA) together to see the restoration of throbin-induced hyperpermeability. Hypothesis: We hypothesize that treatment with the combination Aza and TSA (Aza+TSA) will restore the thrombin induced vascular hyper permeability and increases vascular integrity. Methods and Results: In thrombin induced vascular hyperpermeability, the epigenetic modifiers Aza or TSA alone cannot be able to circumvent the inflammation and restores the cell integrity. However, the Aza+TSA treatment showed a significant level of decrease in vascular hyperpermeability and promoted survival rate. Our Western analysis data suggest that the combination of Aza+TSA treatment suppressed phosphorylation of MLC and ROCK and also Rho1. ROCK-mediated phosphorylation of MLC results in increased actin-filament assembly and actomyosin contraction, which is evident to stress fiber’s formation (Fig. 1A). This was further supported by the immunofluorescence analysis (Fig. 1B). Conclusions: Our data shows that Aza+TSA prevents thrombin induced endothelial hyperpermeability through the inhibition of Rho-ROCK-MLC pathway. Thus Aza+TSA has a substantial therapeutic value during infection, inflammation and acute lung injury.