Critical Role of Non-Muscle Myosin Light Chain Kinase in Thrombin-Induced Endothelial Cell Inflammation and Lung PMN Infiltration

Fabeha Fazal,Kaiser M Bijli,Khandaker N Anwar,Matthew Murrill,Jacob N Finkelstein,D Martin Watterson,Arshad Rahman,Mohammad Minhajuddin,Antony Leonard,Jian Fu

doi:10.1371/journal.pone.0059965

Abstract

The pathogenesis of acute lung injury (ALI) involves bidirectional cooperation and close interaction between inflammatory and coagulation pathways. A key molecule linking coagulation and inflammation is the procoagulant thrombin, a serine protease whose concentration is elevated in plasma and lavage fluids of patients with ALI and acute respiratory distress syndrome (ARDS). However, little is known about the mechanism by which thrombin contributes to lung inflammatory response. In this study, we developed a new mouse model that permits investigation of lung inflammation associated with intravascular coagulation. Using this mouse model and in vitro approaches, we addressed the role of non-muscle myosin light chain kinase (nmMLCK) in thrombin-induced endothelial cell (EC) inflammation and lung neutrophil (PMN) infiltration. Our in vitro experiments revealed a key role of nmMLCK in ICAM-1 expression by its ability to control nuclear translocation and transcriptional capacity of RelA/p65 in EC. When subjected to intraperitoneal thrombin challenge, wild type mice showed a marked increase in lung PMN infiltration via expression of ICAM-1. However, these responses were markedly attenuated in mice deficient in nmMLCK. These results provide mechanistic insight into lung inflammatory response associated with intravascular coagulation and identify nmMLCK as a critical target for modulation of lung inflammation.

Highlights

A hallmark of acute lung injury (ALI) and its more severe form acute respiratory distress syndrome (ARDS) is an exuberant inflammatory response characterized by massive infiltration of polymorphonuclear lymphocytes (PMN) into the lung that leads to disruption of capillary-alveolar barriers and development of pulmonary edema with severe consequences for pulmonary gas exchange [1,2]
Using integrated in vitro and in vivo approaches, we show an important role of non-muscle myosin light chain kinase (nmMLCK) in causing endothelial cell (EC) inflammation by its ability to mediate nuclear translocation and transcriptional capacity of RelA/p65, and PMN infiltration in the lungs of mice challenged with thrombin
To determine if the above effects of nmMLCK inhibition can be recapitulated in vivo in the intact lungs of mice, we developed an in vivo mouse model of lung PMN sequestration associated with intravascular coagulation

Summary

Introduction

A hallmark of acute lung injury (ALI) and its more severe form acute respiratory distress syndrome (ARDS) is an exuberant inflammatory response characterized by massive infiltration of polymorphonuclear lymphocytes (PMN) into the lung that leads to disruption of capillary-alveolar barriers and development of pulmonary edema with severe consequences for pulmonary gas exchange [1,2]. An emerging paradigm is that uncontrolled activation of the coagulation cascade after inflammation and tissue injury plays an important role in the pathogenesis of ALI/ARDS [3,4,5,6]. Studies in animal models have shown that infusion of thrombin induces lung vascular injury and tissue edema and that these responses are critically dependent on PMN sequestration in microvessels [12,13]. The mechanism by which thrombin promotes lung PMN sequestration remains poorly understood, largely due to the lack of a mouse model of thrombininduced lung PMN infiltration

Methods

Results

Conclusion