There is a current view that myosin light chain kinase (MLCK) plays a critical role in endothelial permeability. To investigate the functions of MLCK in endothelial cells in vivo, we generated a mouse model in which MLCK was selectively deleted by crossing Mylk1 floxed mice with Tie2 ⁄ cre transgenic mice. Knocking out Mylk1 from endothelial cells had no effect on the global phenotype of the mice, including body weight and blood pressure. Lipopolysaccharide (LPS)-mediated septic death was also not altered in the knockout (KO) mice. Consistently, LPS-induced inflammatory injury and the increase in microvascular permeability in the main organs, including the lung and the kidney, was not significantly attenuated in KO mice as compared with wild-type mice. However, the LPS-induced microvascular hyperpermeability of the esophagus and the eyeballs was attenuated in KO mice. We also found that the LPS-mediated increase in the number of caveolae in the endothelial cells of the esophagus was significantly reduced in KO mice. Our results do not support a role for endothelial cell MLCK in the pathogenesis of inflammatory diseases.
IntroductionThe vascular endothelium forms a semipermeable barrier between the bloodstream and interstitial space [1,2]. The intact endothelial barrier is restrictive to cells, proteins, fluid, and solutes, thus maintaining vascular homeostasis and the physiological functions of different organs [3]. In the development of a variety of inflammatory diseases, such as sepsis and atherosclerosis, the endothelium is disturbed, resulting in microvascular hyperpermeability [4]. Fluid, molecules and cells move across the endothelium via a transcellular or a paracellular pathway. Small molecules are transported through a structural junction between adjacent endothelial cells (ECs) by a paracellular pathway. Large molecules, e.g. albumin, are transported across the Abbreviations EBD, Evans blue dye; EC, endothelial cell; H&E, hematoxylin and eosin; IF, immunofluorescence; KO, knockout; L-MLCK, long form of myosin light chain kinase; LPS, lipopolysaccharide; MLC, myosin light chain; MLCK, myosin light chain kinase; S-MLCK, short form of myosin light chain kinase; WT, wild-type. Inflammatory mediators, such as lipopolysaccharide (LPS), cytokines, and thrombin, are capable of binding to their receptors on ECs, leading to disruption of the junctions, the formation of gaps between adjacent cells, and, finally, an increase in endothelial permeability and endothelial dysfunction [6]. During this process, a change in cell shape caused by actomyosin contraction is speculated to be the primary determinant of increased endothelial permeability [7]. Actomyosin contraction is primarily mediated by myosin light chain (MLC) phosphorylation, which activates myosin, leading to cross-bridge movement [8,9]. MLC kinase (MLCK) is a dedicated kinase for MLC phosphorylation, and is expressed in ECs [10,11]. MLCK is considered to be important for the regulation of endothelial permeability in response to LPS [12,13]. Therefore, it...
