PURPOSE: To investigate changes in the phosphorylation of myosin light chain (MLC) in response to histamine and its effect on the barrier integrity of corneal epithelial cells. MATERIALS AND METHODS: Experiments were performed in bovine corneal epithelial cells (BCEC). RT-PCR and Western blotting were employed to characterize expression of H1 receptors and MLC kinase (MLCK). Phosphorylation of MLC was assessed by urea-glycerol gel electrophoresis and Western blotting. Barrier integrity was determined as permeability to horseradish peroxidase (HRP; 44 kDa) across monolayers grown on porous filters. RESULTS: Expression of both H1 receptors and MLCK was found in BCEC. Exposure to histamine induced significant MLC phosphorylation concomitant with an increase in HRP permeability. In addition, organization of the cortical actin found in resting cells was disrupted. In contrast to histamine, ATP (a P2Y receptor agonist) induced dephosphorylation of MLC. Pre-exposure to ATP reduced the effect of histamine on HRP permeability and disruption of cortical actin. CONCLUSION: MLC phosphorylation, a biochemical pre-requisite for increased contractility of the actin cytoskeleton, led to histamine-induced breakdown of the barrier integrity in the corneal epithelial cells. This is attributed to weakening of the tethering forces at the tight junctions by the centripetal forces produced by increased actin contractility.
PURPOSE: To investigate changes in the phosphorylation of myosin light chain (MLC) in response to histamine and its effect on the barrier integrity of corneal epithelial cells. MATERIALS AND METHODS: Experiments were performed in bovine corneal epithelial cells (BCEC). RT-PCR and Western blotting were employed to characterize expression of H1 receptors and MLC kinase (MLCK). Phosphorylation of MLC was assessed by urea-glycerol gel electrophoresis and Western blotting. Barrier integrity was determined as permeability to horseradish peroxidase (HRP; 44 kDa) across monolayers grown on porous filters. RESULTS: Expression of both H1 receptors and MLCK was found in BCEC. Exposure to histamine induced significant MLC phosphorylation concomitant with an increase in HRP permeability. In addition, organization of the cortical actin found in resting cells was disrupted. In contrast to histamine, ATP (a P2Y receptor agonist) induced dephosphorylation of MLC. Pre-exposure to ATP reduced the effect of histamine on HRP permeability and disruption of cortical actin. CONCLUSION: MLC phosphorylation, a biochemical pre-requisite for increased contractility of the actin cytoskeleton, led to histamine-induced breakdown of the barrier integrity in the corneal epithelial cells. This is attributed to weakening of the tethering forces at the tight junctions by the centripetal forces produced by increased actin contractility.
Authors: A Kittel; E Kaczmarek; J Sevigny; K Lengyel; E Csizmadia; S C Robson Journal: Biochem Biophys Res Commun Date: 1999-09-07 Impact factor: 3.575
Authors: T Hashikawa; M Takedachi; M Terakura; T Saho; S Yamada; L F Thompson; Y Shimabukuro; S Murakami Journal: J Dent Res Date: 2003-11 Impact factor: 6.116
Authors: Claudia N Emami; Mikael Petrosyan; Stefano Giuliani; Monica Williams; Catherine Hunter; Nemani V Prasadarao; Henri R Ford Journal: Surg Infect (Larchmt) Date: 2009-10 Impact factor: 2.150
Authors: Mikael Petrosyan; Yigit S Guner; Monica Williams; Anatoly Grishin; Henri R Ford Journal: Pediatr Surg Int Date: 2009-03-20 Impact factor: 1.827