Le Varlet B, Ducroc R, Dagonet F B, Pouliquen Y, Vandewalle A, Hirsch M
Laboratoire de Recherches Ophtalmologiques, Unité 86, Institut National de la Santé et de la Recherche Médicale (INSERM), Paris, France.
Invest Ophthalmol Vis Sci. 1995 Nov;36(12):2503-13.
This study describes the effects of the cyclic adenosine monophosphate (cAMP) pathway on the tight junctional barrier of the corneal endothelium, which plays a critical role in maintaining the corneal stroma in an underhydrated, transparent state.
Subcultured bovine corneal endothelial cells grown on filters were used to study the effects of dibutyryl-cAMP and forskolin on transendothelial electrical resistance and [3H]inulin flux. The tight junction-associated protein ZO-1 (zonula occludens protein-1) and F-actin were visualized by indirect immunofluorescence, and the ultrastructural organization of junctional complexes was studied by freeze-fracture electron microscopy.
Cells formed a continuous monolayer of closely apposed hexagonal-type cells separated by a discontinuous belt of tight junctions with a transendothelial electrical resistance of 20.8 +/- 0.6 omega.cm2. Dibutyryl-cAMP (10(-4) M) and forskolin (10(-5) M) increased cell cAMP, significantly decreased the transendothelial resistance by 54% and 43%, respectively, and increased the flux of [3H]inulin from the apical to the basal side of the cells by 56% and 40%, respectively. Both agents also induced condensation of F-actin at the cell borders without any marked changes in the immunostaining of ZO-1 that delineated cell peripheries. However, freeze-fracture studies showed that dibutyryl-cAMP and forskolin induced dispersion of the tight junction network.
These data suggest that activation of the cAMP-dependent pathway, leading to structural changes of the tight junctional network, may modulate the passive fluxes mediated by the paracellular pathway of the corneal endothelial barrier.
本研究描述了环磷酸腺苷（cAMP）途径对角膜内皮紧密连接屏障的影响，该屏障在维持角膜基质处于低水化、透明状态中起关键作用。
使用在滤器上生长的传代培养牛角膜内皮细胞，研究二丁酰 - cAMP和福斯可林对跨内皮电阻和[³H]菊粉通量的影响。通过间接免疫荧光观察紧密连接相关蛋白ZO - 1（闭锁小带蛋白 - 1）和F - 肌动蛋白，并用冷冻蚀刻电子显微镜研究连接复合体的超微结构组织。
细胞形成由紧密排列的六边形细胞组成的连续单层，细胞间由不连续的紧密连接带分隔，跨内皮电阻为20.8±0.6Ω·cm²。二丁酰 - cAMP（10⁻⁴M）和福斯可林（10⁻⁵M）增加细胞内cAMP，分别使跨内皮电阻显著降低54%和43%，并使[³H]菊粉从细胞顶端向基底侧的通量分别增加56%和40%。两种药物还诱导F - 肌动蛋白在细胞边界处凝聚，而勾勒细胞周边的ZO - 1免疫染色无明显变化。然而，冷冻蚀刻研究表明二丁酰 - cAMP和福斯可林诱导紧密连接网络分散。
这些数据表明，cAMP依赖性途径的激活导致紧密连接网络的结构变化，可能调节角膜内皮屏障旁细胞途径介导的被动通量。
