TNF-α disrupts morphologic and functional barrier properties of polarized retinal pigment epithelium

Retinal pigment epithelial (RPE) cells form a blood–ocular barrier, and their polarized property is crucial for maintaining the barrier functions. Tumor necrosis factor alpha (TNF-α), a major pleotropic inflammatory cytokine that disrupts the barrier function and eventual angiogenesis, is expressed in the choroidal neovascularizations of age-related macular degeneration eyes. Thus, it most likely plays an important role in the progression of the disease. The purpose of this study was to compare the effects of TNF-α on the barrier function of polarized RPE cells. Non-polarized RPE cells were used as negative controls. Isolated porcine RPE cells were seeded on Transwell™ membranes. The polarization of the RPE cells was determined by their high transepithelial electrical resistance (TER >150 Ω cm2) and by their differential secretion of vascular endothelial growth factor (lower layer/upper layer >2.5X). Polarized RPE cells were incubated with 10 ng/ml of TNF-α and the TER was measured. TNF-α significantly decreased the TER of polarized RPE cells by 17.6 ± 2.7% (P < 0.001) of the control at 24 h and that of non-polarized RPE cells by 5.4 ± 6.5% (P = 0.401). The p38 mitogen-activated protein kinase (MAPK) inhibitor, SB203580, blocked the effects of TNF-α of decreasing the TER. Cell junction-related molecules, e.g., ZO-1, located between cells in control RPE cells, were disassembled by TNF-α, and this breakdown was suppressed by SB203580 in polarized RPEs. These results indicate that the breakdown of the RPE barrier function was caused exclusively by TNF-α in polarized RPEs, and TNF-α was acting through the p38 MAPK pathways. Investigations of polarized RPE cells should be more suitable for in vitro studies of the pathophysiology of retinochoroidal diseases.

► Barrier functions differ between polarized- and non-polarized RPE cells in vitro.
► TNF-α causes barrier dysfunction in polarized RPE cells.
►  TNF-α does not cause the barrier dysfunction in non-polarized RPE cells.
►  Action of TNF-α is dependent on ZO-1 acting through the p38 MAPK pathways.
► Investigation of polarized RPE is necessary for studying the pathology of diseases.