EFFECT OF CYTOKINES AND NITRIC-OXIDE ON TIGHT JUNCTIONS IN CULTURED RAT RETINAL-PIGMENT EPITHELIUM

PURPOSE. These experiments were designed to study the effect of cytoki nes and nitric oxide (NO) on rat retinal pigment epithelial (RPE) cell tight junctions in vitro. METHODS. Cultures of confluent RPE cells fr om retinas of PVG rats (a strain susceptible to development of experim ental uveitis) were prepared on filters and incubated with various sti mulants. The function of the tight junction was evaluated by measuring the transepithelial electrical resistance (TER) of the cell monolayer and the passive permeation of [H-3]inulin across confluent RPE cells. The morphology of the intercellular junctions was visualized by immun olocalization of the tight junction-associated protein zonula occluden s-1 (ZO-1) and F-actin. RESULTS. Seventy-two hours after plating, the RPE cell monolayer showed a mean TER level of 67.6 +/- 18.8 Ohm/cm(2). A decrease in TER was observed after treatment with interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS). The addition of tumor necros is factor-alpha (TNF-alpha) accelerated the decrease of TER, whereas N -G-monomethyl-L-arginine (L-NMMA) (an NO synthase [NOS] inhibitor) did not further modify the resistance decrease. In contrast, 3-morpholino -sydnonimine (SIN-1), a sydnonimine analog and NO donor, increased the TER. The variations of TER were correlated with the transepithelial f luxes of [H-3] inulin and with tight junction morphologic changes of Z O-1 and F-actin immunostaining. CONCLUSIONS. Incubation with LPS assoc iated with IFN-gamma and TNF-alpha induces alterations of RPE right ju nctions, whereas NO is involved in the maintenance of their integrity. Cytokines and NO production could play a role in regulation of the bl ood-ocular barrier function and of the development of ocular inflammat ion.