The retinal pigment epithelium (RPE) plays an important role in maintenance of homeostasis in the back of the eye by transporting fluid and secreting a variety of molecules including cytokines/chemokines. In ocular inflammation, affected by the elevated pro-inflammatory cytokines, the physiologic properties of RPE such as fluid transport and chemokine secretion may change resulting in exacerbation of the diseases. To investigate this question, we stimulated the cultured human fetal RPE with a mixture of TNF-alpha, IFN-gamma and IL-1beta. Apical addition of the mixture produced no changes in Jv. In contrast, basal addition increased net fluid absorption by 8.6 ? 2.9 ul?cm-2?hr-1 (n=3). Jv increased by 5.5 ? 1.3 ul?cm-2?hr-1 (n=3) after addition to both sides. Stimulation with the mixture, profoundly increased the secretion of the tested 12 chemokines and 3 cytokines. For the majority of these molecules, stimulation from the apical side was much more effective than that from the basal side and the secretion from the apical side was greater by far than that from the basal side. The highly affected chemokines were four CC chemokines (MCP-1, RANTES, MCP-2, MCP-3) and four CXC chemokines (GRO-alpha, IL-8, IP-10, I-TAC). In 3 independent experiments (n=3), for CC chemokines, apical stimulation increased secretion on apical side from 6.9 + 1.2 ng/well to 106.2 + 15.1 ng/well for MCP-1, from 0 to 10.5 + 0.6 ng/well for RANTES, from 69.1 + 1.2 pg/well to 17.1 + 0.7 ng/well for MCP-2, from 6.5 + 1.4 pg/well to 11.2 + 0.7 ng/well for MCP-3. For CXC chemokines, apical stimulation increased secretion on apical side from 15.7 + 3.2 pg/well to 36.2 + 0.3 ng/well for GRO-alpha, from 140.6 + 9.6 pg/well to 55.4 + 9.6 ng/well for IL-8, from 5.1 + 0.3 pg/well to 119.9 + 10.8 ng/well for IP-10, from 9.5 + 2.1 pg/well to 44.6 + 1.0 ng/well for I-TAC. The pro-inflammatory-cytokine-induced activation of JV and the predominant increase of secretion of most chemokine/cytokines to the apical bath suggest a critical role of the RPE in the pathogenesis of acute and chronic phases of ocular inflammatory diseases.