Effects of Inflammatory Cytokines on Human Fetal RPE in vitro

视网膜色素上皮（RPE）通过运输流体和分泌包括细胞因子/趋化因子的各种分子在维持眼睛后部的稳态中起重要作用。在眼部炎症中，受升高的促炎细胞因子的影响，RPE的生理特性如流体运输和趋化因子分泌可能改变，导致疾病的恶化。为了研究这个问题，我们用TNF-α、IFN-γ和IL-1 β的混合物刺激培养的人胎儿RPE。 顶部添加的混合物没有产生任何变化Jv。相比之下，基础添加增加净液体吸收8.6 ± 2.9 μ l·cm-27 hr-1（n=3）。双侧加药后Jv增加5.5 ± 1.3 μ l·cm ~（-27）hr ~（-1）（n=3）。用混合物刺激，显著增加了测试的12种趋化因子和3种细胞因子的分泌。对于这些分子中的大多数，从顶侧的刺激比从基底侧的刺激有效得多，并且从顶侧的分泌远远大于从基底侧的分泌。高度受影响的趋化因子是四种CC趋化因子（MCP-1、RANTES、MCP-2、MCP-3）和四种CXC趋化因子（GRO-α、IL-8、IP-10、I-TAC）。在3个独立的实验（n=3）中，对于CC趋化因子，顶侧刺激增加顶侧分泌，对于MCP-1从6.9 ± 1.2 ng/孔增加到106.2 ± 15.1 ng/孔，对于RANTES从0增加到10.5 ± 0.6 ng/孔，对于MCP-2从69.1 ± 1.2 pg/孔增加到17.1 ± 0.7 ng/孔，MCP-3从6.5 ± 1.4 μ g/孔至11.2 ± 0.7 ng/孔。对于CXC趋化因子，顶端刺激增加顶端侧的分泌，对于GRO-α从15.7 ± 3.2 pg/孔增加到36.2 ± 0.3 ng/孔，对于IL-8从140.6 ± 9.6 pg/孔增加到55.4 ± 9.6 ng/孔，对于IP-10从5.1 ± 0.3 pg/孔增加到119.9 ± 10.8 ng/孔，I-TAC从9.5 ± 2.1 pg/孔至44.6 ± 1.0 ng/孔。 JV的促炎-精氨酸诱导的激活和大多数趋化因子/细胞因子分泌到顶端浴的显著增加表明RPE在眼部炎性疾病的急性和慢性阶段的发病机制中起关键作用。

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 1 2.9 ul7cm-27hr-1 (n=3). Jv increased by 5.5 1 1.3 ul7cm-27hr-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.