Department of Ophthalmology, Kochi Medical School, Nankoku City, Kochi, Japan.
Invest Ophthalmol Vis Sci. 2013 Aug 27;54(8):5817-23. doi: 10.1167/iovs.13-11969.
Severe ocular allergic diseases are characterized by pronounced conjunctival inflammation triggered by T helper 2 (Th2) cells and corneal epithelial damage induced by eosinophils. To examine the role of alarmins released by damaged corneal epithelial cells in tissue eosinophilia, we investigated the effects of a supernatant derived from necrotic human corneal epithelial (HCE) cells on expression of the chemokine CCL11 (eotaxin) and the adhesion molecule VCAM-1 in human corneal fibroblasts.
An alarmin preparation was obtained as the material released from HCE cells after three cycles of freezing and thawing. CCL11 released into culture medium and cell surface expression of VCAM-1 were measured with enzyme-linked immunosorbent assays, and the amounts of CCL11 and VCAM-1 mRNAs were quantitated by reverse transcription and real-time polymerase chain reaction analysis. Signaling by the transcription factor NF-κB was evaluated by immunoblot and immunofluorescence analyses.
The combination of the necrotic HCE cell supernatant and either interleukin (IL)-4 or IL-13 induced synergistic increases in CCL11 release, VCAM-1 expression, and the abundance of CCL11 and VCAM-1 mRNAs in corneal fibroblasts. The necrotic HCE cell supernatant also induced NF-κB activation in corneal fibroblasts, whereas an inhibitor of NF-κB and IL-1 receptor antagonist each attenuated CCL11 release induced by the alarmin preparation and either IL-4 or IL-13.
Alarmins including IL-1 released from necrotic corneal epithelial cells cooperate with Th2 cytokines to induce CCL11 production and VCAM-1 expression in corneal fibroblasts, and may thereby play an important role in tissue eosinophilia associated with ocular allergic diseases.
严重的眼部过敏性疾病的特征是由辅助性 T 细胞 2（Th2）细胞引发的明显结膜炎症和由嗜酸性粒细胞引起的角膜上皮损伤。为了研究受损角膜上皮细胞释放的警报素在组织嗜酸性粒细胞中的作用，我们研究了来自坏死的人角膜上皮（HCE）细胞的上清液对人角膜成纤维细胞中趋化因子 CCL11（嗜酸性粒细胞趋化因子）和黏附分子 VCAM-1 表达的影响。
通过对 HCE 细胞进行三轮冻融处理，获得警报素制剂。通过酶联免疫吸附试验测量培养上清液中释放的 CCL11 和细胞表面 VCAM-1 的表达，通过逆转录和实时聚合酶链反应分析定量 CCL11 和 VCAM-1 mRNA 的量。通过免疫印迹和免疫荧光分析评估转录因子 NF-κB 的信号转导。
坏死的 HCE 细胞上清液与白细胞介素（IL）-4 或 IL-13 的组合诱导 CCL11 释放、VCAM-1 表达以及角膜成纤维细胞中 CCL11 和 VCAM-1 mRNA 的丰度协同增加。坏死的 HCE 细胞上清液还诱导了角膜成纤维细胞中 NF-κB 的激活，而 NF-κB 抑制剂和 IL-1 受体拮抗剂均可减弱警报素制剂和 IL-4 或 IL-13 诱导的 CCL11 释放。
包括 IL-1 在内的来自坏死的角膜上皮细胞的警报素与 Th2 细胞因子合作，诱导角膜成纤维细胞中 CCL11 的产生和 VCAM-1 的表达，因此可能在与眼部过敏性疾病相关的组织嗜酸性粒细胞中发挥重要作用。
