Induction of FGF-2 synthesis by IL-1beta in aqueous humor through P13-kinase and p38 in rabbit corneal endothelium.
Induction of FGF-2 synthesis by IL-1beta in aqueous humor through P13-kinase and p38 in rabbit corneal endothelium.
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DOI：
10.1167/iovs.09-4240
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发表时间：
2010-02
影响因子：
4.4
通讯作者：
Kay EP
中科院分区：
文献类型：
--
作者：
Song JS;Lee JG;Kay EP
目的：探讨经角膜冷冻后房水中IL-1β水平升高是否通过PI-3-K和p38途径上调兔角膜内皮细胞成纤维细胞生长因子-2的合成。对新西兰大白兔进行经角膜冷冻，以诱导损伤介导性炎症。用Bio-Plex蛋白芯片检测细胞中IL-1β的浓度，用Western blotting分析成纤维细胞生长因子-2、p38和Akt的表达。免疫荧光染色确定成纤维细胞生长因子-2和肌动蛋白细胞骨架在细胞内的定位。冰冻后可见大量中性粒细胞(PMN)渗入角膜内皮，冰冻后房水中IL-1β浓度呈时间依赖性升高。类似地，成纤维细胞生长因子-2的表达呈时间依赖性增加。用抗FGF2抗体对角膜内皮细胞进行染色时，冷冻后FGF2主要定位于角膜，而正常角膜内皮细胞FGF2定位于细胞膜。IL-1β作用于体外培养的角膜组织，可上调成纤维细胞生长因子-2的表达，促进其在角膜内皮细胞中的核定位。经角膜冷冻破坏了大脑皮层的肌动蛋白细胞骨架，细胞形状从鹅卵石形状变为不规则形状。冷冻后角膜局部应用LY294002和SB203580分别阻断角膜内皮细胞Akt和p38的磷酸化。这些抑制剂还降低了冰冻后成纤维细胞生长因子-2的水平，并部分阻断了冰冻后的形态变化。这些数据表明，经角膜冷冻后，中性粒细胞释放的IL-1β通过PI3-激酶和p38刺激角膜内皮细胞合成成纤维细胞生长因子-2。
To determine whether the elevated level of interleukin (IL)-1β in aqueous humor after transcorneal freezing upregulates FGF-2 synthesis in rabbit corneal endothelium through PI 3-kinase and p38 pathways. Transcorneal freezing was performed in New Zealand white rabbits to induce an injury-mediated inflammation. The concentration of IL-1β was measured with a Bio-Plex Protein Array System, while expression of FGF-2, p38, and Akt were analyzed by Western blotting. Intracellular location of FGF-2 and actin cytoskeleton was determined by immunofluorescent staining. Massive infiltration of polymorphonuclear leukocytes (PMNs) to the corneal endothelium was observed after freezing, and IL-1β concentration in the aqueous humor was elevated in a time-dependent manner after freezing. Similarly, FGF-2 expression was increased in a time-dependent manner. When corneal endothelium was stained with anti-FGF-2 antibody, nuclear location of FGF-2 was mostly observed in the cornea after cryo-treatment, while FGF-2 in normal corneal endothelium was localized at plasma membrane. Treatment of the ex-vivo corneal tissue with IL-1β upregulated FGF-2 and facilitated its nuclear location in corneal endothelium. Transcorneal freezing disrupted the actin cytoskeleton at the cortex, and cell shapes were altered from cobblestone morphology to irregular shape. Topical treatment with LY294002 and SB203580 on the cornea after cryo-treatment blocked phosphorylation of Akt and p38 in corneal endothelium, respectively. These inhibitors also reduced FGF-2 level and partially blocked the morphologic changes after freezing. These data suggest that after transcorneal freezing, IL-1β released by PMNs into aqueous humor stimulates FGF-2 synthesis in corneal endothelium via PI 3-kinase and p38.
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