Increased clusterin expression in Fuchs' endothelial dystrophy
Increased clusterin expression in Fuchs' endothelial dystrophy
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DOI：
10.1167/iovs.07-1405
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发表时间：
2008-07-01
影响因子：
4.4
通讯作者：
Joyce, Nancy C.
中科院分区：
文献类型：
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作者：
Jurkunas, Ula V.;Bitar, Maya S.;Joyce, Nancy C.
目的。目的探讨糖蛋白簇蛋白/apoJ（CLU）在正常和Fuchs内皮营养不良（FED）角膜内皮中的差异表达，并比较正常和FED组织中各种形式的CLU的表达。 FED和人工晶状体大疱性角膜病（PBK）角膜纽扣在移植过程中被移除，并从组织库中获得正常角膜。从基质中切下人角膜内皮细胞和后弹力层 (HCEC-DM) 复合物。在二维凝胶上分离蛋白质并进行比较蛋白质组分析。通过蛋白质印迹分析比较正常和 FED HCEC-DM 之间分泌前 CLU (pre-sCLU)、分泌性 (s) CLU 和核 (n) CLU 的相对表达。使用RT-PCR比较CLU mRNA的表达。通过免疫细胞化学和共聚焦显微镜比较正常眼和 FED 眼的角膜整体中 CLU 的亚细胞定位。结果。蛋白质组学分析显示，与正常对照相比，FED HCEC-DM 中 CLU 表达明显增加。 Western blot 分析表明，FED 中前 sCLU 蛋白表达比正常样本高 5.2 倍 (P = 3.52E-05)，而分泌修饰的成熟形式 (sCLU) 并未显着升高 (P = 0.092)。 FED 中 nCLU 蛋白的表达显着升高（P = 0.013）。 RT-PCR 分析显示，FED 样品中的 CLU mRNA 显着增加（P = 0.002），但 PBK 样品中则没有。 CLU 在 FED 样品中也有独特的定位，在肠管周围和内皮细胞的细胞核中细胞内染色增强。结论。 CLU 表达在受 FED 影响的组织中显着升高，表明 FED 发病机制中存在一种尚未发现的内皮细胞功能失调形式。
PURPOSE. To investigate the differential expression of the glycoprotein clusterin/apoJ (CLU) in normal and Fuchs' endothelial dystrophy (FED) corneal endothelium and to compare the expression of various forms of CLU in normal and FED tissue.METHODS. FED and pseudophakic bullous keratopathy (PBK) corneal buttons were removed during transplantation, and normal corneas were obtained from tissue banks. Human corneal endothelial cells and Descemet's membrane (HCEC-DM) complex was dissected from the stroma. Proteins were separated on 2-D gels and subjected to comparative proteomic analysis. Relative expression of presecretory CLU (pre-sCLU), secretory (s) CLU, and nuclear (n) CLU were compared between normal and FED HCEC-DM by Western blot analysis. Expression of CLU mRNA was compared by using RT-PCR. Subcellular localization of CLU was compared in corneal wholemounts from normal eyes and eyes with FED by immunocytochemistry followed by confocal microscopy.RESULTS. Proteomic analysis revealed an apparent increase in CLU expression in FED HCEC-DM compared with the normal control. Western blot analysis demonstrated that pre-sCLU protein expression was 5.2 times higher in FED than in normal samples (P = 3.52E-05), whereas the mature form modified for secretion (sCLU) was not significantly elevated (P = 0.092). Expression of nCLU protein was significantly elevated in FED (P = 0.013). RT-PCR analysis revealed that CLU mRNA was significantly increased (P = 0.002) in FED samples, but not in PBK samples. CLU also had a distinctive localization in FED samples with enhanced intracellular staining around the guttae and in the nuclei of endothelial cells.CONCLUSIONS. CLU expression is markedly elevated in FED-affected tissue, pointing to a yet undiscovered form of dysregulation of endothelial cell function involved in FED pathogenesis.