Rungger-Brändle Elisabeth, Conti Alain, Leuenberger Peter M, Rungger Duri
Cell Biology Laboratory, University Eye Clinic, 22, rue Alcide-Jentzer, CH-1211 Genève, Switzerland.
Exp Eye Res. 2005 Nov;81(5):539-50. doi: 10.1016/j.exer.2005.03.012. Epub 2005 Jun 1.
In order to re-evaluate functional implications of alphasmooth muscle actin (alphaSMA) expression in lens epithelial cells (LECs), we assessed its presence in donor lenses without visible opacities (DON), lenses with mature cataract (CAT), and cataractous lenses with posterior subcapsular opacities (PSO) or anterior subcapsular fibrosis (ASF). The levels of alphaSMA and transforming growth factor-beta2 (TGFbeta2) mRNAs were measured by classical and real-time PCR. Expression and structural organisation of alphaSMA protein and beta-catenin were monitored by Western blotting and confocal microscopy. All DON analysed contained measurable amounts of alphaSMA mRNA. In CAT without and with PSO, mRNA expression was increased and, again more than doubled in ASF. TGFbeta2 mRNA expression varied widely between the individual samples but was slightly increased in ASF. No correlation existed between alphaSMA or TGFbeta2 expression and the age of the donors in any of the lens categories. Confocal microscopy revealed that, in DON and CAT, alphaSMA was preferentially expressed in a simple granular pattern in single or small clusters of LECs within a normally shaped cobblestone epithelium. Locally, the granules were merged into short stretches at the cell margin. In CAT, a few abnormally shaped cells contained polygonal alphaSMA structures and short stress fibres. In CAT with PSO and ASF, polygons and stress fibre bundles predominated in spindle-shaped cells. Expression patterns of different complexity were often present in the same epithelium. Apical polygons and basal stress fibres were detected within the same cell and may reflect instability of the interface between epithelium and cortical fibres and changes in adhesion to the capsule, respectively. High levels of betacatenin mRNA and protein were present in all lens types. However, with increasing complexity of alphaSMA organisation, betacatenin staining disappeared from the cell margin and basal infoldings and was shifted towards the cytoplasm and nucleus. The presence of alphaSMA in DON, the absence of any correlation between mRNA level and age, and the modest increase in complexity of alphaSMA-containing structures in CAT argue against an inevitable link between alphaSMA expression and the development of age-related cataract. Low levels of alphaSMA expression may reflect repair of normal wear and tear. In pathologic situations such as PSO and ASF, persisting stimulation and additional incentives may induce increased alphaSMA expression and more elaborate patterning, eventually leading to completion of EMT.
为了重新评估α平滑肌肌动蛋白（αSMA）在晶状体上皮细胞（LEC）中的功能意义，我们评估了其在无可见混浊的供体晶状体（DON）、成熟白内障晶状体（CAT）以及伴有后囊下混浊（PSO）或前囊下纤维化（ASF）的白内障晶状体中的存在情况。通过经典PCR和实时PCR检测αSMA和转化生长因子-β2（TGFβ2）mRNA的水平。通过蛋白质印迹法和共聚焦显微镜监测αSMA蛋白和β-连环蛋白的表达及结构组织。所有分析的DON均含有可测量量的αSMA mRNA。在无PSO和伴有PSO的CAT中，mRNA表达增加，在ASF中再次增加一倍多。TGFβ2 mRNA表达在各个样本之间差异很大，但在ASF中略有增加。在任何晶状体类别中，αSMA或TGFβ2表达与供体年龄之间均无相关性。共聚焦显微镜显示，在DON和CAT中，αSMA优先以简单颗粒模式在正常形状鹅卵石样上皮内的单个或小簇LEC中表达。在局部，颗粒在细胞边缘融合成短片段。在CAT中，一些形状异常的细胞含有多边形αSMA结构和短应力纤维。在伴有PSO和ASF的CAT中，多边形和应力纤维束在梭形细胞中占主导。不同复杂程度的表达模式常常出现在同一上皮中。在同一细胞内检测到顶端多边形和基底应力纤维，分别可能反映上皮与皮质纤维之间界面的不稳定性以及与囊膜黏附的变化。所有晶状体类型中均存在高水平的β-连环蛋白mRNA和蛋白。然而，随着αSMA组织复杂性的增加，β-连环蛋白染色从细胞边缘和基底褶皱消失，并向细胞质和细胞核转移。DON中αSMA的存在、mRNA水平与年龄之间缺乏任何相关性以及CAT中含αSMA结构复杂性的适度增加，均表明αSMA表达与年龄相关性白内障的发生之间不存在必然联系。低水平的αSMA表达可能反映正常磨损的修复。在PSO和ASF等病理情况下，持续的刺激和额外的诱因可能诱导αSMA表达增加和更复杂的模式形成，最终导致上皮-间质转化完成。
