Smith S B, Bora N, McCool D, Kutty G, Wong P, Kutty R K, Wiggert B
Department of Cellular Biology and Anatomy, Medical College of Georgia, Augusta 30912-2000, USA.
Invest Ophthalmol Vis Sci. 1995 Oct;36(11):2193-201.
To determine the mechanism of photoreceptor cell death in the vitiligo mouse, a model of retinal degeneration in which the genetic defect is not retina specific but is instead caused by single point mutation in the microphthalmia (mi) gene that codes for a basic helix-loop-helix DNA transcription factor.
Detection of apoptotic cells was performed in fixed retinal tissue using the TUNEL assay in animals 1, 2, 4, 6, 8, 16, 32, 40, and 52 weeks. Electron microscopic analysis was used to confirm the morphologic hallmarks of apoptosis, and Southern blot analysis was used to detect internucleosomal DNA fragmentation. Additionally, the expression of a gene associated with apoptosis, TRPM-2/clusterin, was examined.
At ages beyond the time of normal retinal programmed cell death, vitiligo retinas had significantly more TUNEL-positive photoreceptor cells and more photoreceptor cells with condensed chromatin than controls. DNA internucleosomal fragmentation ladders were present in vitiligo retinas even as late as 15 weeks, a time well beyond developmental apoptosis in controls. TRPM-2/clusterin mRNA levels in vitiligo neural retinas were similar to controls initially but were two times greater than controls by 12 weeks. Surprisingly, TRPM-2/clusterin mRNA levels were elevated in the retinal pigment epithelium in the mutant; the expression at one week was two times greater than normals and remained elevated for many months, even though retinal pigment epithelial cells showed no morphologic evidence of apoptosis.
The morphologic and biochemical data suggest that photoreceptor cells die by apoptosis in vitiligo mice. The increased retinal TRPM-2/clusterin mRNA levels may be a direct response to these events. The increased expression of this gene in the retinal pigment epithelium, however, may reflect its role in tissue regression and membrane remodeling. Mechanisms by which the mi gene defect might result in the vitiligo retinopathy are proposed.
在白癜风小鼠中确定光感受器细胞死亡的机制。白癜风小鼠是一种视网膜变性模型，其基因缺陷并非视网膜特异性的，而是由编码碱性螺旋-环-螺旋DNA转录因子的小眼症（mi）基因中的单点突变引起的。
在1、2、4、6、8、16、32、40和52周龄的动物中，使用TUNEL法在固定的视网膜组织中检测凋亡细胞。电子显微镜分析用于确认凋亡的形态学特征，Southern印迹分析用于检测核小体间DNA片段化。此外，检测了与凋亡相关的基因TRPM-2/簇集蛋白的表达。
在超过正常视网膜程序性细胞死亡时间的年龄段，白癜风视网膜中TUNEL阳性光感受器细胞和染色质浓缩的光感受器细胞明显多于对照组。即使在15周时，白癜风视网膜中仍存在DNA核小体间片段化梯带，这一时间远远超过了对照组的发育性凋亡时间。白癜风神经视网膜中TRPM-2/簇集蛋白mRNA水平最初与对照组相似，但到12周时比对照组高两倍。令人惊讶的是，突变体视网膜色素上皮中TRPM-2/簇集蛋白mRNA水平升高；1周时的表达比正常高两倍，并在许多个月内持续升高，尽管视网膜色素上皮细胞没有凋亡的形态学证据。
形态学和生化数据表明，白癜风小鼠的光感受器细胞通过凋亡死亡。视网膜中TRPM-2/簇集蛋白mRNA水平的升高可能是对这些事件的直接反应。然而，该基因在视网膜色素上皮中的表达增加可能反映了其在组织退化和膜重塑中的作用。提出了mi基因缺陷可能导致白癜风视网膜病变的机制。
