Kassen Sean C, Ramanan Vijay, Montgomery Jacob E, T Burket Christopher, Liu Chang-Gong, Vihtelic Thomas S, Hyde David R
Department of Biological Sciences, University of Notre Dame, Notre Dame, Indiana 46556, USA.
Dev Neurobiol. 2007 Jul;67(8):1009-31. doi: 10.1002/dneu.20362.
Constant intense light causes apoptosis of rod and cone photoreceptors in adult albino zebrafish. The photoreceptors subsequently regenerate from proliferating inner nuclear layer (INL) progenitor cells that migrate to the outer nuclear layer (ONL) and differentiate into rods and cones. To identify gene expression changes during this photoreceptor regeneration response, a microarray analysis was performed at five time points during the light treatment. The time course included an early time point during photoreceptor death (16 h), later time points during progenitor cell proliferation and migration (31, 51, and 68 h) and a 96 h time point, which likely corresponds to the initial photoreceptor differentiation. Mean expression values for each gene were calculated at each time point relative to the control (0 h light exposure) and statistical analysis by one-way ANOVA identified 4567 genes exhibiting significant changes in gene expression along the time course. The genes within this data set were clustered based on their temporal expression patterns and proposed functions. Quantitative real-time PCR validated the microarray expression profiles for selected genes, including stat3 whose expression increased markedly during the light exposure. Based on immunoblots, both total and activated Stat3 protein expression also increased during the light treatment. Immunolocalization of Stat3 on retinal tissue sections demonstrated increased expression in photoreceptors and Müller glia by 16 h of light exposure. Some of the Stat3-positive Müller cells expressed PCNA at 31 h, suggesting that Stat3 may play a role in signaling a subset of Müller cells to proliferate during the regeneration response.
持续强光会导致成年白化斑马鱼的视杆和视锥光感受器发生凋亡。随后，光感受器会从增殖的内核层（INL）祖细胞再生，这些祖细胞迁移到外核层（ONL）并分化为视杆和视锥。为了确定在这种光感受器再生反应过程中的基因表达变化，在光照处理的五个时间点进行了微阵列分析。时间进程包括光感受器死亡的早期时间点（16小时）、祖细胞增殖和迁移的后期时间点（31、51和68小时）以及一个96小时的时间点，这可能对应于最初的光感受器分化。相对于对照（0小时光照），在每个时间点计算每个基因的平均表达值，通过单因素方差分析进行统计分析，确定了4567个基因在整个时间进程中表现出显著的基因表达变化。该数据集中的基因根据其时间表达模式和推测的功能进行了聚类。定量实时PCR验证了所选基因的微阵列表达谱，包括stat3，其表达在光照期间显著增加。基于免疫印迹，总Stat3蛋白表达和活化的Stat3蛋白表达在光照处理期间也增加。Stat3在视网膜组织切片上的免疫定位显示，光照16小时后，光感受器和Müller胶质细胞中的表达增加。一些Stat3阳性的Müller细胞在31小时时表达增殖细胞核抗原（PCNA），这表明Stat3可能在再生反应期间向一部分Müller细胞发出增殖信号中发挥作用。
