Becker D L, Mobbs P
Department of Anatomy and Developmental Biology, Department of Physiology, University College London, Gower Street, WC1E 6BT, London, UK.
Exp Neurol. 1999 Apr;156(2):326-32. doi: 10.1006/exnr.1999.7027.
During the formation of the eye, high levels of connexin alpha1 (connexin 43) are expressed within the tissues of the cornea, lens, and neural retina. In order to determine whether connexin alpha1 plays a role in the regulation of cell proliferation we have used a novel antisense technique to reduce its expression early in development (embryonic days 2-4). Application of Pluronic gel, containing antisense oligodeoxynucleotides (ODNs) to connexin alpha1, to one eye of early chick embryos results in a rapid and significant reduction of alpha1 protein which lasts for 24-48 h. Embryos grown for 48 h, after ODN application to one eye, showed a marked reduction in the diameter of the treated, compared to that of the contralateral untreated, eye. Sections cut from the treated eyes showed that the retina was also reduced in size. TUNEL labeling and staining with propidium iodide showed that apoptosis within the retinae of both treated and untreated eyes was rare and thus that the reduction in the area of the retina brought about by antisense ODNs directed at connexin alpha1 was unlikely to be the result of increased cell death. However, the number of mitotic figures in the ventricular zone of the antisense-treated retinae revealed by propidium iodide staining was significantly reduced (P < 0.0001) to 53 +/- 3.5% (n = 5) of that in the contralateral untreated control eyes. Embryos in which one eye was sham operated, treated with pluronic gel, or treated with sense ODN showed no significant changes in eye size or in the number of mitotic figures within the neural retina. These results point to a role for connexin alpha1-mediated gap-junctional communication in controlling the early wave of neurogenesis in the chick retina.
在眼睛形成过程中，连接蛋白α1（连接蛋白43）在角膜、晶状体和神经视网膜组织中高水平表达。为了确定连接蛋白α1是否在细胞增殖调节中发挥作用，我们使用了一种新型反义技术在发育早期（胚胎第2 - 4天）降低其表达。将含有连接蛋白α1反义寡脱氧核苷酸（ODN）的普朗尼克凝胶应用于早期鸡胚的一只眼睛，导致α1蛋白迅速且显著减少，持续24 - 48小时。在向一只眼睛应用ODN后生长48小时的胚胎，与对侧未处理的眼睛相比，处理过的眼睛直径明显减小。从处理过的眼睛切取的切片显示视网膜尺寸也减小。TUNEL标记和碘化丙啶染色表明，处理过和未处理过的眼睛视网膜内的细胞凋亡都很少，因此，针对连接蛋白α1的反义ODN导致的视网膜面积减小不太可能是细胞死亡增加的结果。然而，碘化丙啶染色显示，反义处理的视网膜室管膜区有丝分裂细胞数量显著减少（P < 0.0001），降至对侧未处理对照眼睛的53±3.5%（n = 5）。单眼进行假手术、用普朗尼克凝胶处理或用正义ODN处理的胚胎，眼睛大小或神经视网膜内有丝分裂细胞数量均无显著变化。这些结果表明连接蛋白α1介导的缝隙连接通讯在控制鸡视网膜神经发生早期浪潮中发挥作用。
