Division of Molecular and Developmental Biology, Institute of Medical Science, University of Tokyo, Tokyo, Japan.
Invest Ophthalmol Vis Sci. 2011 May 6;52(6):3008-17. doi: 10.1167/iovs.10-6428.
Much attention has been paid to the roles of microRNA in developmental and biological processes. Dicer plays essential roles in cell survival and proliferation in various organs. We examined the role of Dicer in retinal development using retina-specific conditional knockout of Dicer in mice.
Dkk3-Cre expressed the Cre gene in retinal progenitor cells from an early embryonic stage. The authors analyzed Dkk-Cre/Dicer-flox (Dicer-CKO) mice for their survival, proliferation, and differentiation. To analyze the role of Dicer in later stages of retinal development, a Cre expression plasmid was introduced into the neonatal retina by electroporation, and retinal differentiation was examined.
Dicer-CKO mice were born at the numbers we expected, based on Mendelian genetics, but their eyes never opened. Massive death of retinal progenitor cells occurred during embryogenesis, resulting in microphthalmia, and most retinal cells had disappeared by postnatal day 14. In vitro reaggregation culture of Dicer-CKO retinal cells showed that cell death and the suppression of proliferation by Dicer inactivation occurred in a cell-autonomous manner. Cell differentiation markers were expressed in the Dicer-CKO retina; however, these cells localized abnormally, and the inner plexiform layer was absent, suggesting that cell migration and morphologic differentiation, especially process extension, were perturbed. Forced neonatal expression of Cre induced apoptosis and affected the expression of differentiation markers.
Taken together, these results show that Dicer is essential during early retinal development.
人们对 microRNA 在发育和生物过程中的作用给予了极大关注。Dicer 在各种器官的细胞存活和增殖中发挥着重要作用。我们使用视网膜特异性条件性 Dicer 敲除小鼠来研究 Dicer 在视网膜发育中的作用。
Dkk3-Cre 在胚胎早期的视网膜祖细胞中表达 Cre 基因。作者分析了 Dkk-Cre/Dicer-flox(Dicer-CKO) 小鼠的存活、增殖和分化情况。为了分析 Dicer 在视网膜发育后期的作用，通过电穿孔将 Cre 表达质粒引入新生鼠的视网膜，并检测视网膜分化情况。
Dicer-CKO 小鼠的出生数量符合孟德尔遗传规律，但它们的眼睛从未睁开过。大量的视网膜祖细胞在胚胎发生过程中死亡，导致小眼畸形，并且大多数视网膜细胞在出生后第 14 天已经消失。Dicer-CKO 视网膜细胞的体外再聚集培养表明，Dicer 失活导致细胞死亡和增殖受到抑制，这种现象是细胞自主发生的。Dicer-CKO 视网膜中表达了细胞分化标志物；然而，这些细胞的定位异常，内丛状层缺失，这表明细胞迁移和形态分化，特别是突起延伸受到干扰。新生期强制表达 Cre 诱导了细胞凋亡，并影响了分化标志物的表达。
综上所述，这些结果表明 Dicer 在早期视网膜发育中是必需的。
