Department of Otolaryngology-Head and Neck Surgery, Stanford University, CA 94305, USA.
J Assoc Res Otolaryngol. 2011 Aug;12(4):455-69. doi: 10.1007/s10162-011-0267-2. Epub 2011 Apr 7.
The Wnt signaling pathway is a recurring theme in tissue development and homeostasis. Its specific roles during inner ear development are just emerging, but few studies have characterized Wnt target genes. Lgr5, a member of the G protein-coupled receptor family, is a Wnt target in the gastrointestinal and integumentary systems. Although its function is unknown, its deficiency leads to perinatal lethality due to gastrointestinal distension. In this study, we used a knock-in reporter mouse to examine the spatiotemporal expression of Lgr5 in the cochlear duct during embryonic and postnatal periods. In the embryonic day 15.5 (E15.5) cochlear duct, Lgr5-EGFP is expressed in the floor epithelium and overlapped with the prosensory markers Sox2, Jagged1, and p27(Kip1). Nascent hair cells and supporting cells in the apical turn of the E18.5 cochlear duct express Lgr5-EGFP, which becomes downregulated in hair cells and subsets of supporting cells in more mature stages. In situ hybridization experiments validated the reporter expression, which gradually decreases until the second postnatal week. Only the third row of Deiters' cells expresses Lgr5-EGFP in the mature organ of Corti. Normal cochlear development was observed in Lgr5(EGFP/EGFP) and Lgr5(EGFP/+) mice, which exhibited normal auditory thresholds. The expression pattern of Lgr5 contrasts with another Wnt target gene, Axin2, a feedback inhibitor of the Wnt pathway. Robust Axin2 expression was found in cells surrounding the embryonic cochlear duct and becomes restricted to tympanic border cells below the basilar membrane in the postnatal cochlea. Both Lgr5 and Axin2 act as Wnt targets in the cochlea because purified Wnt3a promoted and Wnt antagonist suppressed their expression. Their differential expression among cell populations highlights the dynamic but complex distribution of Wnt-activated cells in and around the embryonic and postnatal cochlea.
Wnt 信号通路是组织发育和稳态中的一个反复出现的主题。其在内耳发育过程中的特定作用刚刚出现,但很少有研究描述 Wnt 靶基因。Lgr5 是 G 蛋白偶联受体家族的成员,是胃肠道和皮肤系统中的 Wnt 靶标。尽管其功能未知,但由于胃肠道扩张,其缺失会导致围产期致死。在这项研究中,我们使用敲入报告小鼠检查 Lgr5 在胚胎期和出生后耳蜗管中的时空表达。在胚胎期 15.5 天 (E15.5) 的耳蜗管中,Lgr5-EGFP 在基板上皮中表达,并与前感觉标记物 Sox2、Jagged1 和 p27(Kip1) 重叠。E18.5 耳蜗管中顶端的新生毛细胞和支持细胞表达 Lgr5-EGFP,该表达在更成熟阶段下调至毛细胞和支持细胞亚群。原位杂交实验验证了报告基因的表达,该表达逐渐降低,直到出生后第二周。在成熟的 Corti 器官中,只有第三排 Deiters 细胞表达 Lgr5-EGFP。在 Lgr5(EGFP/EGFP) 和 Lgr5(EGFP/+) 小鼠中观察到正常的耳蜗发育,其听觉阈值正常。Lgr5 的表达模式与另一个 Wnt 靶基因 Axin2 形成对比,Axin2 是 Wnt 通路的反馈抑制剂。在胚胎耳蜗管周围的细胞中发现了强烈的 Axin2 表达,在出生后的耳蜗中,它被限制在基底膜下方的鼓膜边界细胞中。Lgr5 和 Axin2 都作为耳蜗中的 Wnt 靶标,因为纯化的 Wnt3a 促进了它们的表达,而 Wnt 拮抗剂则抑制了它们的表达。它们在细胞群体中的差异表达突出了 Wnt 激活细胞在胚胎和出生后耳蜗内和周围的动态但复杂的分布。