Table_9_Wnt Signaling Drives Correlated Changes in Facial Morphology and Brain Shape.XLSX

Canonical Wnt signaling plays multiple roles critical to normal craniofacial development while its dysregulation is known to be involved in structural birth defects of the face. However, when and how Wnt signaling influences phenotypic variation, including those associated with disease, remains unclear. One potential mechanism is via Wnt signaling’s role in the patterning of an early facial signaling center, the frontonasal ectodermal zone (FEZ), and its subsequent regulation of early facial morphogenesis. For example, Wnt signaling may directly alter the shape and/or magnitude of expression of the sonic hedgehog (SHH) domain in the FEZ. To test this idea, we used a replication-competent avian sarcoma retrovirus (RCAS) encoding Wnt3a to modulate its expression in the facial mesenchyme. We then quantified and compared ontogenetic changes in treated to untreated embryos in the three-dimensional (3D) shape of both the SHH expression domain of the FEZ, and the morphology of the facial primordia and brain using iodine-contrast microcomputed tomography imaging and 3D geometric morphometrics (3DGM). We found that increased Wnt3a expression in early stages of head development produces correlated variation in shape between both structural and signaling levels of analysis. In addition, altered Wnt3a activation disrupted the integration between the forebrain and other neural tube derivatives. These results show that activation of Wnt signaling influences facial shape through its impact on the forebrain and SHH expression in the FEZ, and highlights the close relationship between morphogenesis of the forebrain and midface.

经典Wnt信号通路（Canonical Wnt signaling）在正常颅面发育中发挥多种关键作用，而其失调已被证实与面部结构性出生缺陷相关。然而，Wnt信号通路究竟在何时、以何种方式影响表型变异（包括与疾病相关的表型变异），目前仍未明确。一种潜在机制是，Wnt信号通路参与调控早期面部信号中心——额鼻外胚层区（frontonasal ectodermal zone, FEZ）的模式建成，并进而调控早期面部形态发生。例如，Wnt信号通路可能直接改变额鼻外胚层区中音猬因子（sonic hedgehog, SHH）表达域的形状与/或表达强度。为验证这一假说，本研究使用编码Wnt3a的复制型禽肉瘤逆转录病毒（replication-competent avian sarcoma retrovirus, RCAS），对面部间充质的Wnt3a表达进行调控。随后，我们通过碘造影显微计算机断层扫描成像与三维几何形态测量学（3D geometric morphometrics, 3DGM），定量比较了处理组与未处理组胚胎的个体发育差异：涵盖额鼻外胚层区SHH表达域的三维形状，以及面部原基与脑组织的形态特征。本研究发现，在头部发育早期升高的Wnt3a表达，会在结构分析与信号分析两个层面均引发相关的形状变异。此外，Wnt3a激活异常会破坏前脑与其他神经管衍生物之间的整合关联。本研究结果表明，Wnt信号通路的激活通过影响前脑以及额鼻外胚层区中的SHH表达来调控面部形态，并凸显了前脑与中面部形态发生之间的紧密关联。