Mouse Model for Eyelid Disease: Congenital Blepharophimosis

描述（由申请人提供）：睑下垂、上睑下垂和上眦内翻综合征（BPES）是一种常染色体显性遗传疾病，以颅面缺陷为特征，主要影响眼睑的发育。BPES有两种类型：I型包括眼睑下垂、上睑下垂、内眦赘肉和远眦赘肉四个主要特征，并伴有卵巢早衰（POF），导致女性不孕。II型仅包括眼睑畸形，无性别偏好。患有BPES的人患近视（近视）或远视（远视）等视力问题的风险更高。他们也可能有眼睛不指向同一个方向（斜视）和影响一只或两只眼睛的弱视（弱视）。转录因子叉头盒L2 （FOXL2）结构基因的突变导致70%的BPES。FOXL2基因为一种蛋白质的生成提供指令，这种蛋白质在出生前参与眼睑和卵巢的发育。大约30%的BPES患者没有FOXL2结构基因突变；这些人患病的原因尚不清楚，但FOXL2基因的调控可能发生了改变。在此，我们研究了Notch1信号激活对眼周间充质细胞（POMC）的影响，POMC有助于眼睑特异性结构的形成，包括提上睑肌、平滑肌、跗骨和睑板腺。Notch1胞内结构域（N1-ICD）在新型三转基因小鼠Kera-rtTA/tetO-Cre/R26floxedN1-ICD （KR/TC/R26fN1- ICD）的POMC（这里简称POMCN1-ICD）中，通过多西环素（Dox）在不同发育阶段的脉冲诱导，有条件地错表达。这些三重KR/TC/R26fN1-ICD小鼠表现出与人类BPES相似的眼睑异常变化。我们的初步数据显示，在眼睑形态发生过程中，N1-ICD的表达导致POMCs中FoxL2和平滑肌分化标记基因α -平滑肌动蛋白（a-SMA）的表达特异性降低。我们的初步研究允许我们提出这样的假设，即持续的Notch信号通过下调POMC细胞中的FoxL2来干扰负责眼睑打开的提上睑肌平滑肌的形成。在本应用中，我们将进一步将KR/TC/R26fN1-ICD三重转基因小鼠作为研究先天性BPES （Aim1）发病机制的新型动物模型。我们还将描述在胚胎眼睑发育过程中导致FoxL2下调和随后提上睑平滑肌畸形的分子途径（Aim2）。

DESCRIPTION (provided by applicant): Blephar-ophimosis, Ptosis, and Epi-canthus inversus Syndrome (BPES) is an autosomal dominant genetic disorder characterized by craniofacial defects that mainly affect the development of the eyelids. There are two types of BPES: Type I consists of the four major features of blepharophimosis, ptosis, epicanthus inversus, and telecanthus plus premature ovarian failure (POF), leading to infertility in woman. Type II consists of only the eyelid malformations without gender preference. People with BPES are at an increased risk of developing vision problems such as nearsightedness (myopia) or farsightedness (hyperopia). They may also have eyes that do not point in the same direction (strabismus) and lazy eye (amblyopia) affecting one or both eyes. Mutations in the transcription factor forkhead box L2 (FOXL2) structure gene cause 70 percent of BPES. The FOXL2 gene provides instructions for making a protein that is involved in the development of the eyelids and the ovaries before birth. Approximately 30 percent of people with BPES do not have an identified FOXL2 structure gene mutation; the cause of the condition in these people is unknown but FOXL2 gene regulation maybe altered. Herein, we have investigated the effects of Notch1 signaling activation in peri-ocular mesenchymal cells (POMC) which contribute to the formation of the lid-specific structures including levator M¿ellersmooth muscle, tarsus, and meibomian glands. Notch1 intracellular domain (N1-ICD) was conditionally mis-expressed in POMC (here refer to as POMCN1-ICD) of a novel triple transgenic mouse strain, namely Kera-rtTA/tetO-Cre/R26floxedN1-ICD (KR/TC/R26fN1- ICD), by pulse induction of doxycycline (Dox) at different developmental stages. These triple KR/TC/R26fN1-ICD mice exhibited variegation of eyelid anomalies resembling BPES in humans. Our preliminary data showed that N1-ICD expression caused specific reduction of FoxL2 and smooth muscle differentiation marker gene, alpha-smooth actin (a-SMA) expressions in POMCs during eyelid morphogenesis. Our preliminary studies allow us to propose the hypotheses that sustained Notch signaling disturbs the formation of levator M¿eller smooth muscle responsible for eyelid opening by down-regulation of FoxL2 in the POMC cells. In this application, we will further characterize KR/TC/R26fN1-ICD triple transgenic mouse as a novel animal model to study the pathogenesis of congenital BPES (Aim1). We will also delineate a molecular pathway that leads to the down-regulation of FoxL2 and subsequent malformation of the levator M¿ellersmooth muscle during embryonic eyelid development (Aim2).