Roles of Growth Factors on Corneal Morphogenesis

脊椎动物眼发育过程中角膜、结膜和眼睑的形态发生涉及神经嵴来源的间充质细胞的迁移和表面外胚层细胞的分化。 通过生长因子的双向间充质-上皮相互作用对于发育期间的形态发生和成体的内环境稳定是必不可少的。 TGF-β和FGF- 7在调节神经嵴来源的间充质细胞的功能和眼形态发生过程中外胚层细胞的分化中起关键作用。 这些功能通过如下观察到的严重临床表现来举例说明：在Tgfb 2-/-小鼠中类似于人类的Axene-Reiger异常;通过角质蛋白聚糖启动子（Kerapr）的双糖蛋白聚糖转基因（KeraprBgn/+）小鼠表现出继发性角膜缘缺陷;以及通过导致角膜表面结膜化的α A-晶体启动子（alphaACPR）的FGF-7转基因小鼠。 这些观察提出了有趣的问题，如：1）。 间充质细胞什么时候转化为内皮细胞？2）。眼睑基质细胞也是神经嵴来源的吗？3）。TGF-β和FGF-7信号在眼睛发育、角膜稳态和伤口愈合中的作用是什么？4）。 在KeraprBgn/+转基因小鼠中观察到的表型的分子和细胞机制是什么？ 为了解决这些问题，我们将建立四环素诱导的转基因小鼠模型，过表达II型TGF-β受体的显性负突变体，Kerapr控制下的活性TGF-β 1和FGF-7，以及alphaCApr控制下的FGF-7。 通过这些模型，我们将研究TGF-β信号传导在发育和稳态过程中对角膜形态发生的作用，并进一步表征KeraprBgn/+小鼠，以研究过量双糖链蛋白聚糖在眼睑形态发生过程中干扰TGF-β信号传导的假设。具体目的1是使用tet-O TbetaRIIdn/+ KeraprrtTA/+和tet-O TGFbeta 1/+ KeraprrtTA/+小鼠阐明TGF-β信号传导在发育、稳态和伤口愈合期间对眼形态发生的作用。 具体目标2将使用过表达FGF-7的alphaACPrrtTA/+ tet-OFGF 7/+和KeraprrtTA/+ tet-OFGF 7/+小鼠阐明眼表面上皮分化。 特异性目的3将阐明过量双糖链聚糖对KeraprBgn/+小鼠发育期间眼睑形成的作用。 拟议的研究将为更好地理解TGF-β和FGF-7对角膜发育和体内平衡的作用提供有用的信息，从而设计出更好的角膜疾病治疗方法，例如，上皮清创术、切开术、碱烧伤。

Morphogenesis of cornea, conjunctiva and eyelids during vertebrate eye development involves the migration of mesenchymal cells of neural crest origin and the differentiation of cells of the surface ectoderm. The bi-directional mesenchyme-epithelium interactions via growth factors are essential for morphogenesis during development and homeostasis in adults. TGF-betas and FGF- 7 play pivotal roles in modulating functions of mesenchymal cells of neural crest origin and differentiation of ectoderm cells during ocular morphogenesis. These functions are exemplified by severe clinical manifestations observed as follows: in Tgfb2-/- mice resembling Axenfeld-Reiger's anomaly in human; by biglycan transgenic (KeraprBgn/+) mice of keratocan promoter (Kerapr) exhibiting secondary limbal deficiency; and by FGF-7 transgenic mice of alphaA-crystalline promoter (alphaACpr) resulting in conjunctivalization of corneal surface. These observations raise intriguing questions such as: 1). When do mesenchymal cells commit to becoming endothelial cells? 2). Are eyelid stromal cells also of neural crest origin? 3). What is the role of TGF- beta and FGF-7 signaling in eye development and corneal homeostasis and wound healing? 4). What are the molecular and cellular mechanisms of phenotypes observed in KeraprBgn/+ transgenic mice? To address these questions, we will create tetracycline inducible transgenic mouse models that overexpress the dominant negative mutant of type II TGF-beta receptor, active TGF-beta1 and FGF-7 under the control of Kerapr, and FGF-7 under the control of alphaCApr. With these models we will examine the role of TGF-beta signaling on corneal morphogenesis during development and homeostasis and further characterize the KeraprBgn/+ mice to examine the hypothesis that excess biglycan perturbs TGF-beta signaling during eyelid morphogenesis. Specific Aim 1 is to elucidate roles of TGF-beta signaling on eye morphogenesis during development, homeostasis and wound healing using tet-O TbetaRIIdn/+ KeraprrtTA/+ and tet-O TGFbeta1/+ KeraprrtTA/+ mice. Specific Aim 2 will elucidate ocular surface epithelial differentiation using alphaACprrtTA/+ tet-OFGF7/+ and KeraprrtTA/+ tet-OFGF7/+ mice that overexpress FGF-7. Specific Aim 3 will Elucidate the Role of Excess Biglycan on Eyelid Formation during Development of KeraprBgn/+ mice. The proposed studies will yield useful information for a better understanding of the role of TGF-beta and FGF-7 on corneal development and homeostasis, leading to the design of better treatments for corneal diseases, e.g., epithelial debridement, incision, alkali burn.