Anopthalmia Spectrum Disorders

描述(由申请人提供)：每5,000至10,000人中就有1人患上无眼症，由于由此导致的视力障碍，这是一种毁灭性的先天缺陷。眼球无力与小眼球(小眼睛)和缺损(脉络膜裂隙或视裂未能闭合)密切相关。我们研究了一名患有严重小眼症的男性患者，他在染色体18q22.1上有一种新的染色体缺失。我们发现18q22.1处缺失的基因之一TXNDC10与眼睛发育有关。原位杂交结果显示，Txndc10在发育中的小鼠视网膜神经上皮和晶状体上皮细胞中均有表达。我们对66例无眼症或小眼球症患者的TXNDC10进行了重新测序，发现了c.260G&gt；A，预示着在一名无关的小眼炎和视网膜缺损患者中存在错义替换p.R39Q。我们使用针对斑马鱼Txndc10同源基因Zgc110025的反义吗啉来显示斑马鱼Txndc10同源基因Zgc110025的形态幼虫存在小眼炎和缺陷症。Pax2原位杂交显示，与对照组相比，脉络膜裂隙染色在变形者中增加，提示Pax2失调参与了缺损的形成。形态视网膜周边抗组蛋白H3染色增强，我们在形态视网膜观察到较少的晚期分化细胞类型。人野生型TXNDC10mRNA与吗啡共注射可挽救变形表型，而人TXNDC10/(p.R39Q)突变型mRNA与吗啡共注射则不能。我们的结果表明，TXNDC10的单倍性不足干扰了眼睛的发育。我们建议从两个方向继续我们对TXNDC10的研究。我们假设Txndc10可能是视网膜腹侧发育的已知途径的一部分，并希望利用斑马鱼来确定Zgc110025是否与其他参与脉络膜裂隙闭合和视网膜腹侧形成的已知基因相互作用，如Vax1、Vax2和Shh(Syu)。我们将使用原位杂交、双次注射反义吗啉和mRNA挽救实验来实现这一目标。我们的目的是试图揭示Txndc10影响眼睛发育的机制。其次，我们建议建立Txndc10功能缺失的小鼠模型，以检测在基因缺失模型中的眼睛表型，而不是在只有部分基因功能丧失的动物模型中。我们假设Txndc10功能的丧失会导致小鼠患上小眼炎和缺陷症，我们将使用得到的敲除小鼠来检查Txndc10缺失动物的眼睛形态，并进行表达阵列来评估Txndc10缺失小鼠和野生型小鼠之间的基因表达差异。我们的最终目标是提高对视网膜腹侧形成和脉络膜裂隙闭合所需基因的现有知识，以便为患有无眼症和相关眼缺陷的人类患者提供更好的遗传咨询和测试。 与公共卫生相关：据估计，出生缺陷每年影响美国出生的12万名婴儿(每33人中就有1人)，是出生后第一年死亡的主要病例。由于视力下降的医疗和社会影响，无眼球(无眼)、小眼球(小眼睛)和缺陷症(脉络膜裂隙或视裂未能闭合)会给受影响的个人及其家庭造成重大负担。这项应用旨在改善与这些眼睛缺陷的遗传原因有关的现有知识，从而改善患者的咨询、管理和治疗。

DESCRIPTION (provided by applicant): Anophthalmia, or absence of the eye, is found in 1 in 5,000 to 10,000 individuals and is a devastating birth defect because of the resulting visual impairment. Anopththalmia is closely related to microphthalmia (small eyes) and coloboma (failure of the choroid fissure or the optic fissure to close). We have studied a male patient with severe microphthalmia who had a novel chromosome deletion at chromosome 18q22.1. We have implicated one of the deleted genes at 18q22.1, TXNDC10, in eye development. In situ hybridization showed that Txndc10 was expressed in the retinal neuroepithelium and lens epithelium in the developing murine eye. We re-sequenced TXNDC10 in 66 patients with anophthalmia or microphthalmia, and found c.260G>A, predicting a missense substitution, p.R39Q, in an unrelated patient with microphthalmia and retinal coloboma. We used antisense morpholinos targeted against the zebrafish Txndc10 orthologue, Zgc110025, to show that the morphant larvae had microphthalmia and coloboma. In-situ hybridization with Pax2 showed increased choroid fissure staining in morphants compared to controls, suggesting that Pax2 dysregulation was involved in the coloboma formation. The morphant retinas had increased anti-histone H3 staining in the peripheral retina and we observed fewer late-differentiating cell types in the morphant retinas. Co-injection of human wild type TXNDC10 mRNA with a morpholino rescued the morphant phenotype, whereas co-injection of human TXNDC10/(p.R39Q) mutant mRNA with morpholino did not. Our results suggest that haploinsufficiency for TXNDC10 perturbs eye development. We propose to continue our research on TXNDC10 in two directions. We hypothesize that Txndc10 may be part of a known pathway for ventral retinal development, and would like to use zebrafish to determine whether Zgc110025 interacts with other known genes that are involved in choroid fissure closure and the formation of the ventral retina, such as Vax1, Vax2 and Shh (Syu). We will use in-situ hybridization, dual injections of antisense morpholinos and mRNA rescue experiments to accomplish this aim. Our purpose is to try to uncover the mechanism whereby Txndc10 affects eye development. Secondly, we propose to make a mouse model of loss of Txndc10 function to examine the eye phenotype in a genetic null model rather than in an animal model with only partial loss of gene function. We hypothesize that loss of Txndc10 function will produce mice with microphthalmia and coloboma, and we will use the resultant knock-out mice to examine eye morphology in a Txndc10 null animal and to perform expression arrays to evaluate for differences in gene expression between Txndc10 null mice and wildtype litter mates. Our ultimate goal is to improve the existing knowledge about the genes that are required for ventral retina formation and choroid fissure closure so that human patients with anophthalmia and related eye defects can be offered improved genetic counseling and testing. PUBLIC HEALTH RELEVANCE: Birth defects affect an estimated 120,000 (1 in 33) babies born in the United States each year, and are the leading case of mortality in the first year of life. Anophthalmia (absent eye), microphthalmia (small eye) and coloboma (failure of the choroid fissure or the optic fissure to close) can result in a significant burden for affected individuals and their families because of the medical and social effects of reduced vision. This application seeks to improve the existing knowledge pertaining to the genetic causes of these eye defects so that patient counseling, management and treatment can be improved.