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Abstract：
OBJECTIVE: Axenfeld-Rieger syndrome (ARS) is an autosomal dominant disorder characterized by ocular anterior segment abnormalities. In the current study, we describe clinical and genetic findings in a Chinese ARS pedigree.
METHODS: An ARS pedigree was recruited and patients were given comprehensive ophthalmic examinations and general physical examinations. DNA from the proband II:2 was used for exome sequencing. Sanger sequencing was utilized to identify and validate PITX2 variations. qPCR and western blotting were performed to detect PITX2 expression in immortalized peripheral blood lymphocytes.
RESULTS: All affected family members showed typical ocular abnormalities, including iris atrophy, corectopia, shallow anterior chamber, complete or partial angle closure, and advanced glaucoma. They also exhibited systemic anomalies, such as microdontia, hypodontia, and redundant periumbilical skin. A heterozygous splice-site variation c.390 + 1G > A in PITX2, which might lead to a truncated PITX2 protein (p.Val131IlefsX127), was found in the proband. Sanger sequencing validated that the variation completely co-segregated with the ARS phenotype within this family and was absent in 100 unrelated controls. Western blotting revealed that the nuclear PITX2 protein was significantly decreased in patients compared with controls. Nonetheless, there was no significant difference in the total PITX2 protein level, consistent with qPCR results showing no alteration in PITX2 mRNA levels in the patient group.
CONCLUSIONS: PITX2 c.390 + 1G > A (p.Val131IlefsX127) was a novel genetic etiology of the ARS pedigree. The mutation leads to decreased nuclear PITX2, indicating lower transcriptional activity.
METHODS: An ARS pedigree was recruited and patients were given comprehensive ophthalmic examinations and general physical examinations. DNA from the proband II:2 was used for exome sequencing. Sanger sequencing was utilized to identify and validate PITX2 variations. qPCR and western blotting were performed to detect PITX2 expression in immortalized peripheral blood lymphocytes.
RESULTS: All affected family members showed typical ocular abnormalities, including iris atrophy, corectopia, shallow anterior chamber, complete or partial angle closure, and advanced glaucoma. They also exhibited systemic anomalies, such as microdontia, hypodontia, and redundant periumbilical skin. A heterozygous splice-site variation c.390 + 1G > A in PITX2, which might lead to a truncated PITX2 protein (p.Val131IlefsX127), was found in the proband. Sanger sequencing validated that the variation completely co-segregated with the ARS phenotype within this family and was absent in 100 unrelated controls. Western blotting revealed that the nuclear PITX2 protein was significantly decreased in patients compared with controls. Nonetheless, there was no significant difference in the total PITX2 protein level, consistent with qPCR results showing no alteration in PITX2 mRNA levels in the patient group.
CONCLUSIONS: PITX2 c.390 + 1G > A (p.Val131IlefsX127) was a novel genetic etiology of the ARS pedigree. The mutation leads to decreased nuclear PITX2, indicating lower transcriptional activity.
摘要：
目的：Axenfeld-Rieger综合征（ARS）是一种以眼眼前节异常为特征的常染色体显性遗传病。在目前的研究中,我们描述了中国ARS家系的临床和遗传发现。
方法：招募ARS家系，对患者进行全面的眼科检查和一般体检。来自先证者II:2的DNA用于外显子组测序。利用Sanger测序来鉴定和验证PITX2变异。进行qPCR和蛋白质印迹以检测永生化外周血淋巴细胞中的PITX2表达。
结果：所有受影响的家庭成员均表现出典型的眼部异常，包括虹膜萎缩,阴形目,浅前房,完全或部分角度闭合，和晚期青光眼。他们还表现出系统异常，比如microdontia,缺省症,和多余的脐周皮肤。PITX2中的杂合剪接位点变异c.3901G>A，这可能导致截短的PITX2蛋白（p。Val131IlefsX127),是在先证者中发现的.Sanger测序验证了该变异与该家族中的ARS表型完全共分离，并且在100个无关的对照中不存在。Western印迹显示，与对照组相比，患者的核PITX2蛋白显着降低。尽管如此，PITX2总蛋白水平无显著差异,与qPCR结果一致，显示患者组中PITX2mRNA水平没有变化。
结论：PITX2c.390+1G>A（p。Val131IlefsX127）是ARS家系的新遗传病因。突变导致核PITX2降低,表明较低的转录活性。
方法：招募ARS家系，对患者进行全面的眼科检查和一般体检。来自先证者II:2的DNA用于外显子组测序。利用Sanger测序来鉴定和验证PITX2变异。进行qPCR和蛋白质印迹以检测永生化外周血淋巴细胞中的PITX2表达。
结果：所有受影响的家庭成员均表现出典型的眼部异常，包括虹膜萎缩,阴形目,浅前房,完全或部分角度闭合，和晚期青光眼。他们还表现出系统异常，比如microdontia,缺省症,和多余的脐周皮肤。PITX2中的杂合剪接位点变异c.3901G>A，这可能导致截短的PITX2蛋白（p。Val131IlefsX127),是在先证者中发现的.Sanger测序验证了该变异与该家族中的ARS表型完全共分离，并且在100个无关的对照中不存在。Western印迹显示，与对照组相比，患者的核PITX2蛋白显着降低。尽管如此，PITX2总蛋白水平无显著差异,与qPCR结果一致，显示患者组中PITX2mRNA水平没有变化。
结论：PITX2c.390+1G>A（p。Val131IlefsX127）是ARS家系的新遗传病因。突变导致核PITX2降低,表明较低的转录活性。
