Abstract：
OBJECTIVE: To explore the molecular mechanisms in lens development and the pathogenesis of Peters anomaly in Smad4 defective mice.
METHODS: Le-Cre transgenic mouse line was employed to inactivate Smad4 in the surface ectoderm selectively. Pathological techniques were used to reveal the morphological changes of the anterior segment in Smad4 defective eye. Immunohistochemical staining was employed to observe the expression of E-cadherin, N-cadherin and α-SMA in anterior segment of Smad4 defective mice and control mice at embryonic (E) day 16.5. Real-time quantitative polymerase chain reaction (qPCR) was performed to detect the expression of Snail, Zeb1, Zeb2 and Twist2 in lens of Smad4 defective mice and control mice at E16.5. Statistical evaluations were performed using the unpaired Student\'s t-test (two-tailed) by SPSS 11.0 software.
RESULTS: Conditional deletion of Smad4 on eye surface ectoderm resulted in corneal dysplasia, iridocorneal angle closure, corneolenticular adhesions and cataract resembling Peters anomaly. Loss of Smad4 function inhibited E-cadherin expression in the lens epithelium cells and corneal epithelium cells in Smad4 defective eye. Expression of N-cadherin was up-regulated in corneal epithelium and corneal stroma. Both E-cadherin and N-cadherin were down-regulated at the future trabecular meshwork region in mutant eye. The qPCR results showed that the expression of Twist2 was increased significantly in the mutant lens (P<0.01).
CONCLUSIONS: Smad4 is essential to eye development and likely a candidate pathogenic gene to Peters anomaly by regulating epithelial-mesenchymal transition. Twist2 can be regulated by Smad4 and plays an essential role in lens development.
METHODS: Le-Cre transgenic mouse line was employed to inactivate Smad4 in the surface ectoderm selectively. Pathological techniques were used to reveal the morphological changes of the anterior segment in Smad4 defective eye. Immunohistochemical staining was employed to observe the expression of E-cadherin, N-cadherin and α-SMA in anterior segment of Smad4 defective mice and control mice at embryonic (E) day 16.5. Real-time quantitative polymerase chain reaction (qPCR) was performed to detect the expression of Snail, Zeb1, Zeb2 and Twist2 in lens of Smad4 defective mice and control mice at E16.5. Statistical evaluations were performed using the unpaired Student\'s t-test (two-tailed) by SPSS 11.0 software.
RESULTS: Conditional deletion of Smad4 on eye surface ectoderm resulted in corneal dysplasia, iridocorneal angle closure, corneolenticular adhesions and cataract resembling Peters anomaly. Loss of Smad4 function inhibited E-cadherin expression in the lens epithelium cells and corneal epithelium cells in Smad4 defective eye. Expression of N-cadherin was up-regulated in corneal epithelium and corneal stroma. Both E-cadherin and N-cadherin were down-regulated at the future trabecular meshwork region in mutant eye. The qPCR results showed that the expression of Twist2 was increased significantly in the mutant lens (P<0.01).
CONCLUSIONS: Smad4 is essential to eye development and likely a candidate pathogenic gene to Peters anomaly by regulating epithelial-mesenchymal transition. Twist2 can be regulated by Smad4 and plays an essential role in lens development.
摘要：
目的：探讨Smad4缺陷小鼠晶状体发育的分子机制和Peters异常的发病机制。
方法:使用Le-Cre转基因小鼠系选择性灭活表面外胚层中的Smad4。采用病理学技术揭示Smad4缺陷眼眼前节的形态学变化。免疫组化染色观察E-cadherin的表达,在胚胎（E）第16.5天，Smad4缺陷小鼠和对照小鼠的前段中的N-钙黏着蛋白和α-SMA。实时定量聚合酶链反应(qPCR)检测Snail的表达,E16.5时Smad4缺陷小鼠和对照小鼠晶状体中的Zeb1，Zeb2和Twist2。通过SPSS11.0软件使用未配对的Studenti'st检验(双尾)进行统计学评价。
结果：眼表外胚层上Smad4的条件性缺失导致角膜发育不良，虹膜角膜角闭合，角膜豆状粘连和白内障类似于彼得斯异常。Smad4功能的丧失抑制了Smad4缺陷眼的晶状体上皮细胞和角膜上皮细胞中E-cadherin的表达。角膜上皮和角膜基质中N-cadherin的表达上调。E-cadherin和N-cadherin在突变眼的未来小梁网区均下调。qPCR结果显示,突变型晶状体中Twist2的表达显著增加(P<0.01)。
结论：Smad4是眼睛发育所必需的，并且可能是通过调节上皮-间质转化导致Peters异常的候选致病基因。Twist2可以由Smad4调节，在晶状体发育中起着至关重要的作用。
方法:使用Le-Cre转基因小鼠系选择性灭活表面外胚层中的Smad4。采用病理学技术揭示Smad4缺陷眼眼前节的形态学变化。免疫组化染色观察E-cadherin的表达,在胚胎（E）第16.5天，Smad4缺陷小鼠和对照小鼠的前段中的N-钙黏着蛋白和α-SMA。实时定量聚合酶链反应(qPCR)检测Snail的表达,E16.5时Smad4缺陷小鼠和对照小鼠晶状体中的Zeb1，Zeb2和Twist2。通过SPSS11.0软件使用未配对的Studenti'st检验(双尾)进行统计学评价。
结果：眼表外胚层上Smad4的条件性缺失导致角膜发育不良，虹膜角膜角闭合，角膜豆状粘连和白内障类似于彼得斯异常。Smad4功能的丧失抑制了Smad4缺陷眼的晶状体上皮细胞和角膜上皮细胞中E-cadherin的表达。角膜上皮和角膜基质中N-cadherin的表达上调。E-cadherin和N-cadherin在突变眼的未来小梁网区均下调。qPCR结果显示,突变型晶状体中Twist2的表达显著增加(P<0.01)。
结论：Smad4是眼睛发育所必需的，并且可能是通过调节上皮-间质转化导致Peters异常的候选致病基因。Twist2可以由Smad4调节，在晶状体发育中起着至关重要的作用。
