Functional Analysis of Novel Genes in Eye Development and Vision

眼睛发育和视力新基因的功能分析

• 批准号: 10930507
• 负责人: graeme j wistow
• 金额: $ 161.67万
• 依托单位: NATIONAL EYE INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10930507
• 关键词: Age related macular degeneration; Animal Model; Behavior; Bioinformatics; Blindness; Bruch' s basal membrane structure; Calcium; Cell Culture System; Cell Culture Techniques; Cell model; Cells; Cholesterol; Collaborations; Deposition; Disease; Eye; Eye Development; FDA approved; Gene Expression Profiling; Genes; Genomics; Human; Hydroxyapatites; Methodology; Modeling; Morphology; Mus; Nonexudative age-related macular degeneration; Pharmaceutical Preparations; Process; Proteins; RPE65 protein; Serum; Small Interfering RNA; Structure; Structure of retinal pigment epithelium; System; Testing; Therapeutic Uses; Tissues; Transgenic Mice; Up-Regulation; Vision; Work; access restrictions; calcification; capillary bed; deprivation; functional genomics; gene product; human model; insight; knock-down; mineralization; molecular modeling; mouse model; novel; patient population; wound

Age-related macular degeneration (AMD) is a major cause of vision loss. We have developed a cell-culture model for serum-deprivation AMD using RPE-derived cells. In AMD, changes at Bruch's membrane and in the capillary bed are likely to restrict access of serum components to the RPE. We have shown that serum deprivation of RPE cells leads to a marked upregulation of cholesterol synthesis and transport and the accumulation of cholesterol in the RPE. This is strongly reminiscent of the accumulation of cholesterol RPE that we and others have seen in human AMD. Analysis of gene expression in the cell model has led to the discovery of the induction of amelotin (AMTN), a protein of calcium/hydroxyapatite (HAP) mineralization. In cell culture AMTN is responsible for HAP deposition and siRNA knockdown of AMTN inhibits calcification. Furthermore, we showed that AMTN is expressed in human donor eyes with dry AMD and is associated with HAP spherules. We have also developed a transgenic mouse model which specifically targets expression of human AMTN to retinal pigment epithelium using a novel cassette based on the RPE65 gene. This efficiently and specifically expresses human AMTN in mouse RPE and causes morphological changes in RPE and Bruch's membrane similar to those seen in AMD. We have now found an experimental wound system that induces HAP/AMTN deposits in RPE in mice. This is being used to test the effects of siRNA inhibition of AMTN for potential therapeutic use. We gave identified several FDA approved drugs that inhibit AMTN function and will be tested as potential therapies to slow AMD progression. We are also collaborating to identify any effects of these drugs on AMD progression in patient populations. We have initiated collaborative structural studies on AMTM. The protein is intrinsically disordered and presents opportunities for new methodological approaches. Our collaborators have already provided new insights into the structure and behavior of AMTN with important implications for work on similar proteins. Combined with molecular modelling, this is giving hints of possible functional mechanisms,

与年龄相关的黄斑变性（AMD）是视力丧失的主要原因。我们已经开发了一种使用RPE衍生的细胞的细胞培养模型，用于血清剥夺AMD。在AMD中，BRUCH的膜和毛细管床的变化可能会限制血清成分进入RPE。我们已经表明，RPE细胞的血清剥夺导致胆固醇合成和转运的明显上调以及胆固醇在RPE中的积累。这强烈让人联想到我们和其他人在人类AMD中看到的胆固醇RPE的积累。细胞模型中基因表达的分析导致发现诱导蛋白质（AMTN），Amelotin（AMTN）是一种钙/羟基磷灰石（HAP）矿化的蛋白质。在细胞培养中，AMTN负责HAP沉积和AMTN的siRNA敲低抑制钙化。此外，我们表明AMTN在干燥AMD的人类供体眼中表达，并且与HAP球形有关。 我们还开发了一种转基因小鼠模型，该模型专门针对人类AMTN对视网膜色素上皮的表达，使用基于RPE65基因的新型盒式盒子。这有效地表达了小鼠RPE中的人类AMTN，并在RPE和BRUCH的膜中引起形态变化，类似于AMD中的膜。 现在，我们发现了一个实验性伤口系统，该系统诱导小鼠RPE中的HAP/AMTN沉积物。这用于测试siRNA抑制AMTN对潜在治疗使用的影响。 我们给出了几种FDA批准的药物，这些药物抑制AMTN功能，并将被视为缓慢AMD进展的潜在疗法。我们还正在合作确定这些药物对患者人群AMD进展的任何影响。 我们已经在AMTM上启动了协作结构研究。该蛋白质本质上是无序的，并为新的方法论方法提供了机会。我们的合作者已经为AMTN的结构和行为提供了新的见解，对类似蛋白质的工作具有重要意义。结合分子建模，这给出了可能的功能机制的提示

项目成果

The klotho-related protein KLPH (lctl) has preferred expression in lens and is essential for expression of clic5 and normal lens suture formation.

• DOI: 10.1016/j.exer.2018.02.001
• 发表时间: 2018-04
• 期刊: Experimental eye research
• 影响因子: 3.4
• 作者: Fan J;Lerner J;Wyatt MK;Cai P;Peterson K;Dong L;Wistow G
• 通讯作者: Wistow G

Molecular studies into cell biological role of Copine-4 in Retinal Ganglion Cells.

• DOI: 10.1371/journal.pone.0255860
• 发表时间: 2021
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Goel M;Aponte AM;Wistow G;Badea TC
• 通讯作者: Badea TC

Retbindin mediates light-damage in mouse retina while its absence leads to premature retinal aging.

• DOI: 10.1016/j.exer.2021.108698
• 发表时间: 2021-08
• 期刊: Experimental eye research
• 影响因子: 3.4
• 作者: Fan J;Rajapakse D;Peterson K;Lerner J;Parsa S;Ponduri A;Sagar V;Duncan T;Dong L;Wistow G
• 通讯作者: Wistow G

Interaction of complement factor h and fibulin3 in age-related macular degeneration.

• DOI: 10.1371/journal.pone.0068088
• 发表时间: 2013
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Wyatt MK;Tsai JY;Mishra S;Campos M;Jaworski C;Fariss RN;Bernstein SL;Wistow G
• 通讯作者: Wistow G

Amelotin is expressed in retinal pigment epithelium and localizes to hydroxyapatite deposits in dry age-related macular degeneration.

Amelotin 在视网膜色素上皮中表达，并定位于干性年龄相关性黄斑变性中的羟基磷灰石沉积物。

• DOI: 10.1016/j.trsl.2020.02.007
• 发表时间: 2020
• 期刊: Translational research : the journal of laboratory and clinical medicine
• 作者: Rajapakse,Dinusha;Peterson,Katherine;Mishra,Sanghamitra;Fan,Jianguo;Lerner,Joshua;Campos,Maria;Wistow,Graeme
• 通讯作者: Wistow,Graeme