Mechanism of Cone Degeneration Resulting from CNG Channel Deficiency

CNG通道缺陷导致锥体退化的机制

• 批准号: 8485614
• 负责人: XI-QIN DING
• 金额: $ 33.74万
• 依托单位: UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8485614
• 关键词: Accounting; Apoptosis; Blindness; Cell Death; Cellular biology; Cessation of life; Color Visions; Complex; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Defect; Development; Disease; Experimental Designs; Gene Expression; Generations; Genes; Genetic; Goals; Guanylate Cyclase; Hand; In Situ Nick-End Labeling; Inherited; Knock-out; Knowledge; Label; Longevity; Mammals; Methods; Microarray Analysis; Modeling; Mus; Mutation; Normal Cell; Pathology; Patients; Photoreceptors; Phototransduction; Population; Proteins; RNA Interference; Retina; Retinal; Retinal Cone; Role; Technology; Testing; Therapeutic Intervention; Vision Disorders; Visual Acuity; achromatopsia; cGMP production; cyclic-nucleotide gated ion channels; cytotoxic; density; gene replacement therapy; improved; inhibitor/antagonist; insight; mouse model; prevent; public health relevance; research study; response; retinal rods; therapeutic development; tool; treatment strategy

DESCRIPTION (provided by applicant): The cone cyclic nucleotide-gated (CNG) channel is essential for central and color vision and for visual acuity. Mutations in the two channel subunits, CNGA3 and CNGB3, are associated with achromatopsia and progressive cone dystrophy. Cones degenerate in mouse models of CNGA3 and CNGB3 deficiency and in patients with achromatopsia or cone dystrophy. There are currently no curative treatments for these diseases, in part due to a lack of understanding of the mechanisms underlying the disease pathobiology. The overarching objective of this study is to understand the mechanism(s) of cone degeneration resulting from loss of CNG channel function. Specifically, we will determine whether the accumulation of cGMP in models of CNG channel deficiency causes cone degeneration. We will perform the proposed studies using the CNGA3-/-/Nrl-/- and CNGB3-/-/Nrl-/- mouse lines. Using these mouse models provides a unique opportunity to study cone degeneration in a retina that contains only cones. Three specific aims are proposed. The first specific aim is to determine whether the accumulation of cGMP is responsible for cone degeneration in CNGA3 deficiency. We will test the ability of guanylate cyclase (GC) inhibitors to reduce cone degeneration in the channel-deficient mice. In a corroborating experiment, we will also test this hypothesis by abolishing expression of retGC-1 (retinal guanlyate cyclase) in channel deficient cones by utilizing RNAi and/or genetic methods (generation of CNGA3-/-/Nrl-/-/retGC-1-/-). The second specific aim is to determine how cones die in CNGA3 deficiency. If cGMP is responsible for cone cell death, we will examine how the accumulation of cGMP leads to cone cell death. We will investigate the role of cGMP- dependent protein kinase (PKG) in cone degeneration. We will determine whether PKG activity is higher in the CNGA3-/-/Nrl-/- retina compared to the Nrl-/- retina, and will test whether the use of PKG inhibitors/knockdown of PKG expression can ameliorate cone degeneration. Alternate mechanisms for cone degeneration will also be investigated by studying altered gene expression in channel deficient mice (compared to WT) by using microarray analysis. The third specific aim is to determine whether CNGB3 deficiency leads to an accumulation of cGMP in cones by using CNGB3-/-/Nrl-/- mice. Upon completion of the proposed study we will have a better understanding of the mechanism(s) underlying cone degeneration resulting from CNG channel deficiency. Improving the lifespan of degenerating cones is necessary for successful treatment of inherited cone diseases. The knowledge gained in this study is crucial for the development of therapeutic strategies to prevent or retard this degeneration, both in cases in which the causative gene is one of the channel proteins and in more general cases of cone dystrophy.

描述（由申请人提供）：锥状环核苷酸门控（CNG）通道对于中央视觉和色觉以及视敏度至关重要。 CNGA3 和 CNGB3 这两个通道亚基的突变与全色盲和进行性视锥细胞营养不良有关。 CNGA3 和 CNGB3 缺陷的小鼠模型以及全色盲或视锥细胞营养不良的患者中，视锥细胞发生退化。目前还没有针对这些疾病的治疗方法，部分原因是缺乏对疾病病理学机制的了解。本研究的首要目标是了解 CNG 通道功能丧失导致视锥细胞退化的机制。具体来说，我们将确定 CNG 通道缺陷模型中 cGMP 的积累是否会导致视锥细胞变性。我们将使用 CNGA3-/-/Nrl-/- 和 CNGB3-/-/Nrl-/- 小鼠系进行拟议的研究。使用这些小鼠模型提供了一个独特的机会来研究仅包含视锥细胞的视网膜中的视锥细胞变性。提出了三个具体目标。第一个具体目标是确定 cGMP 的积累是否是 CNGA3 缺陷中视锥细胞变性的原因。我们将测试鸟苷酸环化酶（GC）抑制剂减少通道缺陷小鼠视锥变性的能力。在确证实验中，我们还将通过利用 RNAi 和/或遗传方法（生成 CNGA3-/-/Nrl-/-/retGC-1-/-）消除通道缺陷视锥细胞中 retGC-1（视网膜鸟苷酸环化酶）的表达来检验这一假设。第二个具体目标是确定 CNGA3 缺乏时视锥细胞如何死亡。如果 cGMP 是导致视锥细胞死亡的原因，我们将研究 cGMP 的积累如何导致视锥细胞死亡。我们将研究 cGMP 依赖性蛋白激酶 (PKG) 在视锥细胞变性中的作用。我们将确定与 Nrl-/- 视网膜相比，CNGA3-/-/Nrl-/- 视网膜中的 PKG 活性是否更高，并将测试使用 PKG 抑制剂/敲低 PKG 表达是否可以改善视锥细胞变性。还将通过使用微阵列分析研究通道缺陷小鼠（与野生型小鼠相比）中基因表达的改变来研究视锥细胞变性的替代机制。第三个具体目标是通过使用 CNGB3-/-/Nrl-/- 小鼠来确定 CNGB3 缺陷是否会导致视锥细胞中 cGMP 的积累。完成拟议的研究后，我们将对 CNG 通道缺陷导致的视锥细胞退化的机制有更好的了解。延长退化视锥细胞的寿命对于成功治疗遗传性视锥细胞疾病是必要的。这项研究中获得的知识对于制定预防或延缓这种退化的治疗策略至关重要，无论是在致病基因是通道蛋白之一的情况下，还是在更一般的视锥细胞营养不良的情况下。