Endothelial Transmigration in Neovascular Age-related Macular Degeneration

新生血管性年龄相关性黄斑变性中的内皮细胞迁移

基本信息

  • 批准号:
    10379608
  • 负责人:
  • 金额:
    $ 43.73万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2007
  • 资助国家:
    美国
  • 起止时间:
    2007-04-01 至 2023-03-31
  • 项目状态:
    已结题

项目摘要

Anti-VEGF therapies, standard care in neovascular age-related macular degeneration (nvAMD), improve outcomes in less than 50% of patients and do not prevent vision-loss progression due to fibrosis or macular atrophy. We used human physiologically relevant models to gain understanding into the coordination of signaling pathways and cross-talk involved in the activation of choroidal endothelial cells (CECs) to migrate and form macular neovascularization in nvAMD. The scaffolding protein, IQGAP1, sustains activation of the GTPase Rac1, which is necessary for CEC migration. Rac1 is activated by AMD-related stresses involving inflammatory, oxidative and angiogenic factors, as well as by the oxysterol, 7-ketocholesterol (7KC), which accumulates in blood and Bruch’s membrane with increased age and in AMD. 7KC causes CECs to change expression of cell markers from endothelial to mesenchymal ones, suggesting endothelial-mesenchymal transition (EndMT). IQGAP1 appears involved. 7KC also causes fibrosis in models of laser induced injury. Our data support the hypothetical framework that will be tested in the next funding period: that (1) IQGAP1 is critical to EndMT induced by the oxysterol 7KC; and that (2) 7KC triggers transcriptional events that render CECs unable to maintain expression of endothelial markers but to develop into a new phenotype of migratory, mesenchymal cells that develop into fibrosis. We will also test two potential therapies to reduce fibrosis: (a) to inhibit TGFβ signaling in combination with anti-VEGF and (b) to target phosphorylation of IQGAP1 in a novel mutant IQGAP1 mouse that we created by CRISPR-Cas9-induced gene mutation. Specific Aim 1 is to test the prediction that IQGAP1 mediates EndMT-induced migration in CECs exposed to 7KC. Specific Aim 2 is to test the prediction that 7KC, mediated through IQGAP1, decreases the proportion of labeled endothelial positive to mesenchymal positive cells after laser in endothelial specific yellow-fluorescent protein reporter mice. Specific Aim 3 is to test predictions that increased age, TGFβ-signaling, or IQGAP1 serine phosphorylation will increase αSMA-labeled lesions after laser in 7KC-treated eyes and to test strategies as possible future treatments. We will also evaluate the involvement of Müller cells, pericyes, and RPE. Tools include isolated human CECs; high throughput RNA sequencing; flow cytometry; spectral domain optical coherence tomography (sdOCT); 7KC-induced models of EndMT and fibrosis; yellow-fluorescent protein endothelial reporter mice; conditional inducible endothelial Iqgap1 knockout mice; a mutant IQGAP1 mouse through CRISPR-Cas9-technology; intravitreal injections of pharmacologic agents; Micron IV laser induced injury to test 7KC-induced EndMT and lesion formation. These studies will test the role of IQGAP1 in 7KC-induced EndMT as a potential cause of fibrosis, which is poorly responsive to anti-VEGF in nvAMD, and will test two novel treatments to reduce nvAMD and EndMT.
抗 VEGF 疗法、新生血管性年龄相关性黄斑变性 (nvAMD) 的标准护理、改善 不到 50% 的患者有结果,并且不能防止纤维化或黄斑导致的视力丧失进展 萎缩。我们使用人类生理相关模型来了解 参与脉络膜内皮细胞 (CEC) 迁移激活的信号通路和串扰 并在 nvAMD 中形成黄斑新生血管。支架蛋白 IQGAP1 维持 GTPase Rac1,这是 CEC 迁移所必需的。 Rac1 被 AMD 相关应激激活,包括 炎症、氧化和血管生成因子,以及氧甾醇、7-酮胆固醇 (7KC), 随着年龄的增长和AMD,它会在血液和布鲁赫膜中积累。 7KC 导致 CEC 发生变化 细胞标志物从内皮到间质的表达,表明内皮-间质 过渡(EndMT)。 IQGAP1 似乎参与其中。 7KC 还会导致激光损伤模型中的纤维化。我们的 数据支持将在下一个资助期测试的假设框架: (1) IQGAP1 是 对氧甾醇 7KC 诱导的 EndMT 至关重要; (2) 7KC 触发转录事件,从而呈现 CEC 无法维持内皮标志物的表达,而是发展成一种新的迁移表型, 间充质细胞发展成纤维化。我们还将测试两种减少纤维化的潜在疗法:(a) 与抗 VEGF 药物联合抑制 TGFβ 信号转导,并 (b) 在一种新型药物中靶向 IQGAP1 的磷酸化 我们通过 CRISPR-Cas9 诱导基因突变创造出突变型 IQGAP1 小鼠。具体目标 1 是测试 预测 IQGAP1 介导暴露于 7KC 的 CEC 中 EndMT 诱导的迁移。具体目标 2 是测试 预测 7KC 通过 IQGAP1 介导,降低标记内皮细胞阳性的比例 内皮特异性黄色荧光蛋白报告小鼠激光照射后的间充质阳性细胞。具体的 目标 3 是检验年龄增长、TGFβ 信号传导或 IQGAP1 丝氨酸磷酸化将影响的预测。 7KC 治疗眼睛激光后 αSMA 标记病变增加,并测试未来可能的策略 治疗。我们还将评估 Müller 细胞、周眼细胞和 RPE 的参与情况。工具包括隔离的 人类 CEC;高通量RNA测序;流式细胞术;谱域光学相干性 断层扫描(sdOCT); 7KC诱导的EndMT和纤维化模型;黄色荧光内皮蛋白 记者鼠;条件诱导型内皮Iqgap1基因敲除小鼠; IQGAP1 突变小鼠 CRISPR-Cas9 技术;玻璃体内注射药剂; Micron IV 激光诱导损伤测试 7KC 诱导 EndMT 和病变形成。这些研究将测试 IQGAP1 在 7KC 诱导的 EndMT 中的作用 作为纤维化的一个潜在原因,纤维化对 nvAMD 中的抗 VEGF 反应较差,并将测试两种新的 减少 nvAMD 和 EndMT 的治疗。

项目成果

期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(1)

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Mary Elizabeth Ruth Hartnett其他文献

Mary Elizabeth Ruth Hartnett的其他文献

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{{ truncateString('Mary Elizabeth Ruth Hartnett', 18)}}的其他基金

Inhibiting Neovascularization and Subretinal Fibrosis in Neovascular Age-Related Macular Degeneration
抑制新生血管性年龄相关性黄斑变性的新生血管形成和视网膜下纤维化
  • 批准号:
    10639785
  • 财政年份:
    2023
  • 资助金额:
    $ 43.73万
  • 项目类别:
Medical Student Research Program in Eye Health and Disease
眼健康与疾病医学生研究计划
  • 批准号:
    9073790
  • 财政年份:
    2016
  • 资助金额:
    $ 43.73万
  • 项目类别:
Endothelial Transmigration in Neovascular Age-related Macular Degeneration
新生血管性年龄相关性黄斑变性中的内皮细胞迁移
  • 批准号:
    8035291
  • 财政年份:
    2007
  • 资助金额:
    $ 43.73万
  • 项目类别:
Endothelial Transmigration in Neovascular Age-related Macular Degeneration
新生血管性年龄相关性黄斑变性中的内皮细胞迁移
  • 批准号:
    7253703
  • 财政年份:
    2007
  • 资助金额:
    $ 43.73万
  • 项目类别:
Endothelial Transmigration in Neovascular Age-related Macular Degeneration
新生血管性年龄相关性黄斑变性中的内皮细胞迁移
  • 批准号:
    7389477
  • 财政年份:
    2007
  • 资助金额:
    $ 43.73万
  • 项目类别:
Endothelial Transmigration in Neovascular Age-related Macular Degeneration
新生血管性年龄相关性黄斑变性中的内皮细胞迁移
  • 批准号:
    8451297
  • 财政年份:
    2007
  • 资助金额:
    $ 43.73万
  • 项目类别:
Endothelial Transmigration in Neovascular Age-related Macular Degeneration
新生血管性年龄相关性黄斑变性中的内皮细胞迁移
  • 批准号:
    8088864
  • 财政年份:
    2007
  • 资助金额:
    $ 43.73万
  • 项目类别:
Endothelial Transmigration in Neovascular Age-related Macular Degeneration
新生血管性年龄相关性黄斑变性中的内皮细胞迁移
  • 批准号:
    7777266
  • 财政年份:
    2007
  • 资助金额:
    $ 43.73万
  • 项目类别:
Endothelial Transmigration in Neovascular Age-related Macular Degeneration
新生血管性年龄相关性黄斑变性中的内皮细胞迁移
  • 批准号:
    7582299
  • 财政年份:
    2007
  • 资助金额:
    $ 43.73万
  • 项目类别:
Endothelial Transmigration in Neovascular Age-related Macular Degeneration
新生血管性年龄相关性黄斑变性中的内皮细胞迁移
  • 批准号:
    10752738
  • 财政年份:
    2007
  • 资助金额:
    $ 43.73万
  • 项目类别:
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