MIR182 and Ocular Hypertension.

MIR182 和高眼压。

• 批准号: 10598874
• 负责人: Yutao Liu
• 金额: $ 23.1万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10598874
• 关键词: 3-Dimensional; Achievement; Affect; Anatomy; Anterior; Aqueous Humor; Blindness; CHEK2 gene; Cell Aging; Cell Count; Cell Death; Cell physiology; Cells; Clinic; Clinical Treatment; Complementary DNA; DNA cassette; Data; Development; Disease; Endothelial Cells; Eye; FOXO1A gene; Female; Fibrosis; Gel; Gene Expression; Genes; Genetic Transcription; Glaucoma; Goals; Histology; Human; In Vitro; Knowledge; MRI Scans; Mass Spectrum Analysis; Mechanics; MicroRNAs; Molecular Target; Monitor; Mus; Mutation; Nerve Fibers; Ocular Hypertension; Optic Disk; Pathogenesis; Pathway interactions; Patients; Periodicity; Phagocytosis; Phagocytosis Inhibition; Physiologic Intraocular Pressure; Physiological Processes; Play; Primary Open Angle Glaucoma; Proteins; Proteomics; Regulation; Reporting; Research; Retina; Retinal Degeneration; Retinal Ganglion Cells; Risk; Risk Factors; Role; Stains; Stretching; Structure of sinus venosus of sclera; Time; Tissues; Trabecular meshwork structure; Training; Transforming Growth Factor beta; Transgenic Mice; Untranslated RNA; Visual Fields; aqueous; cancer cell; comparison control; contrast enhanced; differential expression; high intraocular pressure; in silico; in vivo; innovation; male; multiple reaction monitoring; new therapeutic target; overexpression; particle; pharmacologic; premature; pressure; prevent; promoter; risk variant; senescence; single nucleus RNA-sequencing; therapeutic target; three dimensional cell culture; transcriptome sequencing

Abstract Despite the significant research effort of microRNAs (miRNAs) in the pathogenesis of glaucoma, their specific roles in the aqueous outflow pathway remain unclear. Lowering intraocular pressure (IOP) is the only clinical treatment for primary open-angle glaucoma (POAG), no matter whether the patients have high (IOP > 21 mmHg) or normal-tension glaucoma (IOP ≤ 21 mmHg). Significant achievements have been made in understanding the regulation of IOP and identifying therapeutic targets to lower IOP. We and others have identified the association of rs76481776 in the MIR182 gene with POAG. However, it remains unknown how miR-182-5p contributes to POAG risk. Without such knowledge, the development of miRNA-based glaucoma therapy will likely remain difficult. Our overall objective is to determine how elevated expression of miR-182-5p contributes to POAG with high IOP. The risk allele A of POAG-associated rs76481776 with POAG has been reported to increase mature miR-182-5p expression in vitro, suggesting that elevated miR-182-5p expression may contribute to POAG. We identified 2-fold higher miR-182-5p expression in glaucomatous aqueous humor compared to controls. Our central hypothesis is that elevated expression of miR-182-5p affects IOP by regulating the cellular functions of human trabecular meshwork (TM) and Schlemm's canal (SC) endothelial cells. Human TM cells with induced premature senescence have 7-9 fold higher miR-182-5p expression, and overexpression of miR-182-5p in these HTM cells leads to partial cellular senescence. miR-182-5p has been shown to target genes in many pathways, such as CHEK2, FOXO1, MTSS1, and CYLD. The expression of miR-182-5p could be induced by TGF-β treatment in cancer cells, leading to prolonged NF-κB activation, while TGF-β related pathways have been shown to play essential roles in POAG and fibrosis. The rationale for this proposal is that, once we determine how elevated miR-182-5p expression affects the cellular functions of human TM and SC, certain genes or miRNAs could be manipulated either up or down pharmacologically, resulting in new and innovative approaches to lower IOP and to delay/prevent the progression of glaucoma. In Aim 1, we will identify molecular targets of miR-182-5p in primary human TM cells in vitro. Human TM cells will be subject to cyclic mechanical stretch and tissue train 3D cell culture with uniaxial tissue stretch. We will perform whole proteomics profiling and strand-specific RNA-Seq analysis to identify miR-182-5p target genes. In Aim 2, we will determine the impact of miR-182-5p overexpression in the TM and SC on outflow facility and IOP levels in vivo using transgenic mice. Upon successful completion, we expect to identify the specific molecular targets of miR-182- 5p in human TM and SC to lower IOP more effectively in glaucoma patients.

抽象的 尽管 microRNA (miRNA) 在青光眼发病机制方面进行了大量研究，但它们的 在房水流出途径中的具体作用仍不清楚。降低眼压（IOP）是唯一的方法 原发性开角型青光眼（POAG）的临床治疗，无论患者是否患有高眼压（IOP > 21） mmHg) 或正常眼压青光眼 (IOP ≤ 21 mmHg)。取得了重大成就 了解 IOP 的调节并确定降低 IOP 的治疗靶点。我们和其他人有 确定了 MIR182 基因中的 rs76481776 与 POAG 的关联。然而，目前尚不清楚如何 miR-182-5p 会增加 POAG 风险。如果没有这些知识，基于 miRNA 的青光眼的发展 治疗可能仍然很困难。我们的总体目标是确定 miR-182-5p 的表达升高如何 有助于高 IOP 的 POAG。 POAG 相关 rs76481776 的风险等位基因 A 已被确定 据报道可增加体外成熟 miR-182-5p 表达，表明 miR-182-5p 表达升高 可能会导致 POAG。我们发现青光眼房水中 miR-182-5p 的表达高出 2 倍 与对照相比。我们的中心假设是 miR-182-5p 表达升高通过调节影响 IOP 人小梁网 (TM) 和施累姆氏管 (SC) 内皮细胞的细胞功能。人类 诱导早衰的TM细胞的miR-182-5p表达量高7-9倍，并且过度表达 这些 HTM 细胞中 miR-182-5p 的缺失导致部分细胞衰老。 miR-182-5p 已被证明能够靶向 许多途径中的基因，例如 CHEK2、FOXO1、MTSS1 和 CYLD。 miR-182-5p 的表达可以 TGF-β 在癌细胞中被诱导，导致 NF-κB 激活时间延长，而 TGF-β 相关 通路已被证明在 POAG 和纤维化中发挥重要作用。该提案的理由是， 一旦我们确定了 miR-182-5p 表达升高如何影响人类 TM 和 SC 的细胞功能， 某些基因或 miRNA 可以在药理学上被上调或下调，从而产生新的和 降低眼压和延缓/预防青光眼进展的创新方法。在目标 1 中，我们将确定 体外原代人 TM 细胞中 miR-182-5p 的分子靶标。人类TM细胞将受到循环 机械拉伸和组织训练 具有单轴组织拉伸的 3D 细胞培养。我们将进行整个蛋白质组学 分析和链特异性 RNA-Seq 分析来鉴定 miR-182-5p 靶基因。在目标 2 中，我们将确定 使用 TM 和 SC 中的 miR-182-5p 过表达对体内流出设施和 IOP 水平的影响 转基因小鼠。成功完成后，我们期望鉴定出 miR-182-的特定分子靶点 人类 TM 和 SC 中的 5p 可更有效地降低青光眼患者的 IOP。