Impaired TRVP4-eNOS signaling in TM contributes to glaucoma

TM 中 TRVP4-eNOS 信号传导受损导致青光眼

• 批准号: 10503094
• 负责人: Swapnil K. Sonkusare
• 金额: $ 44.41万
• 依托单位: UNIVERSITY OF NORTH TEXAS HLTH SCI CTR
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2023-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10503094
• 关键词: Aqueous Humor; Blindness; Cell model; Cells; Chemicals; Chronic; Electrophysiology (science); Enzymes; Eye; Functional disorder; Genetic; Glaucoma; Goals; Homeostasis; Human; Image; Impairment; Ion Channel; Laboratories; Measurement; Molecular; Mus; NOS2A gene; NOS3 gene; Nitric Oxide; Nitric Oxide Synthase; Ocular Hypertension; Oxidants; Oxidation-Reduction; Oxidative Stress; Pathologic; Pathology; Pathway interactions; Perfusion; Peroxonitrite; Pharmacology; Physiologic Intraocular Pressure; Post-Translational Protein Processing; Primary Open Angle Glaucoma; Research; Resistance; Risk Factors; Signal Pathway; Signal Transduction; Techniques; Tissues; Trabecular meshwork structure; Vanilloid; aqueous humor flow; endoplasmic reticulum stress; ex vivo perfusion; human model; improved; inhibitor; innovation; insight; knock-down; mouse model; novel; overexpression; patch clamp; receptor; therapeutic target; transcription factor

Abstract Primary Open Angle Glaucoma (POAG) is the most common form of glaucoma that leads to irreversible vision loss. Elevated intraocular pressure (IOP) due to dysfunction of trabecular meshwork (TM) tissue is a hallmark of POAG. However, the pathological mechanisms leading to TM dysfunction and IOP elevation are poorly understood. TM has an intrinsic ability to maintain IOP homeostasis by sensing the changes in the flow of aqueous humor (AH). In this regard, we recently showed that Ca2+ influx through transient receptor potential vanilloid 4 ion channels in the TM (TRPV4TM channels) lowers IOP via activation of endothelial nitric oxide synthase (eNOS)–NO signaling. Importantly, we showed that TRPV4TM-eNOSTM signaling is impaired in glaucomatous primary human TM cells. The major goals of this application are to elucidate the pathological mechanisms that impair TRPV4TM-eNOSTM signaling in glaucoma and to target them for rescuing the TM function. Chronic endoplasmic reticulum (ER) stress is a crucial contributor to TM dysfunction and IOP elevation in glaucoma. In our preliminary studies, we observed that chronic ER stress activates inducible NOS (iNOS), an enzyme commonly associated with the formation of oxidant molecule peroxynitrite (PN). PN levels are elevated in TM tissues from POAG donor eyes and exogenous PN reduced TRPV4TM channel activity in human primary TM cells. Moreover, induction of ER stress also lowered TRPV4TM channel activity. Therefore, we hypothesize that PN-induced inhibition of TRPV4TM-eNOSTM signaling contributes to TM dysfunction and IOP elevation in glaucoma. The major objectives of this application are to determine whether chronic ER stress leads to TM dysfunction and IOP elevation via PN-induced lowering of TRPV4TM-eNOSTM signaling in glaucoma, and to target this pathology for the treatment of glaucoma. In Aim 1, we will determine whether chronic ER stress lowers TRPV4 channel activity in TM cells. In Aim 2, we will determine whether PN levels are elevated in glaucoma and whether PN lowers TRPV4 channel activity in TM. We will also determine whether chronic ER stress underlies elevated PN levels in glaucoma. In Aim 3, we will target PN pathology to lower elevated IOP in mouse and human models of ocular hypertension. This proposal utilizes complementary expertise of Dr. Zode’s laboratory in glaucoma research and ER stress, and Dr. Sonkusare’s laboratory in TRP ion channel imaging and electrophysiology. This study will utilize state-of-art Ca2+ imaging, patch-clamp, eNOS activity, and nitric oxide measurements in primary human TM cells and TM tissues, human perfusion cultured donor eyes, and mouse models of glaucoma. Successful completion of the proposed studies will provide novel pathological mechanisms and therapeutic targets for the treatment of general POAG.

抽象的 原发性开角型青光眼 (POAG) 是最常见的青光眼形式，可导致视力不可逆 损失。由于小梁网 (TM) 组织功能障碍导致的眼内压 (IOP) 升高是 波格。然而，导致TM功能障碍和眼压升高的病理机制尚不清楚。 明白了。 TM 具有通过感知眼内血流变化来维持 IOP 稳态的内在能力。 房水（AH）。在这方面，我们最近表明 Ca2+ 通过瞬时受体电位流入 TM 中的香草酸 4 离子通道（TRPV4TM 通道）通过激活内皮一氧化氮降低 IOP 合酶 (eNOS)-NO 信号传导。重要的是，我们发现 TRPV4TM-eNOSTM 信号在 青光眼原代人 TM 细胞。该应用的主要目标是阐明病理学 损害青光眼中 TRPV4TM-eNOSTM 信号传导的机制并针对它们来挽救 TM 功能。慢性内质网 (ER) 应激是 TM 功能障碍和 IOP 升高的关键因素 在青光眼中。在我们的初步研究中，我们观察到慢性 ER 应激会激活诱导型 NOS (iNOS)，这是一种 通常与氧化剂分子过氧亚硝酸盐 (PN) 的形成有关的酶。 PN 水平升高 POAG 供体眼的 TM 组织和外源 PN 降低了人类原代细胞中 TRPV4TM 通道的活性 TM 细胞。此外，ER应激的诱导也降低了TRPV4TM通道活性。因此，我们假设 PN 诱导的 TRPV4TM-eNOSTM 信号传导抑制导致 TM 功能障碍和 IOP 青光眼升高。该应用的主要目标是确定慢性 ER 应激是否 通过 PN 诱导的青光眼 TRPV4TM-eNOSTM 信号降低导致 TM 功能障碍和 IOP 升高， 并针对这种病理学来治疗青光眼。在目标 1 中，我们将确定慢性 ER 应激是否 降低 TM 细胞中 TRPV4 通道的活性。在目标 2 中，我们将确定 PN 水平是否升高 青光眼以及 PN 是否会降低 TM 中 TRPV4 通道的活性。我们还将确定是否存在慢性 ER 压力是青光眼 PN 水平升高的基础。在目标 3 中，我们将针对 PN 病理学来降低升高的 IOP。 小鼠和人类高眼压模型。该提案利用了 Zode 博士的互补专业知识 青光眼研究和 ER 应激实验室，以及 Sonkusare 博士的 TRP 离子通道成像和 电生理学。这项研究将利用最先进的 Ca2+ 成像、膜片钳、eNOS 活性和一氧化氮 原代人 TM 细胞和 TM 组织、人体灌注培养供体眼睛和小鼠的测量 青光眼模型。拟议研究的成功完成将提供新的病理机制 以及一般 POAG 治疗的治疗靶点。