Adenosine receptor 2A in subretinal fibrosis

腺苷受体2A在视网膜下纤维化中的作用

• 批准号: 10614638
• 负责人: Ruth B Caldwell
• 金额: $ 43.78万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2027-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10614638
• 关键词: Actins; Adenosine; Anatomy; Angioblast; Blood Vessels; Bone Marrow; Cells; Choroidal Neovascularization; Cicatrix; Data; Development; Endothelial Cells; Endothelium; Epithelial Cells; Exudative age-related macular degeneration; Eye diseases; Fibrosis; Genetic; Hypoxia Inducible Factor; Hypoxia-Inducible Factor Pathway; In Vitro; Infiltration; Inflammatory; Lasers; Lesion; Macrophage; Mediating; Mesenchymal; Molecular; Mus; Myelogenous; Myeloid Cells; Myofibroblast; Neuroglia; Oxygen; Pathologic; Patients; Prevention; Production; Profibrotic signal; Purinergic P1 Receptors; Receptor Signaling; Retina; Retinal Diseases; Role; Signal Transduction; Smooth Muscle; Snails; Structure of retinal pigment epithelium; Testing; Therapeutic; Transforming Growth Factor Beta 2; Vascular Diseases; Vascular Endothelial Growth Factors; Visual impairment; cell type; design; in vitro Model; in vivo; inhibitor; mouse model; novel strategies; novel therapeutic intervention; overexpression; pharmacologic; retinal angiogenesis; tool; transcription factor; vascular inflammation

PROJECT SUMMARY Subretinal fibrosis, an end-stage fibrous scar of neovascular age-related macular degeneration (nAMD), compromises highly organized anatomical layers and tightly coordinated cellular interactions, inevitably leading to irreversible visual impairment. The current treatment for subretinal fibrosis is limited and therefore, new therapeutic strategies for the inhibition of subretinal fibrosis are imperative. Multiple cell types, including endothelial cells (ECs), retinal pigment epithelium (RPE) cells, macrophages and glial cells, contribute to subretinal fibrosis by either differentiating into mesenchymal-like cells and further differentiating into α-smooth muscle actin-positive myofibroblasts and/or producing profibrotic and proinflammatory factors. However, the underlying mechanisms for these cellular and molecular activities remain poorly defined. Adenosine receptor 2A (Adora2a) has been implicated in various vascular diseases and inflammation. Our preliminary data here show that (i) the level of Adora2a expression was increased in subretinal lesions of laser-induced CNV in mice; (ii) the size of subretinal fibrosis was markedly decreased in lesions of laser–induced CNV in Adora2a-deficient mice; (iii) endothelial-to-mesenchymal transition (EndMT) occurred to choroidal ECs (CECs) and EndMT participated in the formation of subretinal fibrosis in laser-induced mouse CNV; (iv) Tgfb2-induced EndMT was decreased for Adora2a-deficient CECs; (v) macrophage-to-myofibroblast transition (MMT) in laser-induced subretinal fibrotic lesions was markedly reduced in Adora2a-deficient mice; (vi) Adora2a-deficient bone marrow derived macrophages (BMDMs) had a compromised production of profibrotic factors after stimulation with Tgfb2; and (vii) the levels of hypoxia-inducible factor (Hif) 1a or 2a dynamically correlated with those of Adora2a in the above pathological alterations. Thus, we hypothesize that Adora2a- mediated Hif signaling in CECs and infiltrated macrophages enhance fibrotic effects leading to increased formation of fibrotic lesions in CNV. To test our hypothesis, we have generated mice with inducible global Adora2a deficiency in Vldlr-/- mice, endothelial lineage tracing mice, inducible endothelial Adora2a deficiency in C57BL/6j mice, and myeloid Adora2a deficiency in C57BL/6j mice. We established an ex vivo approach to culture mouse CECs and in vitro approaches to generate BMDMs. We will investigate the effect of Adora2a inactivation in CECs and myeloid cells in subretinal fibrosis using specific genetic and pharmacological tools and assess subretinal fibrosis using an integrated approach of in vivo, ex vivo, and in vitro models. Our study will define the role of Adora2a in the development of subretinal fibrosis and provide the basis for using ADORA2A inhibition as a novel approach in the prevention and treatment of blinding retinal disease.

项目摘要 视网膜下纤维化是新生血管性年龄相关性黄斑变性（nAMD）的终末期纤维性瘢痕， 在高度组织化的解剖层和紧密协调的细胞相互作用之间进行妥协， 不可逆转的视力损伤目前视网膜下纤维化的治疗是有限的，因此， 抑制视网膜下纤维化的治疗策略是必要的。 多种细胞类型，包括内皮细胞（EC）、视网膜色素上皮（RPE）细胞、巨噬细胞 和神经胶质细胞，通过分化成间充质样细胞并进一步 分化成α-平滑肌肌动蛋白阳性肌成纤维细胞和/或产生促纤维化和 促炎因子然而，这些细胞和分子活动的潜在机制仍然存在， 定义不好。腺苷受体2A（Adora 2a）与多种血管疾病有关， 炎症我们的初步数据显示：（i）Adora 2a在视网膜下的表达水平增加， 激光诱导的小鼠CNV病变;（ii）视网膜下纤维化的大小显着减少， Adora 2a缺陷小鼠中激光诱导的CNV;（iii）发生内皮-间充质转化（EndMT）， 脉络膜内皮细胞（CECs）和EndMT参与了激光诱导的小鼠视网膜下纤维化的形成 CNV;（iv）Adora 2a缺陷CEC中Tgfb 2诱导的EndMT减少;（v）巨噬细胞-肌成纤维细胞 在Adora 2a缺陷小鼠中，激光诱导的视网膜下纤维化病变中的MMT显著减少;（vi） Adora 2a缺陷型骨髓源性巨噬细胞（BMDM）的促纤维化蛋白（profibrotic protein）产生受损， 低氧诱导因子（Hif）1a或2a的动态水平 与Adora 2a的上述病理改变相关。我们假设Adora 2a- CEC和浸润的巨噬细胞中介导的Hif信号增强纤维化作用，导致增加 CNV中纤维化病变的形成。为了验证我们的假设，我们已经产生了具有可诱导的全局 Vldlr-/-小鼠中的Adora 2a缺陷，内皮谱系示踪小鼠， C57 BL/6 j小鼠中的骨髓Adora 2a缺乏和C57 BL/6 j小鼠中的骨髓Adora 2a缺乏。我们建立了一种体外培养方法， 小鼠CEC和产生BMDM的体外方法。我们将研究Adora 2a失活的影响， 视网膜下纤维化中CEC和髓样细胞的研究， 使用体内、离体和体外模型的综合方法进行视网膜下纤维化。我们的研究将定义 Adora 2a在视网膜下纤维化发展中的作用，并为使用ADORA 2A抑制剂作为 一种预防和治疗致盲性视网膜疾病的新方法。