A Tissue-Specific Soluble Platelet-Derived Growth Factor Receptor-beta Isoform Retains Functional Capacity

项目摘要/摘要 微血管功能障碍是一系列破坏性疾病的基础，从阿尔茨海默氏症到癌症。 然而，在血管相关的病理中，血管维持的潜在机制变得失调 仍在不断涌现，推动了基于血液的诊断和生物工程疗法的进步。我们 最近发现了一种截短的血小板衍生生长因子受体选择性剪接变异体-β(PDGFRβ) 它编码一种可溶性的PDGFRβ亚型(sPDGFFRβ)，它可能作为一种未来的诊断和 治疗靶点。受体酪氨酸激酶(RTK)，像PDGFRβ一样，通常有可溶的对应物 通过选择性剪接产生，起到“诱饵”受体的作用，负向调节配体诱导的信号转导 全长受体。周细胞(PC)表达全长PDGFRβ，介导其募集到 产生同源配体血小板衍生生长因子BB的微血管内皮细胞 Bb)-PC可提高血管稳定性和调节渗透性。然而，微血管PC密度和血管 组织和特殊血管床之间的通透性不同，血管功能障碍通常与 PC丢失和PDGFFRβ--PDGFR-BB信号失控。因此，PDGFRβ介导的PC招募对于 船只完整性，尽管管理它的确切机制尚不清楚。最近的研究报告了大量的 在脑病理场景中，sPDGFRβ是通过蛋白质分解产生的。然而，我们的数据表明， 在广泛的正常、健康的组织中，通过选择性剪接产生小的sPDGFRβ，尽管它也 可能与疾病状态有关。我们最近阐明了sPdgfrb的完整mRNA序列，使靶向 操纵和分析方法。除了在不同组织中广泛和差异表达外， 我们的初步发现表明，在小鼠脑中，与全长PDGFRb(FPdgfrb)表达细胞重叠，并且 脑实质中存在免疫激活的、非血管相关的sPDGFRβ蛋白信号。这些 研究结果与相关RTK中已建立的配体隔离机制一起考虑，为我们提供了 假设血小板衍生生长因子-BB的生物利用度由选择性剪接的sPDGFRβ调节以介导PC-血管 招募和调整血管的渗透性。因此，使用互补的体外和体内模型，我们 建议研究sPDGFR的β潜能以结合和调节(I)PDGF-BB的生物利用度，(Ii)激活 全长PDGFRβ(fPDGFRβ)，(Iii)PC动力学，(Iv)发展血管形态和通透性。我们 将研究sPDGFRβ的细胞特异性，以及在不同组织中的时空分布，以确定 其职能作用的程度。此外，我们将评估sPDGFRβ作为生物标记物和治疗的潜力。 在涉及PC丢失的血管相关病理中。这项工作将促进我们对机制的理解 基础血管维护和完整性，并为后续合作研究目标奠定基础 开发sPDGFRβ作为心血管疾病的潜在诊断工具和治疗靶点。

PROJECT SUMMARY / ABSTRACT Microvascular dysfunction underlies a wide range of devastating diseases, from Alzheimer’s Disease to cancer. However, mechanisms underlying vessel maintenance that become dysregulated in vascular-related pathologies are still emerging, fueling the advancement of blood-based diagnostics and bioengineered therapeutics. We recently identified a truncated, alternative splice variant of Platelet-Derived Growth Factor Receptor-β (PDGFRβ) that encodes a soluble PDGFRβ isoform (sPDGFRβ), which may harbor potential as a future diagnostic and therapeutic target. Receptor tyrosine kinases (RTKs), like PDGFRβ, often have soluble counterparts that are generated via alternative splicing to function as “decoy” receptors to negatively regulate ligand-induced signaling of the full-length receptor. Full-length PDGFRβ is expressed by pericytes (PCs) to mediate their recruitment to microvascular endothelial cells (ECs) producing the cognate ligand Platelet-Derived Growth Factor BB (PDGF- BB) – where PCs promote vessel stability and tune permeability. However, microvascular PC density and vessel permeability vary between tissues and specialized vascular beds, with vessel dysfunction often associated with PC loss and misregulated PDGFRβ--PDGF-BB signaling. Thus PDGFRβ-mediated PC recruitment is vital to vessel integrity, although the exact mechanisms that govern it remain unclear. Recent studies report a large sPDGFRβ produced via proteolytic cleavage in cerebral pathology scenarios. However, our data indicate that a small sPDGFRβ is generated via alternative splicing in a broad range of normal, healthy tissues, though it is also likely involved in disease states. We recently elucidated the full mRNA sequence of sPdgfrb, enabling targeted manipulation and analysis approaches. In addition to broad and differential expression across various tissues, our preliminary findings indicate overlap with full-length Pdgfrb (fPdgfrb)-expressing cells in mouse brain, and presence of immunolabled, non-vessel associated sPDGFRβ protein signal in the brain parenchyma. These findings, considered alongside established mechanisms of ligand sequestration in related RTKs, inform our hypothesis that PDGF-BB bioavailability is regulated by alternatively spliced sPDGFRβ to mediate PC-vessel recruitment and tune vessel permeability. Therefore, using complementary in vitro and in vivo models, we propose investigation of sPDGFRβ potential to bind and regulate (i) PDGF-BB bioavailability, (ii) activation of full-length PDGFRβ (fPDGFRβ), (iii) PC dynamics, and (iv) developing vessel morphology and permeability. We will investigate sPDGFRβ cell-specificity, and spatio-temporal distribution in various tissues to determine the extent of its functional role. In addition, we will assess the potential of sPDGFRβ as a biomarker and treatment in vascular-related pathologies involving PC loss. This work will advance our understanding of mechanisms underlying vessel maintenance and integrity, and lay the groundwork for follow-on collaborative studies aiming to develop sPDGFRβ as a potential diagnostic tool and therapeutic target in cardiovascular diseases.