Novel Interplay of KILN and MKL1 in Vascular Pathophysiology

KILN 和 MKL1 在血管病理生理学中的新相互作用

基本信息

批准号：
10001073
负责人：
Xiaochun Long
金额：
$ 42.35万
依托单位：
AUGUSTA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-04-01 至 2023-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10001073
关键词：
Abdominal Aortic Aneurysm Aneurysm Angioplasty Angiotensin II Bacterial Artificial Chromosomes Biological Assay Blood Vessels Cell Differentiation process Cells Clustered Regularly Interspaced Short Palindromic Repeats Complex Cre driver Disease Exhibits Functional disorder Genes Genetic Transcription Goals Human IL8 gene Impairment In Vitro Inflammation Inflammatory Inflammatory Response Injury Knockout Mice Knowledge Lead Lesion Link Luciferases Maps Mediating Molecular Mus Myopathy Pathologic Pathology Patients Phenotype Plasma Play Predisposition Proteins Regulation Role Serum Response Factor Site Smooth Muscle Myocytes Stimulus System Therapeutic Transactivation Transgenic Mice Transgenic Organisms Ubiquitin Ubiquitination Untranslated RNA Vascular Diseases Vascular Smooth Muscle Vascular remodeling cancer cell cofactor combat exosome femoral artery genome editing human disease in vivo innovation insight mouse model multicatalytic endopeptidase complex neointima formation novel novel therapeutics p65 programs protein degradation restenosis stem cells therapeutic target transcriptome sequencing vascular inflammation vascular injury

项目摘要

Vascular smooth muscle cells (VSMCs) exhibit remarkable phenotypic plasticity, whereby differentiated contractile VSMCs switch to a synthetic state under pathological conditions. Synthetic VSMCs are manifested by their high susceptibility to inflammatory activation and loss of contractility, contributing largely to various vascular disorders such as post-angioplasty restenosis and aneurysm. Targeting early activation of VSMC inflammation may represent an attractive strategy to block vascular pathologies. However, molecular mechanisms involving key regulator(s) that drive the proinflammatory VSMC phenotype are incompletely understood. One possibility that has yet to be explored is the pervasive class of long noncoding RNAs (lncRNAs). Through an unbiased RNA-seq study, we discovered a novel human-specific lncRNA, KILN, which is enriched in proinflammatory VSMCs and diseased human vessels (aneurysm). RNA-seq revealed that loss of KILN in cultured VSMCs suppresses expression of multiple inflammatory genes. Notably, KILN is highly responsive to inflammatory insults in humanized Bacterial Artificial Chromosome (BAC) transgenic mice carrying either KILN (BAC-KILN) or both KILN and its neighboring gene IL8 (BAC-KILN/IL8). Both transgenic lines display an exacerbated inflammatory response triggered by vascular injury. These results support a critical role of KILN in promoting VSMC inflammation and vascular disease. Mechanistically, KILN interacts with MKL1, a potent transcriptional cofactor with a recognized role in transactivating VSMC contractile genes. Intriguingly, Mkl1 null mice are protected from aneurysm formation and depletion of MKL1 in cultured VSMCs impairs the proinflammatory gene program. These results suggest a potential link between MKL1 and KILN in VSMC inflammation and vascular disease. Indeed, loss of KILN decreases MKL1 protein levels and MKL1/p65 physical interaction, the latter being critical for transactivation of proinflammatory genes. These exciting preliminary findings support a central hypothesis that KILN interacts with MKL1 to stabilize MKL1 protein, which potentiates MKL1/p65-activated VSMC inflammation and vascular pathologies. We propose two specific aims to probe this hypothesis. Aim 1 will elucidate the regulation and function of KILN during pathological vascular remodeling using BAC transgenic mice, and evaluate KILN expression in vessels and plasma exosomes from aneurysm patients. Aim 2 will elucidate the molecular mechanism of KILN-mediated vascular inflammation and disease involving KILN disruption of MKL1 ubiquitination to potentiate MKL1/p65 transactivation of the proinflammatory gene program. Successful completion of the proposed studies will define a new molecular switch comprising a novel VSMC-enriched lncRNA (KILN) that associates with MKL1 for VSMC inflammation and pathological vascular remodeling. These studies will advance our knowledge of lncRNA and MKL1 vascular pathophysiology, potentially providing novel insights into therapeutic strategies for various inflammatory vascular diseases.

血管平滑肌细胞（VSMC）表现出显着的表型可塑性，从而分化在病理条件下，收缩性 VSMC 会转变为合成状态。合成 VSMC 显现由于它们对炎症激活和收缩力丧失高度敏感，在很大程度上导致了各种血管疾病，例如血管成形术后再狭窄和动脉瘤。针对 VSMC 的早期激活炎症可能是阻止血管病变的一种有吸引力的策略。然而，分子涉及驱动促炎 VSMC 表型的关键调节因子的机制不完全明白了。一种尚未探索的可能性是长链非编码 RNA 的普遍存在（lncRNA）。通过一项公正的 RNA-seq 研究，我们发现了一种新型的人类特异性 lncRNA，KILN，它富含促炎性 VSMC 和患病人体血管（动脉瘤）。 RNA-seq揭示了损失 KILN 在培养的 VSMC 中抑制多种炎症基因的表达。值得注意的是，KILN 高度人源化细菌人工染色体（BAC）转基因小鼠对炎症损伤的反应携带 KILN (BAC-KILN) 或同时携带 KILN 及其邻近基因 IL8 (BAC-KILN/IL8)。均为转基因线条显示血管损伤引发的加剧的炎症反应。这些结果支持 KILN 在促进 VSMC 炎症和血管疾病中发挥关键作用。从机制上讲，KILN 相互作用 MKL1 是一种有效的转录辅助因子，在反式激活 VSMC 收缩基因中具有公认的作用。有趣的是，Mkl1 缺失小鼠在培养的 VSMC 中免受动脉瘤形成和 MKL1 消耗的影响损害促炎基因程序。这些结果表明 MKL1 和 KILN 之间存在潜在联系 VSMC 炎症和血管疾病。事实上，KILN 的缺失会降低 MKL1 蛋白水平和 MKL1/p65 物理相互作用，后者对于促炎基因的反式激活至关重要。这些激动人心的初步研究结果支持一个中心假设，即 KILN 与 MKL1 相互作用以稳定 MKL1 蛋白，它增强了 MKL1/p65 激活的 VSMC 炎症和血管病理。我们提出两个具体方案旨在探讨这一假设。目标1将阐明KILN在病理过程中的调节和功能使用 BAC 转基因小鼠进行血管重塑，并评估血管和血浆中 KILN 的表达来自动脉瘤患者的外泌体。目标2将阐明KILN介导的血管生成的分子机制涉及 KILN 破坏 MKL1 泛素化以增强 MKL1/p65 的炎症和疾病促炎基因程序的反式激活。成功完成拟议的研究将定义一种新的分子开关，包含一种新型的富含 VSMC 的 lncRNA (KILN)，可与 MKL1 结合 VSMC 炎症和病理性血管重塑。这些研究将增进我们的知识 lncRNA 和 MKL1 血管病理生理学，可能为治疗策略提供新见解各种炎症性血管疾病。