Switch of Osteogenesis in Vascular Calcification

摘要 血管钙化的治疗进展可能会对公众产生深远的影响。血管钙化是一种 糖尿病并发症多见，发病率和死亡率均呈上升趋势。虽然 确切的机制尚未确定，已知的血管钙化是一个活跃的过程 涉及异位骨形成，其中成骨分化发生在从 其他血统。先前的研究表明，血管内皮细胞(ECs)改变细胞命运以分化 转化为成骨样细胞，促进血管钙化。然而，尚不清楚是否将 内皮细胞来源的成骨样细胞向内皮细胞分化的转录图谱改善 血管钙化。在目前的建议中，我们利用单细胞RNA测序(scRNA-seq)和 连通性地图(Cmap)确定了一种新的方法，旨在改变EC衍生的转录格局 成骨样细胞向内皮细胞分化及其对糖尿病血管钙化的影响 糖尿病。在初步研究中，我们使用谱系追踪、scRNA-seq和Cmap查询来鉴定细胞周期蛋白依赖性 抑制CDK1将EC来源的成骨细胞重定向为内皮细胞分化 显著改善血管钙化。我们发现CDK1在EC来源的成骨细胞样细胞中具有特异性的诱导作用 细胞。我们发现CDK1的缺失或其抑制剂AT7519增加了E-26特异性序列变体2 (ETV2)，负责将EC来源的成骨细胞样细胞的转录格局转变为 内皮细胞分化。此外，ECs或AT7519中CDK1的缺失减少了EC来源的成骨和 在不影响其他组织的情况下减少糖尿病小鼠的主动脉钙化。因此，我们假设 抑制CDK1诱导ETV2将EC来源的成骨细胞重新定向为内皮细胞分化 从而改善糖尿病患者的血管钙化。在具体目标1中，我们将阐明其机制。 潜在的CDK1抑制使EC来源的成骨细胞样细胞向内皮细胞分化。在……里面 具体目标2，我们将确定抑制CDK1是否改善糖尿病小鼠模型中的血管钙化。 如果成功，它将把命运不佳的细胞重新定向到正常化，作为一个新的概念，而CDK1 抑制可能成为一种治疗获得性血管疾病钙化的新方法。

SUMMARY Therapeutic advances in vascular calcification may have far-reaching public benefits. Vascular calcification is a frequent complication of diabetes mellitus and associated with the increase of morbidity and mortality. Although the precise mechanism has not been determined, vascular calcification is known to be an active process involving ectopic bone formation, in which osteogenic differentiation occurs in the cells transdifferentiated from other lineages. Previous studies have shown that vascular endothelial cells (ECs) switch cell fate to differentiate into osteoblastic-like cells to contribute to vascular calcification. However, it is unknown if shifting the transcriptional landscape of EC-derived osteoblast-like cells back to endothelial differentiation ameliorates vascular calcification. In present proposal, we take advantage of single-cell RNA sequencing (scRNA-seq) and connectivity Map (CMap) to identify a novel approach, aiming to shift the transcriptional landscape of EC-derived osteoblast-like cells to endothelial differentiation and investigate its effect on vascular calcification in diabetes mellitus. In preliminary study, we use lineage tracing, scRNA-seq and CMap query to identify cyclin-dependent kinase 1 (CDK1) inhibition to redirect EC-derived osteoblast-like cells to endothelial differentiation and significantly improve vascular calcification. We find a specific induction of CDK1 in EC-derived osteoblast-like cells. We show that CDK1 deletion or its inhibitor AT7519 increases E-twenty-six specific sequence variant 2 (ETV2), which is responsible for shifting the transcriptional landscape of EC-derived osteoblast-like cells to endothelial differentiation. Furthermore, CDK1 deletion in ECs or AT7519 reduces EC-derived osteogenesis and decreases aortic calcification in diabetic Ins2Akita/+ mice without affecting other tissues. Therefore, we hypothesize that CDK1 inhibition induces ETV2 to redirect EC-derived osteoblast-like cells back to endothelial differentiation in turn to ameliorate vascular calcification in diabetes mellitus. In specific Aim 1, we will elucidate the mechanism underlying CDK1 inhibition that shifts EC-derived osteoblast-like cells toward endothelial differentiation. In specific Aim 2, we will determine if CDK1 inhibition ameliorates vascular calcification in a diabetic mouse model. If successful, it will build the redirection of ill-fated cells back to normalization as a new concept, and the CDK1 inhibition may emerge as a new therapeutic approach to treat calcification in acquired vascular diseases.