type I interferon regulates angiogenesis in Down Syndrome

项目总结 21三体(T21)是世界上最常见的染色体异常，可导致唐氏综合征(DS) [1，2]，一种与心肺疾病相关的多系统综合征。肺部疾病包括 肺高压、肺发育不良、肺含铁血黄素沉着症和肺毛细管炎，均为 这可能是由于肺内皮细胞调节失调造成的。此外，T21肺的特征是 肺泡周围未成熟的双毛细血管网络系统持续存在 囊状发育阶段[3]。这表明DS患者的内皮缺陷很可能是始发的 在子宫里。单细胞转录组数据显示存在不同的内皮细胞群 人类成人肺[4，5]。而其他人和我们已经报道了在发展中内皮网络受损 T21肺的特点是毛细血管充血，淋巴管扩张，动脉肌化[6]，性质 这些缺陷以及导致这些缺陷的不同内皮细胞群尚未得到 明白了。此外，发育中的T21人肺显示出抗血管生成因子的表达增加[3， 6-8]。在T21外周血中发现内皮祖细胞减少。此外，T21 iPSC- 与对照组相比，衍生的内皮细胞表现出细胞增殖减少，发芽和管状形成受阻 整倍体细胞[10]。在DS中持续激活的主要途径之一是I型干扰素(IFN)途径 [6]。I型干扰素升高导致内皮祖细胞减少[9]，并抑制血管内皮生长因子诱导的 形成毛细血管样结构[11，12]，从而起到抗血管生成的作用。此外，类型I 干扰素病与肺动脉高压和肺血管病有关[13]。SC-RNASeq数据的 T21和整倍体肺细胞显示有几个内皮细胞群 T21细胞中I型干扰素信号通路基因表达增加。因此，我们假设有缺陷的 肺血管生成发生在发育过程中的T21，是I型干扰素依赖性改变的结果 内皮细胞的增殖、分化和迁移。为了验证这一假设，我们将1)确定内皮细胞 T21与整倍体人类发育肺中的细胞缺陷 用IF染色和原位杂交相结合的方法评价缺陷的时空分布 通过IF和3D图像定量检测各组内皮细胞的增殖和发芽情况；2)定义 I型干扰素信号在T21血管内皮细胞损伤中的作用 干扰素信号对原代和IPSC来源的内皮细胞增殖、迁移、通透性和血管形成的影响 T21细胞和整倍体细胞培养。改进了预防、改善或逆转内皮相关的策略 DS中的肺部疾病将取决于对缺陷和导致这些缺陷的途径的详细了解 可能确定干预的新治疗靶点的缺陷。

PROJECT SUMMARY Trisomy 21 (T21) is the most prevalent chromosomal abnormality worldwide, resulting in Down Syndrome (DS) [1, 2], a multisystem syndrome associated with cardiopulmonary disorders. Pulmonary disorders include pulmonary hypertension, pulmonary hypoplasia, pulmonary hemosiderosis and pulmonary capillaritis, all of which can result from a dysregulated lung endothelium. In addition, T21 lungs are characterized by the persistence of an immature double capillary network system surrounding the alveoli and representative of the saccular stage of development [3]. This suggests that endothelial defects in individuals with DS are likely initiated in utero. Single cell transcriptomics data demonstrated the presence of diverse endothelial cell populations in the human adult lung [4, 5]. While others and we have reported impaired endothelial networks in the developing T21 lung characterized by congested capillaries, lymphatic dilatation, and muscularized arteries [6], the nature of these defects and the different endothelial cell populations contributing to such defects are yet to be understood. Moreover, developing T21 human lungs display increased expression of anti-angiogenic factors [3, 6-8]. A decrease in endothelial progenitor cells is found in T21 peripheral blood [9]. Furthermore, T21 iPSC- derived endothelial cells exhibit decreased cell proliferation, impaired sprouting and tube formation compared to euploid cells [10]. One of the major pathways consistently activated in DS is the type I interferon (IFN) pathway [6]. Elevated type I IFN causes a decrease of endothelial progenitor cells [9] and inhibits VEGF induced development of capillary like structures [11, 12], thus playing an anti-angiogenic role. Additionally, type I interferonopathies are linked to pulmonary hypertension and pulmonary vasculopathy [13]. Sc-RNAseq data of T21 and euploid lung cells demonstrated the presence of several endothelial cell populations displaying increased expression of type I IFN signaling pathway genes in T21. Therefore, we hypothesize that defective lung angiogenesis occurs in T21 during development and is a result of type I IFN-dependent changes in endothelial cell proliferation, differentiation and migration. To test this hypothesis, we will 1) determine endothelial cell defects in T21 vs euploid human developing lungs by defining the different endothelial cell populations and their spatiotemporal distribution using combinatorial in situ hyridization with IF stainings and assessing defects in endothelial cell proliferation and sprouting of each group via IF and 3D image quantifications and 2) define the role of type I IFN signaling in the endothelial defects in T21 by determining the effect of gain and loss of type I IFN signaling on proliferation, migration, permeability and tube formation in primary and iPSC-derived endothelial cell cultures of T21 and euploid cells. Improved strategies to prevent, ameliorate or reverse endothelial related lung disease in DS will be contingent upon a detailed understanding of the defects and pathways driving these defects that may identify novel therapeutic targets for intervention.