Impact of endothelial cells on the smooth muscle foam cell phenotype

血管内皮细胞(ECs)控制着血管平滑肌细胞(VSMCs)表型的许多方面，包括其激活状态和张力。在动脉硬化的背景下，EC功能障碍是在斑块形成之前讨论的，斑块形成包括VSMCs向巨噬细胞样泡沫细胞的转分化。虽然已经描述了参与这种表型转换的几个调控决定因素，但尚不清楚ECs和VSMCs之间改变的旁分泌通讯是否有助于这一过程。为了研究这些细胞的相互作用，我们开发了一种基于球体的3D细胞培养模型，该模型允许生成由覆盖VSMC核心的单层内皮细胞组成的大小定义的血管器官。两种细胞分离后，转录组和蛋白质组分析表明，内皮细胞稳定了VSMCs的分化，减少了泡沫细胞标志物以及与脂质处理和代谢的不同方面相关基因的表达。因此，VSMC基因表达模式的控制似乎至少部分依赖于前列腺素介导的信号(特别是前列环素)。我们进一步研究了与前列腺素系统相关的基因表达是否在前动脉硬化环境中发生了改变，从而刺激了EC功能障碍和VSMC泡沫细胞的形成。为此，我们对scRNAseq数据库进行了荟萃分析，结果显示，在喂食高脂饮食的ApoE缺陷小鼠中，尤其是内皮Ptgs1(环氧合酶1)和Ptgis(前列环素合成酶)的表达(限制前列环素生成的酶的速率)下调。同样，高血压和家族性高胆固醇血症似乎会减少人类冠状动脉内皮细胞中与前列腺素分泌相关的基因的表达。考虑到这些数据，我们假设慢性暴露于高血糖(II型糖尿病)或血脂(血脂异常)等动脉硬化应激源引起的内皮细胞功能障碍限制了内皮细胞维持VSMC分化的有效性，从而促进了有利于VSMC泡沫细胞发育的表型转变。基于三维人体类器官细胞培养模型，本研究将探讨高糖和oxLDL水平对内皮细胞维持VSMC表型能力的影响，ii)临床相关的前列腺素合成抑制剂对动脉硬化前环境中VSMC表型转换的一般影响，以及iii)前列环素依赖的信号转导在这方面的作用。

Endothelial cells (ECs) control many aspects of the phenotype of vascular smooth muscle cells (VSMCs) including their state of activation and tone. In the context of arteriosclerosis, EC dysfunction is discussed to precede plaque formation that includes transdifferentiation of VSMCs to macrophage-like foam cells. While several regulatory determinants involved in this phenotypic switch were already described, it is unknown whether altered paracrine communication between ECs and VSMCs contributes to this process. For the study of those cellular interactions, we developed a spheroid-based 3D cell culture model that allows for the generation of size-defined vascular organoids composed of a single sheet of ECs covering a core of VSMCs. After separating both cell types, transcriptome and proteome analyses revealed that ECs stabilize the differentiation of VSMCs and decrease their expression of foam cell markers as well as of genes associated with different aspects of lipid handling and metabolism. The control of the VSMC gene expression pattern thereby appeared at least in part dependent on prostanoid-mediated signaling (especially prostacyclin). We further investigated whether the gene expression associated with the prostanoid system is altered in a pro-arteriosclerotic environment that stimulates both EC dysfunction and VSMC foam cell formation. To this end, we performed metaanalyses of scRNAseq databases, which revealed that especially endothelial Ptgs1 (cyclooxygenase 1) and Ptgis (prostacyclin synthase) expression (enzymes rate limiting for prostacyclin generation) is downregulated in ApoE-deficient mice fed a high fat diet. Similarly, hypertension and familial hypercholesterinemia appeared to decrease the expression of genes associated with ‘prostaglandin secretion’ in ECs from a human coronary artery. Considering these data, we hypothesize that dysfunction of ECs as evoked by chronic exposure to pro-arteriosclerotic stressors such as high glucose (diabetes type II) or lipid (dyslipidemia) serum concentrations limits the efficacy of ECs to maintain the differentiation of VSMCs thereby facilitating a phenotypic shift favoring the development of VSMC foam cells. Based on a 3D human organoid cell culture model, this study will investigate i) the impact of high glucose and oxLDL levels on the capacity of ECs to maintain the VSMC phenotype, ii) the general influence of clinically relevant pharmacological inhibitors of prostanoid generation on the VSMC phenotype switch in a pro-arteriosclerotic environment and iii) the role of prostacyclin-dependent signaling in this context.