Stem Cell Pluripotency Genes Klf4 and Oct4 Regulate Complex SMC Phenotypic Changes Critical in Late-Stage Atherosclerotic Lesion Pathogenesis.
Stem Cell Pluripotency Genes Klf4 and Oct4 Regulate Complex SMC Phenotypic Changes Critical in Late-Stage Atherosclerotic Lesion Pathogenesis.
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DOI：
10.1161/circulationaha.120.046672
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发表时间：
2020-11-24
期刊：
影响因子：
37.8
通讯作者：
Owens GK
中科院分区：
文献类型：
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作者：
Alencar GF;Owsiany KM;Karnewar S;Sukhavasi K;Mocci G;Nguyen AT;Williams CM;Shamsuzzaman S;Mokry M;Henderson CA;Haskins R;Baylis RA;Finn AV;McNamara CA;Zunder ER;Venkata V;Pasterkamp G;Björkegren J;Bekiranov S;Owens GK
补充数字内容可在文本中找到。晚期动脉粥样硬化病变的破裂和糜烂导致心肌梗死或中风是世界范围内的主要死亡原因。然而，我们对构成晚期动脉粥样硬化病变的许多细胞的身份、起源和功能以及它们控制斑块稳定性的机制的了解有限。我们对晚期人颈动脉内膜切除术样本进行了全面的单细胞RNA测序，并将其与来自小鼠显微解剖的晚期动脉粥样硬化病变的单细胞RNA测序进行了比较，平滑肌细胞（SMC）和内皮细胞谱系追踪调查所有斑块细胞类型并严格确定其来源。我们进一步使用染色质免疫沉淀测序（ChIP-seq），批量RNA测序和创新的双谱系示踪小鼠来了解SMC表型转变影响病变发病机制的机制。我们提供的证据表明，SMC特异性Klf 4敲除与Oct 4敲除显示出几乎相反的基因组特征，并且它们推定的靶基因在调节SMC表型变化中发挥着重要作用。单细胞RNA测序显示小鼠和人类病变之间的转录组簇的显著相似性以及SMC和内皮细胞衍生细胞的广泛可塑性，包括7个不同的簇，大多数对传统标记物呈阴性。特别是，SMC有助于Myh 11-，Lgals 3+群体与软骨细胞样基因签名，这是显着减少SMC-Klf 4敲除。我们观察到，激活Lgals 3的SMC占病变中所有SMC的三分之二。然而，这些细胞中Lgals 3的初始活化并不代表转化为终末分化状态，而是代表这些细胞转变为独特的干细胞标志物基因阳性、细胞外基质重塑的“先锋”细胞表型，其首先在病变内投资，随后在晚期病变内产生至少3种其他SMC表型，包括Klf 4依赖性成骨表型，可能导致斑块钙化和斑块不稳定。综上所述，这些结果提供了证据表明，在晚期小鼠和人动脉粥样硬化病变中SMC衍生的细胞表现出比通常认为的大得多的表型可塑性，Klf 4调节向多种表型的转变，包括Lgals 3+成骨细胞可能对晚期动脉粥样硬化斑块发病机制有害。
Supplemental Digital Content is available in the text. Rupture and erosion of advanced atherosclerotic lesions with a resultant myocardial infarction or stroke are the leading worldwide cause of death. However, we have a limited understanding of the identity, origin, and function of many cells that make up late-stage atherosclerotic lesions, as well as the mechanisms by which they control plaque stability. We conducted a comprehensive single-cell RNA sequencing of advanced human carotid endarterectomy samples and compared these with single-cell RNA sequencing from murine microdissected advanced atherosclerotic lesions with smooth muscle cell (SMC) and endothelial lineage tracing to survey all plaque cell types and rigorously determine their origin. We further used chromatin immunoprecipitation sequencing (ChIP-seq), bulk RNA sequencing, and an innovative dual lineage tracing mouse to understand the mechanism by which SMC phenotypic transitions affect lesion pathogenesis. We provide evidence that SMC-specific Klf4- versus Oct4-knockout showed virtually opposite genomic signatures, and their putative target genes play an important role regulating SMC phenotypic changes. Single-cell RNA sequencing revealed remarkable similarity of transcriptomic clusters between mouse and human lesions and extensive plasticity of SMC- and endothelial cell-derived cells including 7 distinct clusters, most negative for traditional markers. In particular, SMC contributed to a Myh11-, Lgals3+ population with a chondrocyte-like gene signature that was markedly reduced with SMC-Klf4 knockout. We observed that SMCs that activate Lgals3 compose up to two thirds of all SMC in lesions. However, initial activation of Lgals3 in these cells does not represent conversion to a terminally differentiated state, but rather represents transition of these cells to a unique stem cell marker gene–positive, extracellular matrix-remodeling, “pioneer” cell phenotype that is the first to invest within lesions and subsequently gives rise to at least 3 other SMC phenotypes within advanced lesions, including Klf4-dependent osteogenic phenotypes likely to contribute to plaque calcification and plaque destabilization. Taken together, these results provide evidence that SMC-derived cells within advanced mouse and human atherosclerotic lesions exhibit far greater phenotypic plasticity than generally believed, with Klf4 regulating transition to multiple phenotypes including Lgals3+ osteogenic cells likely to be detrimental for late-stage atherosclerosis plaque pathogenesis.