RIG-I sensing of self-RNA in aged endothelial cells links cellular senescence with an interferon gene signature I

Senescent endothelial cells accumulate in blood vessels during aging and contribute to age-related cardiovascular disease. Identification of senescent cells is challenging as molecular changes are cell-type specific. Here, we established, benchmarked, and validated a new gene signature EndoSEN that pinpoints senescent endothelial cells. The EndoSEN signature was enriched for interferon stimulated genes (ISG) and correlated with the senescence-associated secretory phenotype (SASP) signature. SASP establishment is classically attributed to DNA damage and cGAS activation, but our results revealed a pivotal role for RNA accumulation and sensing. Mechanistically, we showed that endothelial senescence hallmarks include self-RNA accumulation, RNA sensor RIG-I upregulation, and an ISG signature. Moreover, a virtual model of RIG-I knockout in endothelial cells underscored senescence as an impacted pathway. We tested and confirmed that RIG-I knockdown was sufficient to extend the lifespan and decrease the SASP in endothelial cells. Our evidence suggests that targeting RNA sensing is a potential strategy to delay vascular aging. Cell suspensions for scRNA-seq were obtained from mouse retinas at 3 ages: 3, 12, and 23 months. Enrichment for CD31+ cells was performed using the AutoMACS.衰老过程中，血管内会蓄积衰老内皮细胞，且此类细胞会促成年龄相关性心血管疾病的发生发展。由于分子变化具有细胞类型特异性，衰老细胞的鉴定颇具挑战性。本研究构建了全新的基因特征标记物EndoSEN，并完成了基准测试与验证，该标记物可精准识别衰老内皮细胞。EndoSEN特征显著富集于干扰素刺激基因（Interferon Stimulated Genes, ISG），且与衰老相关分泌表型（Senescence-Associated Secretory Phenotype, SASP）特征呈显著相关。传统观点认为SASP的形成源于DNA损伤与cGAS通路激活，但本研究结果揭示了RNA蓄积与RNA感知在其中发挥的关键作用。机制层面上，本研究证实内皮细胞衰老的标志性特征包括自身RNA蓄积、RNA感受器RIG-I表达上调，以及ISG特征谱。此外，在内皮细胞中构建RIG-I基因敲除的虚拟模型，进一步证实衰老为受影响的核心通路。本研究验证发现，敲低RIG-I即可延长内皮细胞的存活时长，并降低其SASP分泌水平。本研究结果表明，靶向RNA感知通路或可成为延缓血管衰老的潜在治疗策略。本研究用于单细胞RNA测序（single-cell RNA sequencing, scRNA-seq）的细胞悬液取自三个年龄阶段（3月龄、12月龄及23月龄）的小鼠视网膜，并通过AutoMACS技术对CD31阳性细胞进行富集。