Yale TMC for Cellular Senescence in Lymphoid Organs

项目总结 人体组织中的细胞衰老是在其他增殖细胞中不可逆转的细胞周期停滞状态，这是 是衰老的标志，通过衰老相关的分泌表型改变组织环境 (SASP)，但也可能在组织重塑、再生和伤口愈合方面发挥有益作用。淋巴组织 器官在造血和免疫功能中起着至关重要的作用。这些组织中的细胞衰老是如何 与应激诱导的或年龄相关的功能损害有关的衰老细胞有哪些类型或亚型 它们的空间异质性以及这些细胞如何影响组织环境仍然很差 了解，排除制定针对衰老细胞的战略，以提高健康寿命/寿命 或利用这些细胞或分泌因子来促进组织重塑和修复。最近的一篇评论 NIH在《细胞》杂志上发表的文章确定了五大领域(图谱、成像和可视化、生物标记物、模型 系统、扰动和验证)，这将有助于推动该领域向前发展。我们的应用程序将组装一个 多学科团队处理所有这些领域，特别是作为组织绘图中心(TMC)，将专注于 关于生成细胞衰老和相关组织环境的分子和细胞图4 初级和次级淋巴器官。具体地说，它将(A)收集、分析、注释和分享高 优质未患病人原发(骨髓和胸腺)和继发(扁桃体和淋巴结) 淋巴组织，(2)开发和部署一套高分辨率、高含量、高通量的单核 细胞和空间组学技术来表征这些标本和配对的生物液，以及(3)执行 综合信息学识别衰老细胞异质性的生物标志物并构建全面的 这些器官中细胞衰老和相关环境的分子和细胞图谱。四大 生物分析流水线是：(1)单细胞高复合体(&gt；40)蛋白质分泌组谱，(2)单细胞 蛋白质转录组学测序(scCITE-seq)，(3)细胞水平的空间蛋白质组学测序 (DBiT-seq用于在细胞水平上共同定位整个转录组和一组约300个蛋白质，像素为10微米 大小)，以及(4)单分子FFPE组织中~1000个分子靶点的空间分子成像(SMI 亚细胞分辨率。使用这些独特的工具，我们将(A)表征功能SASP的异构性，并 确定不同细胞类型的SAPS的生物标志物，(B)构建4个人的分子和细胞图谱 淋巴组织，以及(C)确定组织和相关环境中细胞衰老的生物标志物， 为Sennet的资源建设做出贡献。因为免疫功能是所有人生理的中心 主要器官，我们的研究将为衰老免疫细胞在发育、衰老或 其他器官如肠道、肺、肝和肾的疾病，代表了Sennet和 更广泛的科学界。

PROJECT SUMMARY Cell senescence in human tissues is an irreversible cell cycle arrest state in otherwise proliferative cells, which is a hallmark of aging that alters the tissue environments via senescence-associated secretory phenotype (SASP) but may also play a beneficial role in tissue remodeling, regeneration, and wound healing. Lymphoid organs play a vital role in hematopoiesis and immune function. How cellular senescence in these tissues is associated with stress-induced or age-related functional impairment, what types or subtypes of senescent cells are present and their spatial heterogeneity and how these cells impact the tissue environments remain poorly understood, precluding the development of strategies to target senescent cells to improve healthspan/lifespan or harnessing these cells or secreted factors to promote tissue remodeling and repair. A recent commentary published in Cell by NIH identified five broad areas (atlases, imaging &visualization, biomarkers, model systems, perturbation and validation) that would help propel the field forward. Our application will assemble a multidisciplinary team to tackle all these areas and specifically, as a Tissue Mapping Center (TMC), will focus on generating the molecular and cellular maps of cellular senescence and associated tissue environments in 4 primary and secondary lymphoid organs. Specifically, it will (a) collect, analyze, annotate, and share high quality non-diseased human primary (bone marrow and thymus) and secondary (tonsil and lymph node) lymphoid tissues, (2) develop and deploy a suite of high-resolution, high-content and high-throughput single- cell & spatial omics technologies to characterize these specimens and paired biofluids, and (3) perform integrated informatics to identify biomarkers of senescent cell heterogeneity and to construct comprehensive molecular and cellular maps of cellular senescence and associated environments in these organs. Four major biological analysis pipelines are: (1) single-cell high-plex (>40) protein secretome profiling, (2) single-cell proteo-transcriptomic sequencing (scCITE-seq), (3) spatial proteo-transcriptomic sequencing at cellular level (DBiT-seq for co-mapping whole transcriptome and a panel of ~300 proteins at cellular level with 10µm pixel size), and (4) spatial molecular imaging (SMI) of ~1,000 molecular targets in FFPE tissues at single-molecule subcellular resolution. With these unique tools, we will (a) characterize functional SASP heterogeneity and identify biomarkers of SAPS in different cell types, (b) construct molecular and cellular maps in 4 human lymphoid tissues, and (c) identify biomarkers of cellular senescence in tissue and the associated environments, contributing to the resource building of SenNet. Since the immune function is central to the physiology of all major organs, our study will provide insights to the role of senescent immune cells in development, aging, or disease in other organs like gut, lung, liver, and kidney, representing a strong synergy within SenNet and the wider scientific community.