Improvement and standardization of a bioinformatic software suite for multiplexed imaging

项目总结 骨质疏松症，或与年龄相关的肌肉萎缩，困扰着15%的老年人，严重降低生活质量和 发病率和死亡率不断增加。我们的首要目标是阐明石棺减少症的新致病机制。 并利用这些知识来改善衰老的肌肉功能。我们建议利用我们的发现， 15-羟基前列腺素分解代谢导致衰老肌肉中前列腺素PGE2和PGD2的减少 腺苷脱氢酶(15-PGDH)，前列腺素降解酶，是该项目的父母赠款的对象 申请。正如在授予中所描述的，我们得到了意想不到的发现，15-PGDH的表达显著 在老化的小鼠和人的肌肉中增加。此外，我们的数据显示，在体内过表达15-PGDH 年轻的肌肉会导致衰老的表型，伴随着肌肉质量和功能的显著下降， 模仿石棺减少症。父母赠款的中心假设是，在衰老、衰老和炎症期间- Tory细胞聚集在肌肉微环境中，表达15-PGDH，它能降解PGE2和PGD2， 并导致肌肉萎缩。因此，抑制衰老肌肉中的15-PGDH将增加PGE2和PGD2 脂肪代谢和增加肌肉质量和力量。我们的目的是(I)阐明PGE2和PGE2的作用 骨骼肌内稳态中的前列腺素D2，(II)15-前列腺素脱氢酶和前列腺素失调的细胞来源 通过抑制15-前列腺素脱氢酶来恢复衰老肌肉的功能和质量。家长资助目标2 采用尖端的单细胞多路成像技术，称为CODEX(通过索引进行CO检测)。 CODEX允许我们在单个组织切片中同时解析多达60个标记。这种突破性的冰毒- 气味学将使我们能够确定衰老细胞是否包含15-PGDH的细胞来源，并允许我们 以解决衰老肌肉中存在的不同细胞类型之间的空间关系。每个食典实验- MENT生成数TB的高分辨率图像，必须将这些图像去卷积并缝合在一起。我们有 开发了3个软件包(CRISP、HFCLUSTER和SPACE)来进行去卷积、缝合、集群和注释 我们的CODEX数据和计算识别细胞类型之间的信号。我们对这份补充的目标是资助 技术娴熟且敬业的实验室助理，负责本软件的持续改进和维护，具体如下： 改进用户界面，标准化数据输入格式，并为一系列多个 错综复杂的成像工作流程。这些改进不仅将帮助我们实现赠款的第二个目标，而且 它们还将使我们开发的软件更加用户友好，对肌肉生物学来说也更容易访问 并向所有使用CODEX和其他多路成像模式的用户开放。

PROJECT SUMMARY Sarcopenia, or age-related muscle atrophy, afflicts 15% of the elderly, severely diminishing quality of life and increasing morbidity and mortality. Our overarching goal is to elucidate novel causal mechanisms of sarcopenia and use this knowledge to improve aged muscle function. We propose to capitalize on our discovery that a reduction in the prostaglandins PGE2 and PGD2 in aged muscle results from catabolism by 15-hydroxyprosta- glandin dehydrogenase (15-PGDH), the prostaglandin degrading enzyme, the subject of the parent grant of this application. As described in the grant, we made the unexpected finding that 15-PGDH expression is significantly increased in aged mouse and human muscles. Additionally, our data show that overexpressing 15-PGDH in young muscles induces an aging phenotype accompanied by a marked decrease in muscle mass and function, mimicking sarcopenia. The central hypothesis of the parent grant is that during aging, senescent and inflamma- tory cells accumulate in the muscle microenvironment and express 15-PGDH, which degrades PGE2 and PGD2, and causes muscle wasting. Therefore, inhibition of 15-PGDH in aged muscles will increase PGE2 and PGD2 lipid metabolites and augment muscle mass and strength. Our aims are to (i) elucidate the roles of PGE2 and PGD2 in skeletal muscle homeostasis, (ii) identify the cell source of 15-PGDH and prostaglandin dysregulation in aged muscle, and (iii) restore aged muscle function and mass by inhibiting 15-PGDH. Parent grant Aim 2 employs a cutting-edge single cell multiplexed imaging technology called CODEX (CO-Detection by indEXing). CODEX allows us to resolve up to 60 markers simultaneously in single tissue sections. This breakthrough meth- odology will enable a determination of whether senescent cells comprise a cell source of 15-PGDH and allow us to resolve the spatial relationships among the diverse cell types present in aged muscles. Each CODEX experi- ment produces terabytes of high-resolution images that must be deconvolved and stitched together. We have developed 3 software packages (CRISP, HFCluster, and SpaCE) to deconvolve, stitch, cluster, and annotate our CODEX data and computationally identify signaling among cell types. Our goal for this supplement is to fund a skilled and dedicated lab associate for the continued improvement and maintenance of this software, specifi- cally to improve user interface, standardize data input formats, and provide compatibility for a range of multi- plexed imaging workflows. These improvements will not only help us achieve the second aim of our grant, but they will also make the software we develop far more user-friendly and readily accessible to the muscle biology and aging communities, as well as to all users of CODEX and other multiplexed imaging modalities.