权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

High Resolution Spatial Transcriptomics using seq-FISH+

使用 seq-FISH 进行高分辨率空间转录组学

基本信息

批准号：
10404444
负责人：
John E Harris
金额：
$ 38.09万
依托单位：
UNIV OF MASSACHUSETTS MED SCH WORCESTER
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-09-12 至 2027-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10404444
关键词：
Address Affect Antibodies Atlases Autoimmunity Biological Process Biopsy Bulla Cell Communication Cells Clone Cells Communication Complement Complex Data Dendritic Cells Dermal Epidermis Equilibrium Fluorescent in Situ Hybridization Freezing Gene Targeting Genes Genetic Transcription Goals Image Immune In Situ Individual Inflammation Inflammatory Knowledge Label Lead Lesion Ligands Location Maps Messenger RNA Microscope Modification Normal tissue morphology Organ Pathogenesis Pigments Population Positioning Attribute Privatization Process Punch Biopsy RNA Receptor Cell Research Resolution Signal Transduction Skin Spatial Distribution Structure T cell clonality T cell receptor repertoire sequencing T-Cell Receptor T-Lymphocyte Techniques Technology Tissues Transcript Translational Research Tumor-infiltrating immune cells Vitiligo White Spots cell type cellular imaging design health disparity improved in silico innovation insight interest keratinocyte melanocyte multiple omics receptor receptor expression single cell analysis single-cell RNA sequencing skin disorder skin lesion transcriptome transcriptomics treatment strategy

项目摘要

Much has been learned about mechanisms that drive autoimmunity by studying vitiligo, a dis- ease of the skin that results from destruction of pigment-producing melanocytes. This results in disfiguring white spots that are particularly devastating for those with darker skin, which thus leads to health disparities for the most vulnerable of our population. We generated an extensive set of preliminary data using single cell RNA sequencing (scRNA-Seq) that revealed hun- dreds of conversations that are unique to vitiligo lesions involving every cell type of the epider- mis. These data indicate that cells and their signaling networks within affected and unaffected vitiligo skin are in fact more complex than previously appreciated. Yet because scRNA-Seq re- quires disruption of the tissue, the position of each cell and their communications within the skin remains unknown. We hypothesize that cells connected in silico by matching induced ligand-receptor expression are in close physical proximity in situ, and that T cell clones marked by identical T cell receptor (TCR) sequences associate in physical contact with melanocytes. The objective of the High- Resolution Spatial Transcriptomics Research Core using sequential fluorescence in situ hybridi- zation (seqFISH+) is to use our highly innovative platform to provide spatial context to T cell clonality, cellular positioning, and cell-cell communications revealed by the complementary, syn- ergistic Center Projects 1+2. Our seqFISH+ platform provides a cutting-edge, innovative ap- proach to map RNA expression in tissues with high accuracy and sub-diffraction-limit resolution, allowing for identification of cell classes, expressed genes, and spatial organization in tissues. This approach will be applied in the following specific aims: 1) Identify the position and distribu- tion of T cell clones marked by unique TCR sequences within tissue; and 2) Reveal the spatial organization of communicating cells by receptor-ligand mapping directly within vitiligo lesions. We will use punch biopsies taken from the same vitiligo lesions that have been analyzed by sin- gle cell techniques in each Project. We will design seqFISH+ probes that complement TCR se- quences and ligand-receptor pairs for hybridization, which will indicate the spatial distribution of T cell clonality and cell-cell communications through ligand-receptor pairing. This Research core combines a highly innovative conceptual approach with cutting-edge technology. The prelimi- nary data, investigative team, and innovative plan provides a robust and unique opportunity for discovery of the fundamental mechanisms by which immune cells target self-tissues, which may lead to improved treatment strategies.

通过研究白癜风，我们对驱动自身免疫的机制有了很多了解。由于破坏产生色素的黑素细胞而导致皮肤松弛。这导致毁容的白斑对于深色皮肤的人来说尤其具有破坏性，因此导致我们人口中最弱势群体的健康差距。我们生成了广泛的使用单细胞 RNA 测序 (scRNA-Seq) 的一组初步数据揭示了 hun- 白癜风病变所特有的对话，涉及表皮的每种细胞类型错了。这些数据表明受影响和未受影响的细胞及其信号网络白癜风皮肤实际上比以前想象的更复杂。然而，由于 scRNA-Seq 重新需要破坏组织、每个细胞的位置及其在皮肤内的通讯仍然未知。我们假设细胞通过匹配诱导的配体-受体表达在计算机中连接在原位物理上非常接近，并且 T 细胞克隆由相同的 T 细胞受体标记 (TCR) 序列与黑素细胞发生物理接触。高级别会议的目标使用连续荧光原位杂交的分辨率空间转录组学研究核心 zation (seqFISH+) 是利用我们高度创新的平台为 T 细胞提供空间背景互补性、同步性揭示了克隆性、细胞定位和细胞间通信活力中心项目 1+2。我们的 seqFISH+ 平台提供了尖端、创新的应用程序以高精度和亚衍射极限分辨率绘制组织中 RNA 表达图的方法，允许识别细胞类别、表达基因和组织中的空间组织。该方法将应用于以下具体目标：1）确定位置并分配组织内由独特 TCR 序列标记的 T 细胞克隆； 2）揭示空间通过直接在白癜风病变内进行受体-配体定位来组织通讯细胞。我们将使用取自相同白癜风病变的穿刺活检，这些活检已通过 sin- 每个项目中的gle细胞技术。我们将设计 seqFISH+ 探针来补充 TCR se- 用于杂交的序列和配体-受体对，这将表明序列和配体-受体对的空间分布 T 细胞克隆和通过配体-受体配对进行的细胞间通讯。本研究核心将高度创新的概念方法与尖端技术相结合。预赛- 大量数据、调查团队和创新计划为发现免疫细胞靶向自身组织的基本机制，这可能导致治疗策略的改进。