High Resolution Spatial Transcriptomics using seq-FISH+

通过对白癜风的研究，人们已经了解了许多关于驱动自身免疫的机制。 由于破坏产生色素的黑素细胞而导致的皮肤松弛。这将导致 使白色斑点毁容，对深色皮肤的人特别有害，因此 导致我国人口中最脆弱群体的健康差距。我们产生了一个广泛的 一组使用单细胞RNA测序(scRNA-Seq)的初步数据揭示了人类- 白癜风皮损特有的对话，涉及每种细胞类型的表皮- 失踪者。这些数据表明，受影响和未受影响的细胞及其信令网络 事实上，白癜风皮肤比之前想象的要复杂得多。然而，因为scRNA-Seq Re- 停止破坏组织、每个细胞的位置以及它们在皮肤内的通讯 仍然不为人知。 我们假设，细胞通过匹配诱导的配体-受体表达而在电子计算机中连接 在原位物理上非常接近，且T细胞克隆标记有相同的T细胞受体 (TCR)序列与黑素细胞的物理接触有关。高等法院的目标是- 序列荧光原位杂交分辨率空间转录组研究核心 SeqFISH+是使用我们高度创新的平台为T细胞提供空间背景 细胞克隆、细胞定位和细胞间的通讯由互补的、同源的、 ERGISTIC中心项目1+2。我们的seqFISH+平台提供尖端、创新的AP- 以高精度和亚衍射极限分辨率绘制组织中RNA表达图谱的方法， 从而能够识别组织中的细胞类别、表达的基因和空间组织。 这一方法将应用于以下具体目标：1)确定位置和分布- 组织内标记有独特TCR序列的T细胞克隆的检测；以及2)揭示了 白癜风皮损内直接通过受体-配体映射的通讯细胞的组织。 我们将使用从相同的白癜风皮损中提取的穿孔活检，这些皮损已经被SIN- 每个项目中的单元格技术。我们将设计seqFISH+探针来补充TCR se- 序列和配基-受体对的杂交，这将表明空间分布 T细胞的克隆性和通过配体-受体配对的细胞-细胞通讯。本研究的核心 将高度创新的概念方法与尖端技术相结合。开场白： 几乎没有数据、调查团队和创新计划提供了强大而独特的机会 发现免疫细胞攻击自身组织的基本机制，这可能 导致改进治疗策略。

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.