Multiplexed fluorescence microscopy reveals heterogeneity among stromal cells in mouse bone marrow sections
Multiplexed fluorescence microscopy reveals heterogeneity among stromal cells in mouse bone marrow sections
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DOI：
10.1002/cyto.a.23526
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发表时间：
2018-09-01
期刊：
影响因子：
3.7
通讯作者：
Hauser, Anja E.
中科院分区：
文献类型：
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作者：
Holzwarth, Karolin;Koehler, Ralf;Hauser, Anja E.
骨髓（BM）由多个结构化的微环境实体组成，即所谓的壁龛，其中包含造血细胞和基质细胞。这些小生境具有多种功能，如控制造血干细胞池、造血细胞分化和维持免疫记忆等。然而，由于分子和细胞的复杂性以及缺乏合适的组织学多路复用方法，各种BM生态位的组成仍然难以捉摸。在这项研究中，我们应用了多表位配体制图（MELC）对小鼠骨切片。我们将多路免疫荧光组织学数据与各种基于对象的分割方法相结合，以定义不规则形状的网状基质细胞结构。我们证实MELC是一种强大的组织学方法，并使用流式细胞术和人工评估验证了我们的自动分割算法。通过MELC多路复用，我们揭示了瘦素受体（LpR）、BP-1和VCAM-1在基质网络中的异质表达。此外，我们通过定量证明了B细胞亚群和浆细胞与表达cxcl12的基质细胞过程的优先接触，与基质体细胞相比。总之，我们的方法适用于复杂组织结构的空间分析。
The bone marrow (BM) consists of multiple, structured micro-environmental entitiesthe so called niches, which contain hematopoietic cells as well as stromal cells. These niches fulfill a variety of functions, such as control of the hematopoietic stem cell pool, differentiation of hematopoietic cells, and maintenance of immunological memory. However, due to the molecular and cellular complexity and a lack of suitable histological multiplexing methods, the composition of the various BM niches is still elusive. In this study, we apply multiepitope-ligand-cartography (MELC) on bone sections from mice. We combine multiplexed immunofluorescence histology data with various object-based segmentation approaches in order to define irregularly shaped, net-like structures of stromal cells. We confirm MELC as a robust histological method and validate our automated segmentation algorithms using flow cytometry and manual evaluation. By means of MELC multiplexing, we reveal heterogeneous expression of leptin receptor (LpR), BP-1, and VCAM-1 in the stromal network. Moreover, we demonstrate by quantification a preferential contact of B cell subsets as well as of plasma cells to processes of CXCL12-expressing stromal cells, compared with stromal somata. In summary, our approach is suitable for spatial analysis of complex tissue structures.