Imperial College London, London, UK.
The Francis Crick Institute, London, UK.
Methods Mol Biol. 2021;2308:203-222. doi: 10.1007/978-1-0716-1425-9_16.
Haematopoietic stem cells (HSCs) are instrumental in driving the generation of mature blood cells, essential for various functions including immune defense and tissue remodeling. They reside within a specialised bone marrow (BM) microenvironment , or niche, composed of cellular and chemical components that play key roles in regulating long-term HSC function and survival. While flow cytometry methods have significantly advanced studies of hematopoietic cells, enabling their quantification in steady-state and perturbed situations, we are still learning about the specific BM microenvironments that support distinct lineages and how their niches are altered under stress and with age. Major advances in imaging technology over the last decade have permitted in-depth studies of HSC niches in mice. Here, we describe our protocol for visualizing and analyzing the localization, morphology, and function of niche components in the mouse calvarium, using combined confocal and two-photon intravital microscopy, and we present the specific example of measuring vascular permeability.
造血干细胞（HSCs）在驱动成熟血细胞的生成中起着重要作用，这些血细胞对于包括免疫防御和组织重塑在内的各种功能至关重要。它们存在于一个专门的骨髓（BM）微环境中，或称为龛位，由细胞和化学组成部分组成，这些组成部分在调节长期 HSC 功能和存活方面发挥着关键作用。尽管流式细胞术方法极大地推动了造血细胞的研究，使其能够在稳态和扰动情况下进行定量分析，但我们仍在了解支持不同谱系的特定 BM 微环境以及它们的龛位在压力下和随着年龄的增长如何发生变化。过去十年中成像技术的重大进步使得能够在小鼠中深入研究 HSC 龛位。在这里，我们描述了一种使用共聚焦和双光子活体显微镜可视化和分析小鼠颅盖骨中龛位成分的定位、形态和功能的方案，并展示了测量血管通透性的具体示例。
