Three-dimensional imaging and computational quantitation as a novel approach to assess nerve fibers, enteric glial cells, mast cells, and the proximity of mast cells to the nerve fibers in human sigmoid mucosal biopsies from healthy subjects

IF 2.7 4区医学 Q2 BIOCHEMICAL RESEARCH METHODS

Journal of Neuroscience Methods Pub Date : 2025-04-01 DOI:10.1016/j.jneumeth.2025.110436

Pu-Qing Yuan , Tao Li , Swapna Mahurkar-Joshi , Jessica Sohn , Lin Chang , Yvette Taché

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引用次数: 0

Abstract

Background

The visualization and quantitation of nerve fibers (NFs), enteric glial cells (EGCs), mast cells (MCs), and their spatial configurations in the human colonic mucosa represent considerable challenges due to the meshed network of these components and the arborizing of NFs in a three-dimensional (3D) structure.

New method

We developed a novel approach combining tissue clearing, 3D imaging and computerized quantitation of NFs, EGCs and MCs in sigmoid mucosal biopsies of healthy subjects using a modified CLARITY tissue clearing protocol and adapting Imaris Surfaces Rendering Technology.

Results

The cleared colonic biopsies are compatible with immunostaining using 10 marker antibodies and capable of generating 3D images rendering clear spatial views and computational quantitation of NFs, MCs, EGCs, in particular the proximity of MCs to NFs with Imaris 9.7–9.9.

Comparison with existing methods

Our modified tissue clearing protocol shortened the membrane lipid removal time to 1 day from the original 1–2 weeks and total tissue clearing time to 3–4 days from the original 2–4 weeks. The 3D images displayed a clear spatial landscape of NFs, MCs and EGCs in the biopsies which cannot be portrayed with 2D images acquired from sections. Computerized quantitation is faster than measuring manually, allowing us to quantify a larger number of samples with less bias.

Conclusion

The novel approach enables faster tissue clearing/immunolabeling, high-quality 3D imaging and precise computational quantitation of NFs, cells and proximity of MCs to NFs in human sigmoid biopsies which may allow new insight to detect alterations in colonic-related diseases.

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来源期刊

Journal of Neuroscience Methods 医学-神经科学

CiteScore

7.10

自引率

3.30%

发文量

226

审稿时长

52 days

期刊介绍： The Journal of Neuroscience Methods publishes papers that describe new methods that are specifically for neuroscience research conducted in invertebrates, vertebrates or in man. Major methodological improvements or important refinements of established neuroscience methods are also considered for publication. The Journal''s Scope includes all aspects of contemporary neuroscience research, including anatomical, behavioural, biochemical, cellular, computational, molecular, invasive and non-invasive imaging, optogenetic, and physiological research investigations.