3D imaging of nerves, glia, and mast cells in human sigmoid mucosal biopsies
This Learning Wednesday paper note highlights “Three-dimensional imaging and computational quantitation of nerve fibers, glial cells, and mast cells in human sigmoid mucosal biopsies.” The study is relevant to 3D quantification of nerves, glia, and mast cells, as well as spatial proximity, in gut biopsies, with a focus on how three-dimensional tissue context can alter what researchers see and measure.
Selected notes from the paper
“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.”
“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.”
“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.”
“3D imaging was performed using confocal microscopy on 200-μm tissue sections with a 708 × 708 μm field of view, acquired as Z-stacks spanning 150–200 μm in depth.”
“We developed a new protocol applicable for clearing human sigmoid mucosal biopsies and compatible with immunostaining using marker antibodies for NFs from extrinsic sympathetic, parasympathetic, and sensory neurons and intrinsic enteric nervous system (ENS), as well as EGCs and MCs.”
“A computerized approach was established for digital segmentation and computational quantification of NFs, MC, and EGC density as well as the shortest distances of each MC to NFs in 3D images of the whole biopsy tissue samples.”
“These computerized measurements not only reduce biases due to observer/examiner judgment and overcome limitations of 2D images but also are much faster than measuring manually and facilitate the quantification of a large number of samples, increasing statistical accuracy.”
“This new tool for the quantitative assessment of innervation and interactions of MCs and NFs can be applied to biopsies of patients with chronic visceral pain disorders, such as IBS, and has the potential to bring new insight for diagnostic and therapeutic purposes.”
From an Alpenglow perspective, this paper is useful because it connects 3D quantification of nerves, glia, and mast cells, along with spatial proximity, in gut biopsies to a broader need in 3D spatial biology: measuring tissue architecture across depth while preserving context for quantitative analysis.
