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Phenotypic Characterization of Macrophages from Rat Kidney by Flow Cytometry
Published on: October 18, 2016
Ultrastructural and phenotypical diversity of macrophages in the rat ileal mucosa
Youhei Mantani1, Tomohiro Haruta2, Satoki Nakanishi3
1Laboratory of Histophysiology, Department of Bioresource Science, Graduate School of Agricultural Science, Kobe University, 1-1 Rokkodai-cho, Nada-ku, Kobe, Hyogo, 657-8501, Japan. mantani@sapphire.kobe-u.ac.jp.
Insights
This study details macrophage-like cell (MLC) diversity in rat ileal mucosa, revealing distinct ultrastructural and phenotypical differences based on location. Macrophages show varied interactions with intraepithelial lymphocytes (IELs) along the villus-crypt axis.
Area of Science:
- Immunology
- Gastroenterology
- Cell Biology
Background:
- The precise histological localization of diverse macrophage types within the intestinal mucosa remains unclear.
- Understanding macrophage heterogeneity is crucial for deciphering gut immune responses.
Purpose of the Study:
- To elucidate the ultrastructural and phenotypical diversity of macrophage-like cells (MLCs) in the rat ileal mucosa.
- To determine the regional distribution and interactions of MLCs within the intestinal villus and crypt.
Main Methods:
- Immunofluorescent analysis using markers such as CD68, CD163, and CD4.
- Serial block-face scanning electron microscopy (SBF-SEM) for 3D ultrastructural analysis.
- Characterization of MLC types based on morphology, organelle content, and cellular processes.
Main Results:
- Identified three distinct MLC types (I-III) in the ileal mucosa based on ultrastructure and localization.
- CD68+ cells exhibited diverse phenotypes (CD163-, CD4+; CD163+, CD4+; CD163-, CD4-).
- MLCs displayed region-specific localization (villus vs. crypt) and extended cellular processes into the epithelium, with Type I MLCs showing greater interaction with intraepithelial lymphocytes (IELs).
Conclusions:
- Rat ileal macrophages exhibit significant regional heterogeneity in ultrastructure, phenotype, and localization.
- MLCs display distinct interactions with IELs, suggesting specialized roles in different mucosal regions.
- These findings provide a detailed map of macrophage distribution and function in the gut mucosa.
Abstract:
Several types of macrophages have been reported in the intestinal mucosa, but their histological localization remains ambiguous. Here, we obtained detailed information about ultrastructural and phenotypical diversity of macrophage-like cells (MLCs) in the rat ileal mucosa using immunofluorescent analysis and serial block-face scanning electron microscopy (SBF-SEM). The results revealed that the cells immunopositive for CD68, the pan-macrophage marker, included CD163-CD4+, CD163+CD4+, and CD163-CD4- cells in the lamina propria (LP) of the intestinal villus and around the crypt. CD68+CD4+CD163- cells seemed to be preferentially localized in the intestinal villus, whereas CD68+CD163+CD4+ cells were frequently localized around the crypt. SBF-SEM analysis identified three types of MLCs in the ileal mucosa, which were tentatively named types I-III MLC based on aspects of the 3D-ultrastructure, such as the localization, quantity of lysosomes, endoplasmic reticulum, and exoplasm. Type I and II MLCs were localized in the villous LP, while type III MLCs were localized around the crypt, although type II MLCs were a minor population. All three MLC types extended their cellular processes into the epithelium, with type I MLCs showing the greatest abundance of extended processes. Type I MLCs in the upper portion of the intestinal villus showed a higher level of attachment to intraepithelial lymphocytes (IELs) compared to type III MLCs around the crypt. These findings suggest that macrophages of the rat ileal mucosa differed by region along the longitudinal axis of the villous tip-crypt from the perspective of ultrastructure, cellular composition, localization, and interactions with IELs.
