Microglia Immunophenotyping in Paraffin Sections of the Brain

期刊论文
发表日期：

2025年9月30日

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V. A. Razenkova,
O. V. Kirik,
D. E. Korzhevskii

摘要

Abstract

Objective: In neuroinflammation, microglia acquire structural and functional features that complicate its detection and analysis. Moreover, none of the well-established microglial markers on its own is capable to provide comprehensive information about the functional immune status of these cells. The aim of the study was to investigate the possibility of characterizing the microglia immunophenotypes using various functional markers (Iba-1, CD68 and vimentin). Materials and methods: The material for the study was the brain of Wistar and SHR rats (n = 8). Iba-1, P2Y12 and vimentin were used as marker proteins of microglia in a comparative immunohistochemical study. Results and discussion: It was shown that typical resting microglia lacked CD68 and vimentin expression, indicating the absence of active phagocytosis. The cells from those areas of the brain where phagocytosis of pathogens and cellular debris is especially necessary had Iba-1⁺/CD68⁺ phenotype. The simultaneous presence of Iba-1 and vimentin is typical for sub- and supraependymal cells of the lateral and third ventricles. It can be expected that some cells of the third ventricle floor are Iba-1⁺/CD68⁺/vimentin⁺, as evidenced by the analysis of the morphology and localization of both Iba-1⁺/CD68⁺ and Iba-1⁺/vimentin⁺ microglia. The putative polarization type of activated Iba-1⁺/CD68⁺ and Iba-1⁺/vimentin⁺ microglial cells is M2a. It is evidenced not only by the protein markers expression, but also by the cellular morphology of activated, rather than amoeboid microglia. Conclusions: Using confocal laser scanning microscopy, it was shown that double Iba-1/CD68 and Iba-1/vimentin immunofluorescence allows highly efficient characterization of various phenotypes of microglia and macrophages in the rat brain. The combination of different functional markers for a double immunofluorescence in conjunction with morphological analysis allows one to evaluate the activity and putative functions of a particular microglia phenotype.