Molecular and immunohistochemical characterization of intestinal macrophages subsets in goldfish

Macrophage-colony stimulating factor (M-CSF) plays a crucial role in the proliferation and differentiation of the monocyte-macrophage lineage across vertebrates. In this study, we identified and characterized two CSF1 paralogues and their corresponding receptor genes in the intestine of the goldfish (Carassius auratus), showing sequence homology with known teleost species. Immunohistochemical analysis confirmed the presence of CSF, its receptor CSF-R1, and bone morphogenetic protein 2 (BMP2) in intestinal macrophages. These macrophages were localized within mucosal, submucosal, and muscularis layers, suggesting distinct functional subtypes. Quantitative PCR analysis revealed differential gene expression patterns, with csf1a, csf1b, and csf1ra highly expressed in the brain, while csf1rb transcripts were predominant in the intestine. Immunophenotypic characterization using CD14 and CD86 markers further demonstrated macrophage heterogeneity. Additionally, BMP2-expressing macrophages were observed in the muscularis externa, implying a potential role in neuromuscular regulation. These findings provide novel insights into the molecular and immunohistochemical profiles of goldfish intestinal macrophages, highlighting their potential role in immune responses and gut homeostasis.