Single-cell immune atlas of mouse testes unveils metabolic reprogramming of FOLR2 + macrophages in orchestrating testicular immunity during aging.

Immune cells play a crucial role in maintaining tissue homeostasis during aging. However, the dynamics and functions of immune cells in testicular aging have not been well elucidated. In this study, we utilized single-cell RNA sequencing (scRNA-seq) to analyze CD45-enriched immune cells isolated from young and old mice testis. This approach yielded a comprehensive dataset comprising 6622 immune cells, encompassing macrophages, monocytes, and T cells. Our analysis revealed a significant decline in FOLR2 + resident macrophages, accompanied by a corresponding increase in pro-inflammatory CD74 + macrophages, CCR2 + monocytes, and CD8 + T cells in old mice testis. These findings were further validated by multiplex immunofluorescence staining. Notably, during testicular aging, FOLR2 + macrophages underwent a phenotypic transition towards a pro-inflammatory state. This transition subsequently facilitated the recruitment of monocytes and CD8 + T cells via the CCL8-CCR2/CCR5 axis. Furthermore, we discovered that mitochondrial metabolic dysfunction was a key driver of FOLR2 + macrophage activation. Specifically, inhibition of IDH2, a key catalytic enzyme in the TCA cycle, significantly induced this activation. Collectively, our findings provide a detailed immune atlas of testicular aging and suggest a potential role for FOLR2 + macrophages in maintaining testicular immune homeostasis.