FOXO1 Pathway Restored by Senescence-Resistant Stem Cells to Reverse Primate Reproductive Aging

Male infertility increases with age, and while testicular decline is well-studied, the epididymis — essential for sperm maturation and storage — has received far less attention. This study addresses that gap using a rigorous, multimodal approach in non-human primates to characterize epididymal aging at cellular resolution and identify actionable therapeutic targets.

The researchers analyzed epididymal tissue from young and aged cynomolgus monkeys using histology, bulk transcriptomics, and single-nucleus RNA sequencing (snRNA-seq). Histological analysis revealed age-associated epithelial thinning, luminal dilation, fibrosis, and chronic inflammatory infiltration. Transcriptomic profiling identified widespread upregulation of senescence markers, inflammatory cytokines (SASP), and extracellular matrix remodeling genes, alongside suppression of epididymal secretory and transport functions.

SnRNA-seq resolved the epididymal cellular landscape into distinct epithelial and stromal populations. Principal cells (PCs) — the dominant secretory epithelial type — emerged as the most transcriptionally perturbed with aging. Among downregulated genes in aged PCs, FOXO1, a well-established longevity-associated transcription factor, showed the most striking and consistent decline. Functional experiments in human epididymal epithelial cells confirmed that FOXO1 knockdown induced hallmarks of cellular senescence (p21 upregulation, SA-β-gal activity, DNA damage foci). Mechanistically, the study showed that FOXO1 directly transcriptionally activates LHX1, a homeodomain transcription factor, and that this FOXO1-LHX1 axis is necessary and sufficient to suppress senescence in epididymal epithelial cells.

For therapeutic intervention, the team employed senescence-resistant mesenchymal progenitor cells (SR-MPCs) — a previously developed cell line engineered to resist replicative senescence. Injection of SR-MPCs into aged monkey epididymides led to measurable reductions in senescence burden, fibrosis, and inflammatory signaling, while restoring FOXO1 and LHX1 expression. Importantly, exosomes derived from SR-MPCs recapitulated these effects both in vivo and in vitro, suggesting the therapeutic benefit is mediated at least partly through paracrine signaling rather than direct cell replacement.

This work is notable for establishing the FOXO1-LHX1 axis as a central protective pathway against primate epididymal aging, providing the first cell-type-resolved aging atlas of the primate epididymis, and demonstrating that a clinically translatable cell therapy can partially reverse this aging phenotype. Caveats include the small number of animals studied, the use of non-human primates rather than humans, and uncertainty about the long-term durability and safety of SR-MPC or exosome treatment.