Testicular aging is marked by declining testosterone, structural deterioration of the seminiferous tubules, and reduced sperm quality, yet the molecular circuits driving this decline have remained poorly defined. A new study published in the Journal of Molecular Cell Biology now provides a high-resolution, multi-omic map of the aging human testis and identifies WLS (Wntless), a Wnt transporter, as a molecule that can actively accelerate senescence in testicular cells. The findings were first reported by ScienceAlert and are detailed in the primary paper by Sun et al. (doi:10.1093/jmcb/mjag001).

What the Researchers Did

The team collected testicular tissue from two young donors (aged 23 and 30) and two older donors (aged 62 and 91), all of whom had fathered children and had undergone orchiectomy for various medical conditions. Using the 10x Genomics Chromium platform, they performed single-nucleus RNA sequencing (snRNA-seq) and single-nucleus ATAC sequencing (snATAC-seq) on the same nuclei—capturing both gene expression and chromatin accessibility from individual cells. After quality filtering, 10,410 high-quality cells were retained for analysis.

Seven major testicular cell types were identified: germ cells (DAZL⁺, DDX4⁺), Sertoli cells (SOX9⁺, WT1⁺), immune cells (PTPRC⁺), myoid cells (PTHC1⁺, MYH11⁺), endothelial cells (FLT1⁺, VWF⁺), and two Leydig cell subtypes—Leydig1 (IGF1⁺, DLK1⁺, immature) and Leydig2 (CYP11A1⁺, CYP17A1⁺, adult). As expected, intratesticular testosterone was significantly higher in young donors, and aged seminiferous tubules showed basement membrane thickening and germ cell depletion.

Spermatogonial Stem Cells Decline with Age

Germ cells were subdivided into seven stages along the spermatogenesis trajectory, including two spermatogonial stem cell (SSC) populations: SSC1 (ZBTB16⁺/PLZF⁺) and SSC2 (GFRA1⁺). Aged SSC1 showed 28 upregulated and 22 downregulated genes; aged SSC2 showed 41 upregulated and 26 downregulated genes. Downregulated pathways in SSC1 were enriched for spermatogenesis, while SSC2 downregulated genes clustered in G protein-coupled receptor signaling.

Integrating ATAC-seq with expression data, the team identified key transcription factors—NR2C2, NRF1, NKX6-1, and MYBL1—whose regulatory activity declined with age. Chromatin accessibility at their downstream targets, including FNDC3A and DLGAP2, was also reduced, suggesting an epigenetic contribution to SSC aging.

Sertoli Cells Are Highly Sensitive to Aging

Sertoli cells from different donors clustered into distinct groups despite sharing common marker genes, indicating that these nurse cells are “highly dynamic and particularly responsive to changes in the testicular microenvironment.” Aged Sertoli cells showed dysregulated genes enriched for actin filament–based movement and cell adhesion—processes critical for the blood–testis barrier (BTB). Consistent with this, the tight-junction protein ZO-1 was significantly reduced in aged testes, as confirmed by Western blot and immunofluorescence, pointing to BTB disruption and increased immune infiltration with age.

The WLS Discovery

By comparing differentially expressed genes across three independent human testis aging datasets (this study, Huang et al. 2022, and Nie et al. 2022), the team identified shared aging signatures in Sertoli cells. Among the most consistent was WLS (also known as GPR177 or Evi), a conserved membrane protein required for Wnt secretion. WLS was primarily expressed in Sertoli and Leydig cells and was significantly upregulated in aged Sertoli cells, a finding confirmed by fluorescence in situ hybridization (FISH).

The chromatin data added a mechanistic layer: aged samples showed enhanced ATAC-seq peaks within the chr1:68192900–68193300 interval of the WLS locus, indicating aging-related epigenetic remodeling at WLS enhancer sites. Downstream Wnt/β-catenin target genes, such as AXIN2, also showed more open chromatin at their regulatory regions in aged Sertoli cells, linking elevated WLS to potentiated Wnt signaling.

WLS Overexpression Accelerates Senescence

To test causality, the researchers overexpressed WLS in cultured Sertoli cells. The WLS-overexpressing cells showed higher SA-β-Gal positivity and increased protein levels of aging markers P16, P21, and γ-H2A. They also secreted significantly less Inhibin B, a hormone produced by healthy Sertoli cells. Together, these results suggest that WLS is not merely a marker of aging but can actively promote senescence in the testicular niche.

Leydig Cells and Disrupted Cell Communication

Leydig cells showed 496 upregulated and 254 downregulated genes with age. Downregulated genes were enriched for cell adhesion, while upregulated genes clustered in cell differentiation, growth hormone signaling, and apoptosis. Pseudotime analysis revealed distinct differentiation trajectories: young Leydig cells progressed from LeydigB to LeydigE, whereas old cells followed a LeydigA-to-LeydigD path. Key transcription factors with altered activity included MTF2 and ETV1 in Leydig2 cells, ZNF521 across Leydig cells, and SOX5 and MEIS2 in Leydig1 cells.

Cell–cell communication analysis using CellChat showed that both the number and strength of interactions diminished with age. Activin signaling from Leydig1 cells to Sertoli cells was reduced, while TGF-β signaling—a driver of fibrosis and inflammation—increased across nearly all cell types in aged donors. Ligand–receptor pairs INHBA–(ACVR1B+ACVR2A) and TGFB1–(ACVR1+TGFBR1) were both reduced in aged tissue, and qPCR confirmed downregulation of TGFBR1, ACVR1B, and ACVR2A. Sertoli-to-germ-cell signaling (FGF1–FGFR2, PTN–PTPR21) also declined, as did Leydig interactions such as GAS6–AXL and LGR4–RSPO3.

Other Somatic Cells Also Change with Age

• Endothelial cells: Five differentially expressed genes—FLT1, FOS, and LIFR upregulated; MPP7 and ZNF521 downregulated. FOS is implicated in cellular senescence, suggesting pro-aging signaling in the vasculature.
• Myoid cells: Upregulation of ACTA2, MYH11, TAGLN, FHL2, JUN, and RGN, with enrichment for smooth muscle contraction and fibroblast activation—consistent with increased contractility and potential fibrosis. ERBB4, linked to sperm motility, was downregulated.
• Immune cells: Macrophages showed upregulation of HERPUD1, an inflammatory-response gene, suggesting stronger inflammation in aged testicular tissue.

Why This Matters

Previous work on testicular aging was largely descriptive. This study moves toward mechanistic causality by combining unbiased multi-omic profiling with functional validation. The convergence of elevated WLS expression in aged Sertoli cells, epigenetic opening at WLS enhancers, and WLS-driven senescence in vitro makes a compelling case that Wnt/WLS signaling is an active contributor to the aging phenotype.

The authors place their finding in the context of prior mouse work: conditional deletion of Gpr177 (the mouse orthologue of WLS) in germ cells caused testicular atrophy and compromised spermatogenesis in aged mice, whereas Sertoli-specific knockout did not impair fertility in young or aged mice. That contrast, the authors suggest, raises the possibility that WLS knockout in Sertoli cells may offer a protective effect against age-related deterioration—positioning WLS as a potential therapeutic target for preserving testicular function during aging.

Limitations

The authors are candid about the study's constraints. The multi-omic analysis rests on a small sample of four donors, reflecting the practical and ethical difficulty of obtaining high-quality aged human testicular tissue. The detailed downstream mechanisms by which WLS promotes aging were not extensively investigated, and the team calls for future work to develop WLS inhibitors and evaluate their potential to intervene in testicular aging.

Read the Primary Study and the News Coverage

Primary source: Sun, W., Liu, H., Ren, G., Pang, P., Feng, Z., Wang, L., Piao, S., He, M., Shen, H., Li, J., Li, X., Yang, C., Wang, L., and Liu, Z. “Integrated single-nucleus transcriptomic and chromatin accessibility analysis reveals key molecular basis of human testicular aging.” Journal of Molecular Cell Biology, mjag001, published 09 March 2026. https://doi.org/10.1093/jmcb/mjag001

News summary: Scientists Discover a Potential Driver of Aging in The Testicles — ScienceAlert