Oral Presentation ESA-SRB-ANZOS 2025 in conjunction with ENSA

Mapping the effects of chronic and acute activin A elevation on spermatogonial fate throughout life: implications for male fertility (127675)

Penny A. F. Whiley 1 2 , Benedict Nathaniel 2 , Robin M Hobbs 1 2 , Kate L Loveland 1 2
  1. Department of Molecular and Translational Sciences, SubFaculty of Clinical and Molecular Medicine. Monash University, Clayton, VIC, Australia
  2. Centre for Endocrinology and Reproductive Health, Hudson Institute of Medical Research, Clayton, VIC, Australia

This study addresses the hypothesis that the risk of developing testicular cancer and/or male infertility is heightened by exposure to elevated activin A (AA), which occurs in pathological conditions throughout life (pre-eclampsia1, infections2, cachexia3). AA influences somatic and germ cell proliferation and viability, limits steroid production, and governs somatic cell differentiation in fetal mouse testes4,5,6,7. Here we examine AA cell-specific actions in fetal, neonate and adult testes, and highlight consequences for germ cell development.

We performed single-cell RNAseq (scRNAseq) on testes from E13.5, E15.5, P0 and P6 (n=2/age/genotype) InhaKO mice (chronic unopposed/elevated AA signalling8). To study acute effects, E17.5 WT testis fragments were cultured (72hr) with 50ng/mL AA or 10µM SB431542 (activin/TGFβ/Nodal inhibitor; n=5-7 fragments/treatment). Spermatogonia were examined using immunofluorescence and RNAseq in neonate (P0, P3, P6; n>3/age/genotype) and adult InhaKO testes, and cultured adult WT undifferentiated spermatogonia (5, 25ng/ml AA or BMP4, 6/24hr; n=4/treatment/timepoint).

InhaKO testes lose 50% of fetal germ cells by E15.5. Amongst surviving germ cells, scRNAseq identified cell cycle defects (Ccnd2 & Cdk1 retention) at E15.5 and premature cell cycle re-entry (Mki67+) at P0. Evidence of advanced development includes early expression of differentiation genes (Stra8, Sohlh2) and higher germ cell migration in E17.5 testis cultured with AA. At P6, when the SSC population is fully established, more spermatogonia in InhaKO testes were GFRA1+, indicating enhanced SSC formation. Surprisingly, SSC-associated transcripts were unaltered in AA-cultured spermatogonia, while BMP4 elevated several (Id1-3), and altered Wnt signalling potential. A greater abundance and proliferation of SSCs in adult InhaKO testes suggests chronic AA elevation promotes a niche that supports long-term SSC self-renewal.

These data indicate elevated AA exposure in early life may increase the risk of testicular cancer/infertility by disrupting germ cell maturation. Ongoing scRNAseq interrogation is focussed on revealing candidate niche factors regulated by AA that determine spermatogonial fate.

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  7. 7. Whiley, Luu et al. Front Endocrinol (2023)
  8. 8. Matzuk et al. Nature (1992)