Background: Spermatogonial stem cells (SSCs) hold the potential to be used for ground-breaking technologies, spanning from infertility treatments for childhood cancer survivors to biobanking for endangered species conservation. Progress in these areas requires a better understanding of how SSC fate is regulated. In this study we aimed to characterise the unique properties of SSC metabolism, which we hypothesise actively influences the regenerative capacity of germline stem cells.
Methods: An Id4-eGfp transgenic mouse line was used to capture populations of mouse SSCs and progenitor spermatogonia. Mitochondrial respiratory capacity and glycolytic flux were measured using Seahorse bioanalysis (n=4). Mitochondrial ultrastructure and cristae architecture were assessed using Transmission Electron Microscopy, and metabolic and mitochondrial protein expression was quantified by proteomics (n=4). Statistical analyses were performed using a Student’s t-test or Mann–Whitney test.
Results: The SSC-to-progenitor transition was accompanied by substantial mitochondrial remodelling. SSC mitochondria exhibited wider and less elongated cristae (p<0.001) at a lower density than progenitors (p<0.001), consistent with a less structurally mature mitochondrial state and reduced inner-membrane area available for electron transport in vivo. Proteomic comparison revealed coordinated differences in expression of proteins regulating mitochondrial dynamics, cristae junction composition, and glycolytic flux, with enrichment of proteins involved in pyruvate-lactate interconversion forecasting plasticity in SSCs. Strikingly, bioenergetic analyses revealed that, when released from their hypoxic niche into normoxia, SSCs display markedly greater basal and maximal respiration and higher spare respiratory capacity than progenitors (p<0.05), while maintaining comparable glycolytic flux. These finding suggest that SSCs are metabolically poised to interconvert between glycolysis and oxidative phosphorylation in response to environmental cues.
Conclusion: Metabolic plasticity is a previously unrecognised feature of SSCs. The intrinsic respiratory reserve of these stem cells may support transitions between self-renewal and differentiation during germline regeneration. This process thus represents a potential target for advancing SSC-based reproductive and conservation technologies.