Phenotypic Heterogeneity During Sperm Capacitation: A Compositional Analysis

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ID: 329508
2026
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Abstract
Mammalian sperm must undergo capacitation, a process marked by substantial phenotypic heterogeneity among sperm. How heterogeneity relates to fertilizing potential remains unresolved, in part because sperm subpopulations are typically analyzed independently despite being intrinsically interdependent. We combine flow cytometry with statistical modeling to quantify how sperm subpopulation compositions respond to a range of capacitation signal intensity in vitro. Using cauda epididymal mouse sperm, we systematically varied extracellular bicarbonate and free calcium-key regulators of capacitation-and classified millions of individual cells into four irreversible physiological states. We show that bicarbonate and calcium interact nonlinearly to redistribute sperm across these subpopulations, revealing responses that would be otherwise obscured in measurements lacking single-cell resolution. Elevated intracellular calcium was associated with increased cell death, while the highest proportions of live, acrosome-reacted sperm occurred under relatively low extracellular calcium conditions. We then applied a hierarchical Dirichlet-multinomial regression model that accounts for multinomial sampling noise and between-male variability, yielding posterior probability surfaces that describe how sperm subpopulations reallocate across functional states as signaling conditions change. We then tested the model predictions using IVF and found that the proportion of live, acrosome reacted sperm was not predictive of early embryonic developmental success. Together, these results suggest that capacitation, though tightly regulated at the single cell scale, is a stochastic process shaped by structured phenotypic heterogeneity at the cell population scale. This framework provides a quantitative foundation for linking sperm subpopulation composition to measures of fertility competence.
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Authors Benjamin M. Brisard, Aishwarya P Halder, Aidan Charles, Amy L. Ward, Maryam Asadi, David Hart, Debajit Bhowmick, Paul de Vos, Cameron A. Schmidt
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Journal biology of reproduction
Year 2026
DOI
10.1093/biolre/ioag207
URL
https://doi.org/10.1093/biolre/ioag207
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