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Application of Flow Cytometry Using Advanced Chromatin Analyses for Assessing Changes in Sperm Structure and DNA
Estíbaliz Lacalle1,2, Estela Fernández-Alegre2, Belén Gómez-Giménez1
1Institute of Animal Health and Cattle Development (INDEGSAL), University of León, 24071 León, Spain.
International Journal of Molecular Sciences
|February 24, 2024
Summary
Assessing pig sperm chromatin structure using multivariate analysis reveals storage impacts DNA fragmentation and protamine status. This comprehensive approach offers insights beyond simple DNA damage assessment for fertility prediction.
Area of Science:
- Reproductive Biology
- Spermatozoa Analysis
- Chromatin Structure
Background:
- Sperm chromatin integrity is crucial for fertility and reflects spermatogenesis.
- Assessing pig sperm chromatin complexity requires advanced multivariate approaches.
- Standard assessments often focus solely on DNA damage, neglecting other critical factors.
Purpose of the Study:
- To evaluate a multivariate approach for assessing pig sperm chromatin structure.
- To analyze the impact of storage on various sperm chromatin parameters.
- To identify key indicators of sperm nucleus status for fertility prediction.
Main Methods:
- Analyzed sperm doses from 36 boars over 11 days of storage at 17 °C.
- Utilized CASA for motility, and flow cytometry for SCSA (%DFI, %HDS), monobromobimane (mBBr), chromomycin A3 (CMA3), and 8-oxo-dG.
- Employed linear models, hierarchical clustering, and principal component analysis (PCA).
Main Results:
- Storage reduced motility and negatively impacted chromatin status (%DFI, 8-oxo-dG increased; mBBr decreased).
- Boar significantly affected most chromatin variables, while storage affected most except %HDS.
- Multivariate analysis showed mBBr significantly influenced variance, particularly with storage; SCSA and 8-oxo-dG affected between-sample variability.
Conclusions:
- A combination of DNA fragmentation, oxidation, protamination, and tiol status provides a comprehensive view of pig sperm nucleus status.
- CMA3 was the least informative parameter in this study.
- Further research is needed to validate this multivariate approach for predicting fertility in pigs.
