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. 2021 Aug 25:8:719319.
doi: 10.3389/fvets.2021.719319. eCollection 2021.

Direct but Not Indirect Methods Correlate the Percentages of Sperm With Altered Chromatin to the Intensity of Chromatin Damage

Jordi Ribas-Maynou  1   2 , Marc Llavanera  1   2 , Yentel Mateo-Otero  1   2 , Estela Garcia-Bonavila  1   2 , Ariadna Delgado-Bermúdez  1   2 , Marc Yeste  1   2
Affiliations

Direct but Not Indirect Methods Correlate the Percentages of Sperm With Altered Chromatin to the Intensity of Chromatin Damage

Jordi Ribas-Maynou et al. Front Vet Sci. .

Abstract

Although sperm chromatin damage, understood as damage to DNA or affectations in sperm protamination, has been proposed as a biomarker for sperm quality in both humans and livestock, the low incidence found in some animals raises concerns about its potential value. In this context, as separate methods measure different facets of chromatin damage, their comparison is of vital importance. This work aims at analyzing eight techniques assessing chromatin damage in pig sperm. With this purpose, cryopreserved sperm samples from 16 boars were evaluated through the following assays: TUNEL, TUNEL with decondensation, SCSA, alkaline and neutral sperm chromatin dispersion (SCD) tests, alkaline and neutral Comet assays, and chromomycin A3 test (CMA3). In all cases, the extent of chromatin damage and the percentage of sperm with fragmented DNA were determined. The degree of chromatin damage and the percentage of sperm with fragmented DNA were significantly correlated (p < 0.05) in direct methods (TUNEL, TUNEL with decondensation, and alkaline and neutral Comet) and CMA3, but not in the indirect ones (SCD and SCSA). Percentages of sperm with fragmented DNA determined by alkaline Comet were significantly (p < 0.05) correlated with TUNEL following decondensation and CMA3; those determined by neutral Comet were correlated with the percentage of High DNA Stainability (SCSA); those determined by SCSA were correlated with neutral and alkaline SCD; and those determined by neutral SCD were correlated with alkaline SCD. While, in pigs, percentages of sperm with fragmented DNA are directly related to the extent of chromatin damage when direct methods are used, this is not the case for indirect techniques. Thus, the results obtained herein differ from those reported for humans in which TUNEL, SCSA, alkaline SCD, and alkaline Comet were found to be correlated. These findings may shed some light on the interpretation of these tests and provide some clues for the standardization of chromatin damage methods.

Keywords: DNA fragmentation; SCSA; TUNEL; chromatin; comet assay; sperm.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Correlation matrix between methods regarding the degree of sperm chromatin damage. Figures in each box represent Spearman correlation coefficients. Asterisks and bold figures indicate statistically significant correlations (p < 0.05).
Figure 2
Figure 2
Statistically significant correlations between parameters analyzing the degree of chromatin damage and obtained through different methods.
Figure 3
Figure 3
Correlation matrix between methods regarding the percentages of sperm with chromatin damage. Figures in each box represent Spearman correlation coefficients. Asterisks and bold figures indicate statistically significant correlations (p < 0.05).
Figure 4
Figure 4
Statistically significant correlations between parameters analyzing the percentages of sperm with chromatin damage obtained through different methods.
Figure 5
Figure 5
Correlation matrices of motility parameters with (A) the degree of chromatin damage, and (B) the percentages of sperm with fragmented DNA. Asterisks and bold figures indicate statistically significant correlations (p <0.05).
See this image and copyright information in PMC

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