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DNA fragmentation in concert with the simultaneous assessment of cell viability in a subfertile population: establishing thresholds of normality both before and after density gradient centrifugation

  • Gamete Biology
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Journal of Assisted Reproduction and Genetics Aims and scope Submit manuscript

Abstract

Purpose

TUNEL assay is the most common, direct test for sperm chromatin integrity assessment. But, lack of standardized protocols makes interlaboratory comparisons impossible. Consequently, clinical thresholds to predict the chance of a clinical pregnancy also vary with the technique adopted. This prospective study was undertaken to assess the incidence of sperm DNA fragmentation in a subfertile population and to establish threshold values of normality as compared to a fertile cohort, both before and after density gradient centrifugation in the total and vital fractions.

Method

Men presenting at a university hospital setup for infertility treatment. DNA damage via TUNEL assay was validated on fresh semen samples, as conventional semen parameters, to reduce variability of results.

Results

Total DNA fragmentation in the neat semen was significantly higher in the subfertile group, but the vital fraction was not significantly different between the two cohorts. After gradient centrifugation, DNA fragmentation increased significantly in the total fraction of the subfertile group but decreased significantly in the vital fraction. In the fertile cohort, there was a non-significant increase in total fragmentation and in the vital fraction the trend was unclear.

Conclusions

Estimating total and vital sperm DNA fragmentation, after density gradient centrifugation, increased both the sensitivity and the specificity, thereby lowering the number of false negatives and false positives encountered. These findings provide opportunities to investigate the significance of the total and the vital fractions after different assisted reproductive technologies.

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Acknowledgments

The authors acknowledge Christine Croes, Els Smet, Julie Biltjes, and Zehra Kara for preparing the sperm samples.

Funding

This work was supported by funds from the Centre for Reproductive Medicine, Antwerp University Hospital and by the Fonds Wetenschappelijk Onderzoek (FWO), Research Foundation Flanders (grant number T007016N).

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Authors and Affiliations

  1. Centre for Reproductive Medicine, Antwerp University Hospital, Wilrijkstraat 10, 2650, Edegem, Belgium

    U. Punjabi, H. Van Mulders, I. Goovaerts, K. Peeters & D. De Neubourg

  2. University of Antwerp, Universiteitsplein 1, 2610, Wilrijk, Belgium

    U. Punjabi & D. De Neubourg

  3. Clinical Trial Center (CTC), CRC Antwerp, Antwerp University Hospital, University of Antwerp, Wilrijkstraat 10, 2650, Edegem, Belgium

    E. Roelant

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  1. U. Punjabi
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Contributions

U.P. substantial contributions to conception, acquisition of data, analysis and interpretation of data, drafting the article. D.D.N. revising it critically for important intellectual content. H.V.M. and I.G. conducted the DNA damage analyses. E.R. was involved in statistical analysis and interpretation of data. All authors revised and approved the final version of the manuscript.

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Correspondence to U. Punjabi.

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Punjabi, U., Van Mulders, H., Goovaerts, I. et al. DNA fragmentation in concert with the simultaneous assessment of cell viability in a subfertile population: establishing thresholds of normality both before and after density gradient centrifugation. J Assist Reprod Genet 36, 1413–1421 (2019). https://doi.org/10.1007/s10815-019-01476-z

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