Oxidative DNA damage in human sperm can be detected by Raman microspectroscopy

Capsule:
Raman microspectroscopy was able to identify oxidative sperm nDNA damage. The differences in spectral profiles were corroborated by Fourier transform infrared spectroscopy, and the levels of fragmentation were consistent with flow-cytometric assessment.

Authors:
Victoria Sánchez, B.Sc, Klaus Redmann, Ph.D., Joachim Wistuba, Ph.D., Frank Wübbeling, Ph.D., Martin Burger, Ph.D., Harriette Oldenhof, Ph.D., Willem F. Wolkers, Ph.D., Sabine Kliesch, M.D., Stefan Schlatt, Ph.D., Con Mallidis, Ph.D.

Volume 98, Issue 5, Pages 1124-1129.e3, November 2012

Abstract:

Objective:
To determine whether Raman microspectroscopy can identify different levels of oxidative sperm nDNA damage and to corroborate the findings using an established method and an alternative but complementary spectroscopic technique.

Design:
Three-way comparison of Raman profiles, FTIR spectra and flow cytometric assessments of sperm nDNA damage.

Setting:
University-based research laboratory.

Patients:
Thirty-eight men attending the infertility clinic at the Centre of Reproductive Medicine and Andrology.

Interventions:
Induction of oxidative damage by Fenton’s reaction on semen samples.

Main Outcome Measure(s):
Raman profiles, FTIR spectra, and flow cytometric analysis of DNA fragmentation.

Result(s):
Raman and FTIR spectra contained distinctive differences between untreated and fragmented nDNA sperm which were indicative of oxidative attack. The changes in Raman profiles were similar to those previously seen and corresponded to the DNA backbone. The peak attributions were corroborated by the FTIR spectra. PCA of the entire Raman spectra distinguished samples with varying degrees of damage. After determination of a cut off value (0.63), estimation of the percentage of sperm with nDNA damage using the intensity ratio of Raman peaks (1050/1095 cm-118) correlated linearly to the flow cytometric assessment.

Conclusion(s):
Raman microspectroscopy still requires further validation but may potentially provide a means of assessing nDNA status of a living sperm.

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