Study of mitochondrial membrane potential, reactive oxygen species, DNA fragmentation and cell viability by flow cytometry in human sperm

TLDR

Sperm cell death is a recognized contributor to male infertility. The study aimed to identify the most reliable method for assessing sperm quality during IVF preparation. Conventional semen analysis was compared with several cytofluorometric techniques that detect death-associated changes, evaluating neat semen from infertile patients and sperm prepared by PureSperm® gradient for mitochondrial membrane potential, reactive oxygen species, DNA fragmentation, and viability. Mitochondrial membrane potential correlated positively with standard semen parameters and fertilization rates, was the only detectable change in prepared sperm, and thus emerged as the most sensitive test for sperm quality and a promising predictor of IVF success.

Abstract

BACKGROUND: Sperm cell death appears to be a cause of male infertility. The objective of this study was to determine the most reliable method for the evaluation of sperm quality in semen samples during sperm preparation for IVF. METHODS: Conventional analysis of semen samples was compared with several cytofluorometric methods detecting death-associated changes. Neat semen from infertile patients and sperm prepared by PureSperm® gradient were studied by conventional microscopy and analysed for mitochondrial membrane potential (ΔΨΨm), generation of reactive oxygen species, DNA fragmentation and cell viability. RESULTS: In neat semen, a positive correlation was found between the percentage of ΔΨΨmhigh sperm cells and standard semen parameters (concentration/motility). Sperm cells depicting ΔΨΨmhigh and cells with low DNA fragmentation displayed high fertilization rate after IVF. The only changes that could be detected in prepared sperm were changes in ΔΨΨm, with ΔΨΨmhigh sperm positively correlated with forward motility and also with high fertilization rates after IVF. CONCLUSION: Analysis of mitochondrial membrane potential is the most sensitive test by which to determine sperm quality. These findings promise development of a test that may help to predict successful IVF.