A double-blinded comparison of in situ TUNEL and aniline blue versus flow cytometry acridine orange for the determination of sperm DNA fragmentation and nucleus decondensation state index.

The impact of sperm DNA fragmentation on assisted reproductive technology (ART) successes, in terms of outcome, is now established. High levels of DNA strand breaks severely affect the probability of pregnancy. The importance of sperm nucleus condensation in early embryogenesis and, subsequently, on the quality of the conceptus is now emerging. In this article we have compared in situ analyses with terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labelling (TUNEL) (for DNA fragmentation) with aniline blue (AB) (for nucleus decondensation), versus flow cytometry (FC) after acridine orange staining, in a double-blinded analysis. In our hands, TUNEL and acridine orange give perfectly comparable results. For decondensation the results are also comparable, but the double-stranded green fluorescence obtained with acridine orange seems to slightly underestimate the decondensation status obtained with AB.

Correlation between DNA damage and sperm parameters: a prospective study of 1,633 patients.

OBJECTIVE: To investigate DNA fragmentation by using terminal deoxyribonucleotidyl transferase-mediated dUTP nick-end labeling in relation to World Health Organization parameters and computer-aided sperm analysis (CASA) in sperm to determine the possibility of obtaining a correlation among CASA parameters, sperm morphology, and DNA fragmentation. DESIGN: Sperm analysis according to World Health Organization parameters, terminal deoxyribonucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) for sperm DNA fragmentation, and CASA for sperm movement. Prospective study. SETTING: All the patients were under clinical management, consulting for hypofertility at a fertility center in France. PATIENT(S): One thousand six hundred thirty-three men who were referred for infertility investigation, including a complete sperm analysis. INTERVENTION(S): Sperm analysis and DNA damage testing. MAIN OUTCOME MEASURE(S): Sperm morphology, DNA fragmentation, and movement characteristics. RESULT(S): One third of the patients had a TUNEL rate of >30%. Analysis of the 21 semen parameters tested revealed that 7 of them were significantly correlated with the TUNEL results. CONCLUSION(S): World Health Organization sperm parameters and DNA damage are complementary, rather than strongly linked. This should be considered to more fully understand the paternal contribution in assisted reproductive technologies failures.

The Murine Prepuberal Oviduct Supports Early Embryo Development In Vitro: (mouse embryo coculture/2 cell-block/immature oviduct/protein secretion/early embryo signal).

Swiss albino mice were used to evaluate the ability of explanted murine oviducts to support development through the block stage. One cell eggs develop as well when cultured 50 hours in oviducts explanted from pregnant mice or in oviducts from immature mice: The blastocyst formation occurs at a similar rate in both cases. The viability of the blastocysts was high (8/9) when transferred to pseudopregnant (C 57 Black) recipient mice. Only a few difference was observed in the polypeptide pattern of immature and pregnant explanted oviduct. In immature oviduct, the polypeptide secretory profile is not modified by the presence of fertilized embryos transferred into it, and so nor is it directly egg dependent. It is concluded that oviduct's ability to sustain normal early embryonic developments is not dependent upon sexual maturity.