In this study we investigated whether morphology and chromatin anomalies in human spermatozoa can influence fertilization after intracytoplasmic sperm injection (ICSI). We examined unfertilized oocytes, using the flnorochrome Hoechst 33342, to determine whether a relationship exists between failure of fertilization and sperm chromatin quality. Sperm chromatin packaging quality was assessed using the chromomydn A 3 (CMA 3 ) fluorochrome, and the presence of DNA damage in spermatozoa, using in-situ nick translation. Normal males present sperm parameters with a normal morphology of >20%, CMA 3 fluorescence of <30% and exhibit endogenous nicks in <10% of their spermatozoa. When patients were separated according to these values no difference was observed hi their fertilization rates after ICSL When the unfertilized ICSI oocytes were examined, we found that patients with CMA 3 fluorescence of <30% and nicks in <10% of their spermatozoa had only 17.5 and 21.6% respectively of their unfertilized oocytes containing spermatozoa that remained condensed. In contrast, patients with higher CMA 3 and nick values had a significantly higher number, 412 and 48.9%, of their unfertilized oocytes containing condensed spermatozoa. Sperm morphology did not show any such pattern. The percentage of spermatozoa which had initiated decondensation in unfertilized oocytes was not influenced by morphology, CMA 3 fluorescence or nicks. In light of these results we postulate that poor chromatin packaging and/or damaged DNA may contribute to failure of sperm decondensation after ICSI and result in failure of fertilization.
A number of non-invasive methods have been proposed to evaluate embryo viability in human in-vitro fertilization programmes. In addition to biochemical analyses, a common method for the selection of embryos prior to transfer involves assessment of embryo quality and morphology. We propose a new method to evaluate embryo viability based on the timing of the first cell division. Fertilized embryos that had cleaved to the 2-cell stage 25 h post-insemination were designated as 'early cleavage' embryos while the others that had not yet reached the 2-cell stage were designated as 'no early cleavage'. In all cases the early cleavage embryos were transferred when available. Early cleavage was observed in 27 (18.9%) of the 143 cycles assessed. There were significantly (chi2 = 4.0; P = 0.04) more clinical pregnancies in the early cleavage group, 9/27 (33.3%), compared with the no early cleavage group, 17/116 (14.7%). No difference was found when comparing key parameters (age, stimulation protocol and semen characteristics) of couples belonging to both groups, pointing to an intrinsic property or factor(s) within the early cleaving embryos. We propose 'early cleavage' as a simple and effective non-invasive method for selection and evaluation of embryos prior to transfer.
In-vitro fertilization (IVF) embryos are selected for transfer on the basis of morphology and rate of development. However, when a number of embryos have similar characteristics, the selection of the best embryos is left to chance. Recently, we proposed a simple, novel method to overcome this problem, based on pre-selection of embryos cleaving early to the two-cell stage. In this study we have adopted the same method to choose embryos fertilized after intracytoplasmic sperm injection (ICSI). Fertilized embryos that had cleaved to the two-cell stage by 27 h post-injection were designated as 'early cleavage' embryos, while those that had not yet reached the two-cell stage were designated as 'no early cleavage'. In all cases, the early cleavage embryos were transferred when available. Early cleavage was observed in 54 (61.4%) of the 88 cycles assessed. There were significantly (P = 0.04) more clinical pregnancies in the early cleavage group, 14/54 (25.9%), compared with the no early cleavage group 2/34 (3.2%). No differences between the groups were found when comparing key parameters (age, stimulation protocol and semen characteristics) of the couples. Using the ICSI technique, we have shown that early cleavage to the two-cell stage is not influenced by the timing of fertilization, and is more likely due to intrinsic factors within the oocyte or embryo that promote embryo cleavage after fertilization.
We report on 332 infertile couples who underwent 1115 cycles of intrauterine insemination (IUI) with washed husband's semen. The indication for IUI was an abnormal post-coital test due to either a male or cervical infertility factor. The mean number of IUI cycles per patient was 3.4, the overall pregnancy rate 18.7%, and the pregnancy rate per cycle 5.6%. The cumulative pregnancy rate calculated by life table analysis showed that 16.0% of pregnancies occurred in the first three treatment cycles, while the cumulative pregnancy rate was 26.9% by the sixth cycle. The outcome of the therapy was adversely affected if the woman's age was >39 years and/or total motile sperm count per insemination was <1X1O*. No pregnancy occurred in women older than 44 years or in cases with a total motile sperm count before semen preparation of <1X1O*.
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