Factors affecting activation and fertilization of human oocytes following intracytoplasmic injection.

Intracytoplasmic sperm injection (ICSI) has dramatically altered the treatment of severe male factor infertility, resulting in improved fertilization and pregnancy rates. The purpose of this study was to investigate oocyte activation and fertilization in aged human oocytes following ICSI. Non-viable spermatozoa were injected into 24 h old human oocytes in the presence and absence of calcium and were assessed for evidence of activation and fertilization 16-19 h after the injection procedure. Sham injections were also carried out to assess the effect of the injection procedure itself and the presence of calcium in the injection medium on oocyte activation. Non-viable spermatozoa injected in the presence of 1.78 mM calcium were capable of normally fertilizing aged human oocytes and the resulting zygotes underwent cleavage. None of the oocytes injected with non-viable spermatozoa in the absence of calcium were fertilized normally, although the rates of activation following all treatments were similar.

Pharmacological stimulation of sperm motility.

The treatment of male factor infertility is a rapidly developing field. The introduction of microsurgical fertilization techniques allows assisted conception units to treat couples who previously would not have benefited from in-vitro fertilization techniques. However, these techniques are only used for the minority of subfertile men in andrological practice. Many subfertile men are still treated pharmacologically or by sperm selection methods to enhance sperm fertilizing ability. Numerous pharmacological compounds have been described that enhance sperm motility and thus, potentially, sperm fertilizing capacity. This paper attempts to review these compounds and assess their role in treatment of the subfertile male.

Artificial induction of the acrosome reaction in human spermatozoa.

This study investigated the use of human follicular fluid and pentoxifylline as inducers of the human sperm acrosome reaction in vitro. Motile sperm suspensions were prepared using a discontinuous Percoll gradient, preincubated for 3 h, divided into aliquots and exposed to various concentrations of non-heat-inactivated follicular fluid for 1 and 24 h and pentoxifylline for 30 min. Detection of the acrosome reaction involved the combined use of a fluorescent vital stain, H33258, and fluorescein isothiocyanate-conjugated peanut agglutinin (FITC-PNA). A short (1 h) exposure to follicular fluid at concentrations of 50% or more, did not compromise sperm motility and significantly increased the proportion of spermatozoa having completed the acrosome reaction. Similarly, a 30 min exposure to pentoxifylline also significantly increased the proportion of spermatozoa having completed the acrosome reaction.

Cytogenetic and fluorescent in-situ hybridization chromosomal studies on in-vitro fertilized and intracytoplasmic sperm injected 'failed-fertilized' human oocytes.

This study was undertaken to establish baseline data on the chromosomal status of 'failed-fertilized' oocytes derived from in-vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) procedures. A cytogenetic analysis was undertaken on 162 IVF and 51 ICSI oocytes. In all, 82.1% (133/162) of the IVF and 78.4% (40/51) of the ICSI oocytes had metaphase II (MII) plates, of which 50.4% of the IVF and 47.5% of the ICSI oocytes were analysed further. Chromosomes of the G-group (21-22) were identified with the majority of the anomalies. No overall significant difference in the aneuploidy rate was found for the IVF (37.3%) of ICSI (31.6%) oocytes, or with maternal age. However, chromosome anomalies, e.g. diploidy, fragmented and broken chromatids, single sperm and oocyte chromatids, were found in oocytes from IVF patients aged > 36 years and in the ICSI oocytes throughout the maternal age range (31-38 years). The status of the polar body chromatin indicated that there was no overall significant difference in the maturation of the IVF and ICSI oocytes. Evidence of successful sperm delivery was found in 72.5% (37/51) of the ICSI failed-fertilized oocytes. In this group there was a significant increase in the incidence of premature chromosome condensation: 19.6% (10/51) contained sperm chromosomes, 7.8% (4/51) had swollen sperm heads, and the remaining 45.0% had condensed sperm heads. The presence of both sperm and MII oocyte chromosomes was found in 19.6% (10/51) of the ICSI and 8.6% (14/162) of the IVF failed-fertilized oocytes. Specific fluorescent in-situ hybridization DNA probes were used to re-analyse the chromosomes of karyotyped 'failed-fertilized' IVF oocytes and, for the first time, applied to the karyotyped chromosomes of failed-fertilized ICSI oocytes. The hybridization efficiency was 86-95% for the centromere probe and 100% for probes 21 and 18.