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PubMed2002

[Role of the urologist in the management of infertile men at the time of sperm intracytoplasmic injection].

Although intracytoplasmic sperm injection (ICSI) revolutionized treatment and prognosis of male infertility, checkup (case history, clinical and paraclinical examinations) practiced by urologist in infertile man keeps all its place. Varicocele, congenital or acquired seminal duct obstruction, urogenital tract infection, or ejaculation disorder must be sought, because these affections remain accessible to treatment.

Humans↗
PubMed1999

Relationship between the sperm motility index assessed by the sperm quality analyzer and the outcome of intracytoplasmic sperm injection.

PURPOSE: Intracytoplasmic sperm injection (ICSI) has been validated as a useful treatment in severe male-factor patients who could not achieve fertilization and live births by conventional in vitro fertilization treatment. To examine the impact of male factors on ICSI outcome, clinical laboratory data were retrospectively analyzed. METHODS: One hundred two cycles of ICSI treatment indicated by severe male-factor infertility were entered into this study. Sperm parameters including sperm motility, sperm concentration, and sperm motility index assessed by the Sperm Quality Analyzer were evaluated. RESULTS: Five hundred seventy-six metaphase II oocytes retrieved were manipulated. The normal fertilization (2 PN) rate per oocyte was 64.9 +/- 26.0% (mean +/- SD). Of the 99 transfers, 31 clinical pregnancies were obtained, yielding an average pregnancy rate of 31.3% per transfer. The mean sperm motility, sperm concentration, and sperm motility index were 20.3 +/- 16.1% (range, 0 to 50%), 18.2 +/- 25.1 x 10(6)/ml (range, < 1 to 150 x 10(6)/ml), and 31.2 +/- 45.0 (range, 0 to 220), respectively. Sperm concentration did not have a significant impact on fertilization rate by ICSI. In four cases, ICSI was performed using totally immotile sperm and the fertilization rate was 43.5%, which was significantly lower than that of some of the other sperm motility groups, and no pregnancy could be achieved. In 14 cases in which the sperm motility index assessed by the Sperm Quality Analyzer was 0, the fertilization rate (50.0%) was significantly lower than in most of the other sperm motility index groups. CONCLUSIONS: These findings suggest that in severe male-factor cases with totally immotile sperm or a sperm motility index of 0, the selection of good-quality sperm should be verified before injection.

Female↗
PubMed2004

Use of Mammalian eggs for assessment of human sperm function: molecular and cellular analyses of fertilization by intracytoplasmic sperm injection.

PROBLEMS: Intracytoplasmic sperm injection (ICSI ) has been described as the 'cure' for male sterility because a single sperm can now be directly introduced into an egg with some chance of pregnancy. While ICSI has revolutionized the practice of assisted reproductive techniques (ART), there are few molecular and cellular studies about its safety and efficacy. Even by using ICSI, fertilization in humans succeeds only if the sperm effectively accomplishes a number of tasks including 'post-ICSI events' in fertilization. To assess the function of human sperm after ICSI, we used heterologous ICSI with human sperm into animal eggs. Egg activation, sperm decondensation and sperm centorosomal function were examined in sperm from fertile men and infertile patients. METHODS: Sperm from fertile men and infertile patients were injected into hamster, rabbit and bovine eggs by Piezo micromanipulator, and studied in decondensation of sperm nuclei, egg activation and microtubule organization. RESULTS: Decondensation human sperm head following ICSI into hamster eggs occurred initially form basal lesion, and apical portion of sperm nuclei which is surrounded by acrosome and perinuclear theca, still condensed in early pronuclear stage. Radial array of microtubules from sperm centrosome 'sperm aster' which is essential for pronuclear movement was observed in 30% rabbit eggs following ICSI with human sperm. By heterologous ICSI system with fertile human sperm and bovine eggs, 83.3% of eggs was activated and 60% eggs had sperm aster, indicating that bovine Piezo ICSI system is appropriate for assessing human sperm oocyte activation ability and human sperm centrosomal function. Oocyte activation and sperm centrosomal function were significantly low in sperm from men with globozoospermia and men with dysplasia of fibrous sheath. CONCLUSION: These assays indicate differences of the process of fertilization between in vitro fertilization and ICSI, and reflect the human sperm function especially for the 'post-ICSI events' in fertilization. More molecular and cellular analyses in fertilization by ICSI are needed for improvement of ART.

Acrosome↗
PubMed2006

Sperm retention site and its influence on cleavage rate and early development following intracytoplasmic sperm injection.

BACKGROUND: Intracytoplasmic sperm injection (ICSI) has risen to the forefront of reproductive technology. In the present study, the location of the sperm injection was noted, and a prospective study was conducted to evaluate the effect of the sperm retention site on cleavage rates and embryo quality after ICSI. METHODS: This study involved 336 ICSI patients (age 27-44; average 37.4) where 1545 oocytes were observed. An oocyte was divided into nine sites and the sperm retention site was observed microscopically after injection. The polar body was placed at either the twelve or six o'clock position. The injection pipette was introduced at the three o'clock position and oolemma rupture was ascertained by mild suction. The main outcome measures were the relationship of sperm remaining in position in the oocyte to fertilization rate and embryo quality. RESULTS: When the injection pipette was introduced at the three o'clock position, about 80% of the sperm remained in the center or left of center. The fertilization rate was significantly lower (p < 0.05) when the sperm remained near the site of introduction. Embryo quality was not significantly affected by the sperm retention site. CONCLUSIONS: About 12-14% of the spermatozoa remained near the introducing position, and in these cases the fertilization rate was low. However, once fertilization occurred, the sperm retention site had minimal impact on embryo quality. Injecting sperm near the spindle site may improve embryo quality.

Adult↗
PubMedDate not supplied

Spatiotemporal analysis of [Ca2+]i rises in mouse eggs after intracytoplasmic sperm injection (ICSI).

Intracytoplasmic sperm injection (ICSI) into mammalian eggs induces repetitive rises in intracellular Ca2+ concentration ([Ca2+]i) which are the pivotal signal in fertilization. Spatiotemporal aspects of [Ca2+]i rises following ICSI into the periphery of mouse eggs were investigated with high-speed confocal microscopy. The first Ca2+ response was generated 25-30 min after ICSI, when [Ca2+]i increased slowly and reached a certain level. The [Ca2+]i rise occurred synchronously over the ooplasm, attained the peak in 40-70 s, and lasted for 5-7 min. Succeeding Ca2+ responses occurred at intervals of 20-30 min, associated with the faster rate of [Ca2+]i rise and the shorter duration as Ca2+ oscillations progressed. The [Ca2+]i rises took the form of a wave that started from an arbitrary cortical region, but not from the vicinity of the injected sperm head. The Ca2+ wave became more pronounced and propagated across the egg faster in the later Ca2+ responses. An artifactual [Ca2+]i rise was inevitably produced during the ICSI procedure. The larger artifact affected the subsequent first Ca2+ response, resulting in the faster [Ca2+]i rise (time to peak, 10-20 s), slight spatial heterogeneity of [Ca2+]i rise in the ooplasm (but not a wave) and the shorter duration (3-4 min). The artifact slightly affected the amplitude of the second Ca2+ response, but little affected the later Ca2+ responses. It is suggested that the factor(s) that leaked out of the injected spermatozoon diffuses to a wide area and sensitizes Ca2+ channels of the endoplasmic reticulum to induce Ca2+ release synchronously over the ooplasm. The enhanced sensitization leads to propagating Ca2+ release initiated from the cortex that is more sensitive to the sperm factor.

Animals↗
PubMed1998

Strong preference for non-invasive prenatal diagnosis in women pregnant through intracytoplasmic sperm injection (ICSI)

Intracytoplasmic sperm injection (ICSI) is a new and most powerful technique to treat severe forms of human infertility. While the follow-up studies have not shown an increased malformation risk so far, the genetic implications of ICSI are still not fully understood. For this reason, many institutions routinely recommend or even enforce invasive prenatal tests after successful intracytoplasmic sperm injection. We have counselled 107 women pregnant through ICSI about prenatal diagnosis. Sixty-five had already received genetic counselling prior to the treatment (group 1), while 42 had not attended our clinic before (group 2). They were free to choose between invasive and non-invasive diagnosis or no prenatal tests at all. Fifty-four per cent of these patients had an indication for prenatal karyotyping or other invasive procedures independent of ICSI. Only 17 per cent of the total cohort made use of amniocentesis or fetal blood sampling, 82 per cent opted for non-invasive tests (ultrasound, serum screening), and one couple did not wish any prenatal studies. The preference for non-invasive procedures was stronger in group 1 (94 per cent) than in group 2 (65 per cent). We suggest that if patients pregnant through ICSI have the option to choose freely between invasive and non-invasive prenatal tests, they strongly favour the latter.

Adult↗
PubMed1997

Is paternal mitochondrial DNA transferred to the offspring following intracytoplasmic sperm injection?

During intracytoplasmic sperm injection (ICSI) the whole sperm, including head, midpiece and tail, is injected into the middle area of the oocyte. To find out what happens to the sperm mitochondria after ICSI, we checked the first six children born after ICSI treatment for occurrence of paternal mitochondrial DNA (mtDNA). The difference between maternal and paternal mtDNA in the investigated couples in our study was confined to single-base pair substitutions and we had to rely on restriction enzyme cleavage to differentiate between the mitochondrial genomes of the parents. With this kind of assay we were able to reach a sensitivity of about 0.2% for the paternal mtDNA. However, as uneven partition between tissues of heteroplasmic mtDNA is expected to occur, it would not be unlikely that an enrichment to 0.2% would occur in a given tissue if paternal mtDNA was transmitted by the ICSI procedure. We did not detect this level in the blood in any of the six children.

Adult↗
PubMed1999

Unilateral or bilateral testicular biopsy in the era of intracytoplasmic sperm injection.

PURPOSE: Intracytoplasmic sperm injection has significantly improved the treatment of male infertility. Since only single vital spermatozoa are required for successful fertilization, the value of unilateral or bilateral diagnostic testicular biopsies in patients with azoospermia is controversial. We evaluated differences in bilateral testicular biopsies in azoospermic patients with regard to testicular histology and focal spermatogenesis. MATERIALS AND METHODS: Histopathological results of 100 testicular biopsies from 50 patients (mean age 33.3 years) were reviewed. In all cases azoospermia was the indication for diagnostic testicular biopsy. Intra-individual differences of bilateral testicular biopsies were retrospectively reviewed by determining the latest stage of spermatogenesis. RESULTS: After bilateral biopsy a difference in testicular histology was found in 28% and identical histopathology was noted in 70% of patients. An unsuspected burned out seminoma with maturation arrest in the contralateral testis was seen in 2% of cases. Testicular symmetry determined by a Prader orchidometer was noted in 54.8% of patients whereas 45.2% had asymmetrical testis. The frequency of divergent histopathologies in relation to testicular symmetry was 21.7 and 26.3%, respectively. Spermatozoa were found in 42% of right and 44% of left testes (p >0.05), and spermatids as the latest stage of differentiation were detected in 14 and 16%, respectively (p >0.05). Differentiation of testicular histologies according to the side of biopsy revealed spermatozoa and/or spermatids in 56% of right and 58% of left testes (p >0.05). Bilateral biopsies increased the detection of focal spermatogenesis to 68%. If only unilateral diagnostic testicular biopsies had been performed, in 20% of patients focal spermatogenesis in the contralateral testis would have been missed. CONCLUSIONS: Bilateral testicular biopsies are superior to unilateral biopsies in the evaluation of patients with azoospermia. A 28% intra-individual difference in testicular pathology was seen after bilateral biopsies, and in 20% of patients focal spermatogenesis would have been missed after unilateral biopsy only. Due to the prognostic relevance of testicular biopsies for successful sperm retrieval before assisted reproduction, bilateral diagnostic testicular biopsies are recommended in the evaluation of patients with azoospermia.

Adult↗
PubMed1998

Medical and developmental outcome at 1 year for children conceived by intracytoplasmic sperm injection.

BACKGROUND: Intracytoplasmic sperm injection (ICSI) was introduced as a new form of in-vitro fertilisation (IVF) in 1993 and is now accepted as the treatment of choice for severe male infertility in many centres around the world. However, there is little information about the long-term outcome of children conceived by ICSI. We aimed to find out the medical and developmental outcome of children conceived by ICSI at age 1 year. METHODS: In this prospective study, we compared the medical and developmental outcome at 1 year of 89 children conceived by ICSI with 84 children conceived by routine IVF, and with 80 children conceived naturally. Formal developmental assessment was done with Bayley Scales of Infant Development (2nd edition) from which a mental development index (MDI) was derived. FINDINGS: There was no significant difference in the incidence of major congenital malformations or major health problems in the first year of life. However, the mean Bayley MDI was significantly lower for the children conceived by ICSI than for the children conceived by routine IVF or naturally (95.9 [SD 10.7], 101.8 [8.5], and 102.5 [7.6], respectively, p < 0.0001). 15 (17%) of 89 children conceived by ICSI experienced mildly or significantly delayed development (MDI < 85) at 1 year compared with two (2%) of the 84 children conceived by IVF and one (1%) of the 80 children conceived by natural conception (p < 0.0001). INTERPRETATION: Although most children conceived by ICSI are healthy and develop normally, there is an increased risk of mild delays in development at 1 year when compared with children conceived by routine IVF or conceived naturally. These findings support the need for ongoing developmental follow-up of children conceived by ICSI to see whether they are at increased risk of intellectual impairment or learning difficulties at school age.

Adult↗
PubMed1996

Biochemical study of individual zonae from human oocytes that failed to undergo fertilization in intracytoplasmic sperm injection.

Successful intracytoplasmic sperm injection (ICSI) is dependent upon the competence of the oocyte to respond to the injection of a spermatozoon in the presence of calcium. This study determined if oocytes that failed to become fertilized (absence of any pronuclei) failed to undergo cytoplasmic activation, as ascertained by an electrophoretic shift in the zona pellucida (zona) protein, huZP2. Of 48 zonae individually analysed, 58% did not have a detectable huZP2 shift. Three patterns were observed. In seven patients, none of the unfertilized oocyte huZP2 shifted (16 out of 48); in two, all zonae exhibited a huZP2 shift (five out of 48); in eight, there was a mixture of shifted and unshifted (27 out of 48) in each case. There was no clear relationship between the presence of the huZP2 shift and maternal age, pregnancy outcome, male-factor infertility, or fertilization rate. However, in six couples diagnosed as having no detectable male-factor infertility, 73% (11 out of 15) of the zonae had unshifted huZP2, suggesting that an oocyte defect is involved in some cases of failed fertilization after ICSI. One likely cause for the absence of the huZP2 shift is a failure of cytoplasmic activation. Since oocytes were injected at metaphase II, we hypothesize that some oocytes that failed to fertilize have completed nuclear (meiotic) without cytoplasmic maturation.

Adult↗
PubMed1997

[Value of a genetic evaluation before an intracytoplasmic sperm injection procedure].

Intracytoplasmic sperm injection (ICSI) still needs to be validated as far as safety is concerned. Results concerning foetal karyotypes currently seem to show an increase in sexual chromosomal abnormalities. In view of the heterogeneity of publications concerning genetic tests, it seems necessary to define a protocol with a limited time span, which will allow the extensive collection of data. The analysis of the results will secondarily allow a common procedure to be established. We propose a protocol limited in time, aiming to evaluate the chromosomal status at birth by a systematic karyotype of umbilical cord blood.

Adult↗
PubMed1998

[The ICSI (Intracytoplasmic Sperm Injection)].

ICSI (Intracytoplasmic Sperm Injection) is the latest known assisted reproduction technique (ART) and it already appears to be mature. In fact the analysis of the results presented by the researchers over the years has shown that the most specific indication for this ART is the sterility of the couple with serious male pathology, up to ejaculatory azoospermia where it is possible to perform MESA, PESA or TESE. Any kind of sterility could be solved with ICSI whose only limit presently known is the high technology and therefore high costs involved. The percentage of oocytes that undergo ICSI without being damaged varies from 87% to 94% and the percentage of fertilization varies from 33% to 71%. The transfer rate is 59-100%. The rate of pregnancies per couple ranges from 12% to 40% of the couples, from 11% to 41% for the transfers. Since abortions are similar to the values of the normal population (10-15%), ICSI is actually the assisted fertilization technique with the highest incidence of pregnancies and "take home babies". The percentage incidence of the two sexes, of the malformations and the typologies of malformations corresponds to those observed in the population with spontaneous pregnancies. Since there is no natural selection of the gametes in ICSI, one may be sure that when spermatozoa with any kind of pathology are injected, the pregnancy does not take place at all.

Cytoplasm↗
PubMed1998

Percutaneous testicular sperm aspiration: a convenient and effective office procedure to retrieve sperm for in vitro fertilization with intracytoplasmic sperm injection.

PURPOSE: After intracytoplasmic sperm injection was established to facilitate in vitro fertilization in men with the most severe semen abnormalities, the use of testicular sperm to achieve conception became feasible. We investigated the use of a method of percutaneous needle aspiration previously used for diagnostic purposes to obtain testicular sperm for intracytoplasmic sperm injection. MATERIALS AND METHODS: A method of percutaneous aspiration of sperm was developed to facilitate intracytoplasmic sperm injection. A total of 69 testicular aspirations were performed for diagnostic purposes and 179 to obtain sperm on the day of egg retrieval for couples undergoing in vitro fertilization with intracytoplasmic sperm injection. The procedures were performed in an outpatient facility. Most patients received intravenous sedation and a few received only local anesthesia. RESULTS: Sperm adequate for intracytoplasmic sperm injection were obtained in all men with obstructive azoospermia, including those with significant testicular atrophy and those with anejaculation or necrospermia. Adequate numbers of sperm for intracytoplasmic sperm injection were retrieved less reliably in men with nonobstructive azoospermia. The number of sperm correlated positively with testicular size. Morbidity and discomfort were nonexistent. Sperm were obtained from 43 of 69 men undergoing diagnostic and 170 of 179 men undergoing therapeutic aspiration. Sperm motility ranged from 0 to 20% and viability from 55 to 85%. CONCLUSIONS: Percutaneous testicular sperm aspiration is a cost-effective method to retrieve sperm for intracytoplasmic sperm injection in select men with obstructive azoospermia, anejaculation and necrospermia, and some with nonobstructive azoospermia.

Equipment Design↗
PubMed2000

Intracytoplasmic sperm injection in bovine: effects of oocyte activation, sperm pretreatment and injection technique.

Intracytoplasmic sperm injection (ICSI) is a very important technique for treating male subfertility and for basic research. The efficiency of ICSI in bovine is very limited because of the necessity for additional oocyte activation before or after the ICSI procedure. In this study, we compared the effects of seven different protocols on activation and fertilization rates of bovine oocytes after ICSI and on their subsequent development under in vitro conditions. The protocols include 1) different chemical activation of oocytes, 2) pretreated or nonpretreated sperm, and 3) conventional or Piezo-driven injection techniques. In all three groups, ICSI, sham-injected, and noninjected, the highest activation rates were obtained after treatment of oocytes with ionomycin followed by 6-dimethylaminopurine (6-DMAP). Using this treatment for oocyte activation, 59% of oocytes were activated and 31% of oocytes were fertilized using dithiothreitol (DTT) pretreated spermatozoa and Piezo-driven injection. Using the protocols with the same oocyte activation or activation with calcium ionophore (Ca-I) and cycloheximide (CHX), nonpretreated sperm, and conventional injection technique, early cleavage rate (79.6% and 77.6%, respectively) were significantly (P <0.01) higher when compared with all other protocols. The latter protocol resulted in 8% blastocyst and 90% of the obtained blastocysts were found to be diploid. Our results demonstrate that activation of oocytes, sperm treatment, and injection technique separately or together could improve the success of bovine ICSI.

Adenine↗
PubMed1999

Clinical application of intracytoplasmic sperm injection using in vitro cultured testicular spermatozoa obtained the day before egg retrieval.

OBJECTIVE: To study the effect of in vitro culture on the quality of human testicular sperm and the efficiency of intracytoplasmic sperm injection with in vitro cultured testicular sperm. DESIGN: Clinical study. SETTING: A private IVF center. PATIENT(S): Twenty consecutively seen IVF patients undergoing testicular biopsies for ICSI. INTERVENTION(S): The testicular specimens were cultured in vitro for 24 hours and the isolated spermatozoa were microinjected. MAIN OUTCOME MEASURE(S): Preincubation and postculture sperm motility, and fertilization, implantation, and pregnancy rates after intracytoplasmic sperm injection. RESULT(S): Motility increased from initial nonmotile or twitching sperm to free motile sperm in 18 of 20 cases. The injection of in vitro cultured testicular sperm resulted in a fertilization rate of 58%, an implantation rate of 20%, and a pregnancy rate of 45%. CONCLUSION(S): A testicular biopsy procedure can be performed the day before egg retrieval. Despite the low initial sperm quality, a high percentage of the prepared testicular sperm showed increased motility after 24 hours of culture. The injection of in vitro cultured testicular sperm into matured oocytes resulted in fertilization, implantation, and pregnancy rates comparable to those obtained with ejaculated sperm.

Adult↗
PubMed2005

Influence of motility and vitality in intracytoplasmic sperm injection with ejaculated and testicular sperm.

The vitality of spermatozoa used for intracytoplasmic sperm injection (ICSI) is a crucial factor for fertilization, establishment and outcome of a pregnancy in assisted reproductive technique cycles. The sperm origin may also be a limiting factor, although little is known about this issue. It is known that the motility of injected spermatozoa and their origin from ejaculate or testicular biopsies are important predictors in terms of fertilization, pregnancy and birth rates. Oocytes of patients in 2593 cycles were retrieved in our in vitro fertilization programme and inseminated via ICSI. We used motile (group 1, n = 2317) or immotile ejaculated spermatozoa (group 2, n = 79), motile sperm retrieved from testicular biopsies (group 3, n = 62) and immotile spermatozoa from testicular biopsies (group 4, n = 135). Female age and number of oocytes retrieved did not differ significantly among the groups. The fertilization rates were as follows: 67.1% in group 1, 49.8% in group 2, 68.3% in group 3 and 47.8% in group 4. The pregnancy rates in cases where three embryos had been transferred amounted to 35.7% in group 1, 17.3% in group 2, 38.3% in group 3 and 20.5% in group 4. The embryo quality showed no differences between groups 1 and 3 (14.5), and between groups 2 (11.8) and 4 (10.8). The abortion rate was similar in groups 1-3, but increased in group 4 (26.6%, 27.3%, 31.6% and 55.5%). Irrespective of their origin, the fertilization potential of injected spermatozoa was found to be influenced by motility. The resulting pregnancy and birth rates, i.e. the potential of the resulting embryos to implant and to achieve viable pregnancies, seem to be additionally dependent on the sperm origin. This was well shown by declining rates when spermatozoa in a relatively early stage of maturity had been used. We see increasing evidence that the degree of sperm maturity has an important impact on the outcome of ICSI. In obstructive azoospermia, spermatozoa retrieved from the epididymis should be used rather than testicular biopsy spermatozoa, or testicular sperm should be preincubated in culture medium before ICSI.

Adult↗
PubMed2005

Active demethylation of individual genes in intracytoplasmic sperm injection rabbit embryos.

Intracytoplasmic sperm injection (ICSI), as an assisted reproduction technique, has been widely used in animal and human. However, its possible effect on epigenetic changes has not been well studied. To investigate whether ICSI can induce aberrant DNA methylation changes in rabbit preimplantation embryos, we examined the methylation status of the SP-A promoter region and the satellite sequence Rsat IIE by bisulfite-sequencing technology. The SP-A promoter region was extensively demethylated before the first round of DNA replication commences, and the unmethylated status was maintained until morula when dynamic remethylation occurred. A similar but more moderate demethylation process was observed in satellite sequence Rsat IIE. These results are in contrast with the previous reports of no active demethylation in normal rabbit embryos, suggesting that the active demethylation we observed may be induced by ICSI.

Animals↗