Intracytoplasmic Sperm Injection Group Research Articles

Standard in vitro fertilization or intracytoplasmic sperm injection in advanced female age – what may be expected?

This study was conducted to evaluate the current results of standard in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) in the elderly (≥ 40 years of age) female population. Oocyte recovery, fertilization, embryo transfer, pregnancy and cumulative pregnancy rates were assessed. The results were analyzed for: the entire elderly population; the standard IVF group (group 1); all those in the ICSI group (group 2); and ICSI for severe male-factor category (group 3). A total of 330 IVF and 158 ICSI treatment cycles were carried out in 249 women. Forty-five (9.2%) clinical pregnancies were achieved. This rate was not statistically different from those achieved for groups 1, 2 and 3 (9.1%, 9.5%) and 6.8%, respectively). The cumulative pregnancy rate for a total of five cycles was 19.2% and 26.4% for groups 1 and 2, respectively. For those who started their treatments at ≥ 40 years, the cumulative pregnancy rate for three cycles was 26.5% and 36.5% in groups 1 and 2, respectively. These results clearly demonstrate that female age is a major success determinant, with similar influence on both standard IVF and ICSI therapy modalities.

Does female age affect embryo morphology?

Deteriorating oocyte quality is commonly believed to be the primary determinant of the decreased implantation potential in older women. We assessed the influence of age on embryo morphology in standard in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) modalities. All 6350 consecutive embryos (2990 IVF, 3360 ICSI) obtained in our Assisted Reproductive Technology Unit from January 1996 through June 1991 were included. High quality embryos were defined as those with equal-sized blastomeres and < 10% fragmentations and a cleavage rate of four cells on day 2 or eight cells on day 3 transfers. The results were analyzed for the standard IVF group, the ICSI group, and the ICSI subgroup with severe male factor infertility (≤ 1 × 106 total motile spermatozoa in the ejaculate). For standard IVF, a positive association was observed between female age and increased proportion of good quality embryos. No such association was detected for the ICSI cycles (whole group or subgroup). We conclude that in standard IVF, embryo quality, as reflected by embryo morphology, does not deteriorate with increased maternal age.

Incidence of Sex Chromosome Abnormalities in Spermatozoa From Patients Entering an IVF or ICSI Protocol

Objective. The objective of this study was the determination of sex chromosome aneuploidy frequency in spermatozoa from patients included in an in vitro fertilization (IVF) or intra cytoplasmic sperm injection (ICSI) protocol. Methods. Spermatozoa from nineteen patients, including patients with normal seminal parameters according to World Health Organization (WHO) criteria and patients exhibiting abnormal seminal parameters, were analyzed by dual color fluorescence in situ hybridization (FISH) for aneuploidies of the X and Y chromosomes. Our technique, using only probes for sex chromosomes and not for autosomes, does not discriminate between hyperhaploid and diploid sperm nuclei. The results were analyzed in two different ways: in relation to the semen status, denoted normal or abnormal and with regard to the ability of the sperm to fertilize oocytes when IVF or ICSI was performed. Results. Abnormal semen showed a significant increase in the overall rate of sperm nuclei with XY, XX and YY sex chromosome complements, 1.59% compared to normal semen, 0.78% (p<0.02). Semen shown to be able to fertilize oocytes only by ICSI showed a higher incidence of XY-bearing spermatozoa, 1.26%, compared to semen able to fertilize oocytes by conventional IVF, 0.37% (p<0.001). The incidence of XX-or YY-bearing sperm nuclei was also significantly elevated in the ICSI group (0.25% XX, 0.50% YY) (p<0.02) as compared to the IVF group (0.06% XX, 0.16% YY). Conclusions. We concluded that infertile men requiring ICSI treatment showed a higher incidence of sex chromosome aneuploidy, due to meiosis I and II nondisjunction, in their spermatozoa as compared to men requiring IVF for reasons of predominantly female infertility.

Development of embryos produced by intracytoplasmic sperm injection of cat oocytes.

Development of cat oocytes following intracytoplasmic sperm injection (ICSI) and in vitro fertilization (IVF) was compared in two experiments. Domestic cat donors (used as a model for wild felids) were treated with 150 IU equine chorionic gonadotrophin (eCG) on treatment day 1 or a total of 10-15 IU of follicle-stimulating hormone (FSH) over four days, followed by 100 IU human chorionic gonadotrophin (hCG) on day 5 and follicular aspiration 24-26 h later. A jaguarundi (Herpailurus yaguarondi) female was stimulated twice with FSH (20 IU) or eCG (300 IU) and hCG (250 or 300 IU) before oocyte recovery. After storage at 4 degrees C, domestic cat semen was washed and processed. For ICSI, denuded oocytes were each injected with an immobilised spermatozoon. IVF oocytes were co-incubated with 5 x 10(4) motile spermatozoa/0.5 ml for 4-6 h. Noncleaving oocytes were fixed and stained 24-28 h after injection or insemination. Presumptive zygotes were cultured before transfer on day 5 (experiment I only) or evaluation on day 7 (experiments I and II). In experiment I, fertilization frequency was 67.9% (72/106) and 58.1% (122/210) for IVF and ICSI oocytes, respectively (P > 0.05). Most noncleaving ICSI oocytes (71/88, 80.7%) at 24 h were at metaphase II, of which half (35/71, 49.3%) had an activated spermatozoon (n=4) or premature chromatin condensation (PCC, n=31) of the sperm head. All 69 day 7 IVF embryos developed to morulae (> 16-cells, 46.7%) or blastocysts (53.3%), and 59/63 (93.7%) ICSI embryos reached the morula (50.8%) or blastocyst (42.9%, P > 0.05) stage. Mean cell number in IVF and ICSI embryos was 136 and 116 (P > 0.05); morulae had 77 and 46 (P < 0.05) and blastocysts had 187 and 209 (P > 0.05) cells, respectively. After transfer of 10 or 11 day 5 ICSI morulae to each of four recipients, a total of three kittens were born to two dams at 66 or 67 days. Of 18 fair-to-good quality oocytes recovered from a jaguarundi on two occasions, 10 (55.6%) embryos were produced by ICSI with fresh (n=5) or frozen (n=5) conspecific spermatozoa, but no jaguarundi kittens were born after transfer of these embryos to domestic cat recipients. In experiment II, cleavage frequency following IVF (15/17, 88.2%) and ICSI (31/38, 81.6%) was higher (P < 0.05) than following sham ICSI (13/35, 37.1%). Mean cell number (27 cells) and blastocyst development (0%) on day 7 was lower (P < 0.05) in the sham ICSI group than in the ICSI group (45 cells, 15.6% blastocysts) which, in turn, was lower (P < 0.05) than the IVF group (94 cells, 46.7% blastocysts). We have demonstrated that ICSI can be applied successfully in domestic felids and suggest that the technique will effectively augment other biotechniques being developed for enhancing reproduction in endangered felids.

Comparison of cryopreservation outcome following intracytoplasmic sperm injection and conventional in vitro fertilization.

Our purpose was to compare the success rate of transferring frozen-thawed embryos generated from either intracytoplasmic sperm injection (ICSI) or conventional in vitro fertilization (IVF). A retrospective review of all frozen-thawed embryo transfer (ET) cycles between January 1995 and April 1997 was performed. There were 83 and 204 transfer cycles of frozen-thawed multicellular embryos generated from conventional IVF (group A) and ICSI (group B), respectively. The survival rate of frozen-thawed embryos and the outcome following ET in both groups were assessed. The groups did not differ in age (31.7 +/- 4.6 and 30.6 +/- 6.0; mean +/- SD) or number of embryos transferred (3.5 +/- 1.1 and 3.8 +/- 1.3 for groups A and B, respectively). An acceptable pregnancy rate per ET was achieved in both groups, but the rate was significantly higher (P = 0.04) for group A than group B, 32.5 and 20%, respectively. Group A included frozen embryos of a higher quality than those of group B, but the proportion of embryos surviving after thawing was significantly higher for group B than group A (92.5 and 85.6%, respectively; P = 0.0004). The abortion rate did not differ between the two groups: 22 and 26.8% for groups A and B, respectively. Although an overall high pregnancy rate was achieved following frozen-thawed ET, it was lower for cycles in which embryos had been generated from ICSI. This difference may be attributed to a lower prefreezing embryo quality in the ICSI group. Embryos originating from ICSI were not vulnerable to cryopreservation and, when implanted, resulted in a comparable abortion rate to thawed embryos of conventional IVF.

Chromosome Abnormalities in Embryos Obtained After Conventional In Vitro Fertilization and Intracytoplasmic Sperm Injection

Objective: To compare the rate of numerical chromosome abnormalities in embryos derived from bipronucleated zygotes produced by intracytoplasmic sperm injection (ICSI) and conventional IVF. Design: Embryos were classified by maternal age and morphological and developmental characteristics to avoid bias when comparing chromosome abnormalities in ICSI and IVF embryos. Setting: The Institute for Reproductive Medicine and Science of Saint Barnabas Medical Center, West Orange, New Jersey. Patient(s): Seventy-nine couples undergoing IVF and 53 couples undergoing ICSI. Intervention(s): Embryos donated for research were fully biopsied, and their cells were analyzed by fluorescence in situ hybridization with specific probes for chromosomes X, Y, 13, 18, and 21 and some with also a probe for chromosome 16. Main Outcome Measure(s): Embryo chromosome abnormalities. Result(s): A total of 245 embryos obtained through conventional IVF and 136 embryos obtained through ICSI were analyzed. There were no statistical differences between the rates of numerical chromosomal abnormalities detected in the IVF (61%) and ICSI (52%) embryos analyzed. Regarding gonosomal aneuploidy, the same rate was found in both ICSI (1%) and IVF groups (2%). Conclusion(s): If the parents are chromosomally normal, the results indicate that, at the embryo level and before any embryo selection has occurred in utero, ICSI does not produce more numerical chromosomal abnormalities than conventional IVF.

Incidence of sex chromosome abnormalities in spermatozoa from patients entering an IVF or ICSI protocol

The objective of this study was the determination of sex chromosome aneuploidy frequency in spermatozoa from patients included in an in vitro fertilization (IVF) or intra cytoplasmic sperm injection (ICSI) protocol. Spermatozoa from nineteen patients, including patients with normal seminal parameters according to World Health Organization (WHO) criteria and patients exhibiting abnormal seminal parameters, were analyzed by dual color fluorescence in situ hybridization (FISH) for aneuploidies of the X and Y chromosomes. Our technique, using only probes for sex chromosomes and not for autosomes, does not discriminate between hyperhaploid and diploid sperm nuclei. The results were analyzed in two different ways: in relation to the semen status, denoted normal or abnormal and with regard to the ability of the sperm to fertilize oocytes when IVF or ICSI was performed. Abnormal semen showed a significant increase in the overall rate of sperm nuclei with XY, XX and YY sex chromosome complements, 1.59% compared to normal semen, 0.78% (p<0.02). Semen shown to be able to fertilize oocytes only by ICSI showed a higher incidence of XY-bearing spermatozoa, 1.26%, compared to semen able to fertilize oocytes by conventional IVF, 0.37% (p<0.001). The incidence of XX-or YY-bearing sperm nuclei was also significantly elevated in the ICSI group (0.25% XX, 0.50% YY) (p<0.02) as compared to the IVF group (0.06% XX, 0.16% YY). We concluded that infertile men requiring ICSI treatment showed a higher incidence of sex chromosome aneuploidy, due to meiosis I and II nondisjunction, in their spermatozoa as compared to men requiring IVF for reasons of predominantly female infertility.

Cytogenetic abnormalities of unfertilized oocytes generated from in-vitro fertilization and intracytoplasmic sperm injection: a double-blind study.

In the present study we have assessed the cytogenetic abnormalities of unfertilized oocytes from in-vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) programmes during a one year period (July 1995 to July 1996) with the cytogenetic analysis being carried out in a double-blind manner. A total of 88 unfertilized ICSI and 85 unfertilized IVF oocytes were used for the study and of these 51 and 62 oocytes, in each respective group, were suitable for analysis. The haploidy, diploidy and aneuploidy rates between ICSI (62.7, 7.8 and 5.9%) and IVF (61.3, 9.7 and 14.5%) groups were similar. A significant inter-patient variation in the incidence of hypohaploidy was observed within the IVF group. Chromosomal fragmentation or breakage was observed at a similar rate in both groups of unfertilized oocytes (23.5 and 14.5% for ICSI and IVF respectively). A significantly higher proportion of ICSI oocytes contained sperm nuclei (27/51, 52.9%) than did IVF oocytes (20/62, 32.3%, P < 0.01). The distribution and state of sperm head chromatin in relation to oocyte chromosomal complement was studied in both groups. ICSI oocytes contained decondensed or swollen sperm nuclei in association with haploid oocyte chromosomes (12/27, 44.4%) or condensed sperm heads in oocytes showing no chromosomal complements (7/27, 25.9%). In IVF oocytes sperm heads were either arrested in the condensed state (5/20, 25%), metaphase stage (3/20, 15%) or had undergone premature chromosome condensation (PCC; 6/20, 30%) in association with haploid oocyte chromosomes. The incidence of PCC was similar in the two groups. A marked variation in the incidence of total chromosomal abnormality was observed between patients within both ICSI (0-75%) and IVF (0-71%) groups indicating a possible similarity in oocyte quality between the majority of male factor and tubal infertility patients. The type of sperm used in the two fertilization procedures showed an increased incidence of chromosomal breakage with ICSI-MESA (microepididymal sperm aspiration) spermatozoa (4/6, 67%) compared to the ICSI-ejaculated (6/35, 17.1%; P < 0.05), ICSI-testicular biopsy (2/10, 20%) and IVF-normospermic (9/62, 14.5%; P < 0.01) spermatozoa. Chromosomal fragmentation may be associated with the degree of difficulty experienced at sperm injection, especially with sperm retrieved from the reproductive tract. Thus chromosomal fragmentation in ICSI may need further investigation using a larger sample size in order to assess the possible causative factors.

The relationship between sperm morphology and rates of fertilization, pregnancy and spontaneous abortion in an in-vitro fertilization/intracytoplasmic sperm injection programme.

The morphological normality of a spermatozoon is considered to be an important factor in relation to its ability to fertilize an oocyte. We examined the influence of morphology (strict criteria) on the rates of fertilization, pregnancy and spontaneous abortion obtained following conventional in-vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) in our clinical programme. We found our fertilization cut-off values for conventional IVF to be slightly different from those of the Kruger group (10 and 5%, compared to 14 and 5%). We also found the pregnancy rate per transfer to be as good or better in the groups with < 5% normal forms: 36% of these men had a fertilization rate > 50% using conventional IVF, showing that fertilization capacity is not necessarily impaired even in this 'poor prognosis' group. With the exception of the ICSI group with 5-9% normal forms, the rate of spontaneous abortion in this study was similar to or lower than in our IVF/ICSI programme overall. When the 5-9% normal spermatozoa group was divided into those with teratozoospermia as the only factor and those with additional sperm factors, the increased abortion rate was found in the group with multiple sperm factors (67% spontaneous abortions).

Morphology and clinical outcomes of embryos after in vitro fertilization are superior to those after intracytoplasmic sperm injection

Objective: To compare embryos obtained after IVF and intracytoplasmic sperm injection (ICSI) regarding morphology and the likelihood of achieving clinical pregnancy. Design: Case-control study. Setting: An IVF unit controlling 1,310 cycles in 1996. Patient(s): Women having a total of 477 IVF and 475 ICSI consecutive cycles. Intervention(s): Ovarian stimulation, IVF-ET, or ICSI-ET for all couples. Main Outcome Measure(s): Number of grade-A embryos transferred, preclinical pregnancy losses, and clinical pregnancy rates in IVF and ICSI cycles. Result(s): In comparison with the ICSI group, the IVF group showed significantly more grade-A embryos available for transfer (mean, 2 ± 1.6 versus 1.8 ± 1.5), significantly fewer preclinical pregnancy losses (1.6% versus 4%), and significantly higher clinical pregnancy rates (25% versus 19.1%). Conclusion(s): Embryos obtained after IVF are superior to those obtained after ICSI in relation to embryo morphology and the likelihood of achieving clinical pregnancy.

Clinical outcome of cryopreserved human pronuclear stage embryos resulting from intracytoplasmic sperm injection

Objective: To compare the survival rate and pregnancy rate (PR) of embryos from intracytoplasmic sperm injection (ICSI) or conventional IVF, which were cryopreserved at the pronuclear stage in cycles where fresh transfer was deferred. Design: Comparative observational study. Setting: University-associated IVF center. Patient(s): Ninety-nine patients who deferred ET and had all their embryos cryopreserved at the pronuclear stage after 153 oocyte retrievals. Thirty-nine patients had their oocytes inseminated by ICSI and 60 patients had conventional IVF insemination. Intervention(s): All embryos were frozen-thawed at the two pronuclear stage and allowed to cleave for 2 days before transfer. Main Outcome Measure(s): Survival rate (morphologically intact after thaw), cleavage rate (cleaved by time of transfer), and the clinical PR after frozen ET. Result(s): In the ICSI group, 205 embryos were thawed for use in 57 frozen ETs; in the IVF group, there were 527 embryos thawed for use in 149 frozen ETs. There was no significant difference in any of the outcome measures by insemination method: survival rates (ICSI, 93.2%; IVF, 94.8%); cleavage rates (ICSI, 95.2%; IVF, 94.7%), and clinical PR (ICSI, 14.0%; IVF, 17.4%). Conclusion(s): Pronuclear embryos resulting from ICSI can be cryopreserved successfully, thawed, and the survival rate and PR are comparable to conventional IVF.

Andrology: Preferences for intracytoplasmic sperm injection versus donor insemination in severe male factor infertility: a preliminary report

With the advent of intracytoplasmic sperm injection (ICSI), our programme noted a drop in the number of couples using donor insemination (DI) for severe male factor infertility. Over the first 8 months in which our infertility programme offered both treatments, 27 consecutive couples scheduled for ICSI and 15 consecutive couples scheduled for DI were evaluated Since all patients in our infertility programme beginning in-vitro fertilization (IVF) with planned ICSI or starting DI undergo a semi-structured psychological interview, the psychologist's clinical notes as well as the medical chart were reviewed and coded retrospectively to determine factors related to a couple's treatment choice. Couples who chose IVF-ICSI over DI had a higher occupational status and included husbands with higher educational levels. Their most common motivation was to have the husband's biological child (93% of couples in the ICSI group). The most common motivation for choosing DI (60% of DI couples) was that IVF was not financially affordable. Choice of treatment was not related to psychological adjustment, the husband having prior biological children, or his risk of passing on a genetic defect to offspring. These preliminary data raise the concern that, with the success of ICSI, DI may change in the USA from being an option dictated by semen quality to a second choice treatment utilized for economic reasons.

Endometrial thickness as a predictor of pregnancy after in-vitro fertilization but not after intracytoplasmic sperm injection

An ultrasonographic evaluation of the endometrium was performed in 158 patients undergoing ovarian stimulation for an in-vitro assisted reproduction programme. Endometrial thickness was evaluated in 109 patients undergoing in-vitro fertilization (IVF) for female indications and in 49 patients undergoing intracytoplasmic sperm injection (ICSI) for male indications. The maximal endometrial thickness was measured on the day of human chorionic gonadotrophin (HCG) administration by longitudinal scanning of the uterus on the frozen image using electronic callipers placed at the junction of the endometrium-myometrium interface at the level of the fundus. Cases in which the endometrial thickness was >/=10 mm were included in group A; cases in which the endometrial thickness was <10 mm were assigned to group B. The age of the patients, serum 17-beta oestradiol concentrations on the day of HCG administration, the length of follicular stimulation, the number of follicles, 17-beta oestradiol concentrations per follicle on the day of HCG and the number of embryos transferred were analysed in each case. When comparing endometrial thickness and results in IVF and ICSI patients, an endometrium <10 mm predominated in IVF patients (27.5%) compared with those undergoing ICSI (16.7%) (P = 0.05); conversely an endometrium >=10 mm was more frequent in ICSI than in IVF patients. The incidence of pregnancy was higher in IVF group A patients (32/79; 41%) than in IVF group B patients (5/30; 17%) (P = 0.03), whereas no significant difference was found between ICSI group A (13/42; 31%) and ICSI group B (3/7; 43%) patients. Thus, a higher percentage of IVF patients had thin endometrium when compared with ICSI patients; thin endometrium was a prognostic indicator of pregnancy only in the case of a female indication for infertility (IVF). A thin endometrium in cases of female infertility may reflect a previous or present uterine pathology, whereas in indications of male infertility (i.e. cases using ICSI), in the absence of any associated uterine pathology, the presence of a thin endometrium is not predictive.

Relationship between anticomplement in seminal plasma and complement in follicular fluid associated with outcome in an in vitro fertilization program.

The study was conducted to evaluate levels of anticomplement in seminal plasma and levels of complement in follicular fluid, in correlation with fertilization and pregnancy rate after in vitro fertilization and intracytoplasmic sperm injection programs. Anticomplement levels were determined in 70 couples undergoing in vitro fertilization therapy. In 15 of these couples, complement levels were measured. Anticomplement and complement levels were also determined in an additional 21 couples (apart from the 70 couples) undergoing intracytoplasmic sperm injection treatment. A correlation was found between fertilization rate and anticomplement levels in the seminal plasma (r = 0.4, P < 0.01) after standard in vitro fertilization. No correlation was found in the intracytoplasmic sperm injection group, or observed between complement levels and any parameter examined in both groups. Pregnancy occurred only in those couples with an anticomplement:complement ratio below 0.49. Determination of anticomplement and complement levels may contribute to the assessment of a successful outcome of in vitro fertilization/intracytoplasmic sperm injection.

A comparative analysis of embryo implantation potential in patients with severe teratozoospermia undergoing in-vitro fertilization with a high insemination concentration or intracytoplasmic sperm injection

The objective of this study was to assess fertilization, implantation and pregnancy rates in infertile patients with severe teratozoospermia [P (poor prognosis) pattern sperm morphology assessed by strict criteria] treated by in-vitro fertilization (IVF) using a high insemination concentration (HIC), or by intracytoplasmic sperm injection (ICSI). This was a retrospective cohort study performed in an academic tertiary institution. The outcome of 115 consecutive ICSI cycles was compared to that of a similar number of cycles of IVF with HIC performed during a similar time frame and matched by woman's age and basal serum (cycle day 3) follicle stimulating hormone concentrations. The inclusion criteria were sperm morphology </=4% normal forms (P pattern) and >/=1x10(6) total motile spermatozoa per ejaculate. The diploid fertilization rate in the HIC-IVF group was 86% and in the ICSI group 68% (P < 0.05). Importantly, an equal number of embryos was transferred to both groups of patients. The morphological quality of the embryos (proportion of transfers having superior morphology embryo scores) was significantly better in the ICSI group than in the patients receiving HIC-IVF. Although there was a clear trend for better implantation and pregnancy rates in the ICSI group, these differences were not statistically significant. We conclude that, although HIC-IVF resulted in a higher fertilization rate than ICSI in patients with severe teratozoospermia, ICSI produced a significantly higher proportion of morphologically superior embryos with a tendency towards a higher implantation potential. Therefore, teratozoospermic patients having adequate numbers of motile spermatozoa should be offered ICSI as an alternative to modified (HIC) IVF treatment.

Cryopreservation of human oocytes and fertilization by two techniques: in-vitro fertilization and intracytoplasmic sperm injection

Human oocyte cryopreservation results in poor survival and subsequent fertilization rates. It has been suggested that freeze-thaw-induced changes in the zona pellucida may impair sperm penetration or attachment. The aim of this study was to compare fertilization and cleavage rates in cryopreserved oocytes inseminated by conventional in-vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI). A total of 220 oocytes, obtained from volunteers who had undergone ovarian stimulation, were cryopreserved using a slow freeze-rapid thaw protocol with 1.5 M propanediol as the cryoprotectant. Surviving oocytes (n = 74, 34.4%) were randomly allocated for fertilization by conventional IVF (group 1) or ICSI (group 2) using cryopreserved spermatozoa from a single donor of proven fertility. Fertilization was achieved in five (13.5%) of the oocytes in group 1 and 17 (45.9%) in group 2 (P < 0.005), with only one oocyte in group 1 exhibiting normal fertilization as opposed to 16 (43.2%) in group 2 (P < 0.001). Similarly, one oocyte fertilized by IVF cleaved, while all fertilized with ICSI cleaved (P < 0.001). We conclude that although the survival of oocytes is poor following cryopreservation, fertilization and cleavage rates can be enhanced significantly using ICSI. These data also suggest that the method of cryopreservation used in this study affected the zona pellucida, such that normal sperm attachment or penetration was impaired.

Fertilization and early embryology: Intracytoplasmic sperm injection of mouse oocytes with 5 mM Ca2+ at different intervals

The objective of this investigation was to determine whether intracytoplasmic sperm injection (ICSI) can be performed in the mouse. Metaphase II oocytes were obtained from F1 hybrid mice (C57BL x CBA) by i.p. injections of 10 IU pregnant mare's serum gonadotrophin (PMSG) and human chorionic gonadotrophin (HCG) administered 48 h apart. Oocytes with cumulus oophorus were retrieved 13-14 h post HCG. Cumulus was dispersed with 0.1% hyaluronidase. Mouse spermatozoa were obtained from the cauda epididymides of males of the same strain. The spermatozoa were processed by the standard swim-up procedure. The harvested spermatozoa were then incubated for 1.5 h to allow capacitation. Healthy oocytes were injected with 3-4 pl 5 mM Ca2+, followed by one live morphologically normal spermatozoon into the cytoplasm at intervals of 0, 0.5, 1, 2 and 3 h. The proportion of 2-cell embryos that developed from oocytes injected with Ca2+ and spermatozoa ranged between 29.5 and 36.5% in all groups, with no statistical difference between treatments. Chromosomal analysis showed that two-thirds of the ICSI-derived 2-cell embryos were diploid. The proportion of parthenogenetically activated embryos in the ICSI groups was similar to that in the control group (8-10%) which was injected with Ca2+ and polyvinyl pyrrolidone only. The proportion of blastocysts that developed in culture from the ICSI-derived 2-cell embryos was of the order of 36-42%. Some blastocysts were used for cell number counts. There was a significant increase in total and inner cell mass counts of blastocysts in which the spermatozoon was injected at 2 and 3 h following Ca2+.(ABSTRACT TRUNCATED AT 250 WORDS)

Experience with subzonal insemination (SUZI) and intracytoplasmic sperm injection (ICSI) on unfertilized aged human oocytes

The aim of this study was to assess the fertilizability of unfertilized aged human oocytes from failed in vitro fertilization (IVF) cycles using SUZI and ICSI. A total of 363 oocytes which showed no fertilization after conventional IVF was subjected to assisted fertilization using SUZI or ICSI. The microinjected oocytes which were derived from 72 patients undergoing their first IVF treatment had an intact polar body and no signs of degeneration. SUZI was carried out in 265 oocytes and ICSI in the remaining 98. Significantly more oocytes were damaged after ICSI (9 vs 0.3%, P < 0.01). Normal fertilization rates were higher at 24 hr in both groups and occurred more frequently after ICSI, although the difference did not reach statistical significance. Abnormal fertilization occurred significantly more often after SUZI at 48 hr (P < 0.005), but not at 24 hr. Cleavage rates were significantly higher after ICSI (94.4 vs 57.1%, P < 0.025) at 24 hr, but this was not observed at 48 hr, although the ICSI group still showed better cleavage rates (33.3 vs 19.1%). There was no difference in embryo quality in either group. Our results indicate that micromanipulation rather than reinsemination should be carried out on unfertilized human oocytes from failed IVF attempts. Both techniques can be used to achieve fertilization which occurs more often after ICSI. However, the trauma from the former technique on the microinjected oocytes may impair the potential of the generated embryos to achieve pregnancy compared to SUZI. Prospective randomized trials are necessary to address the problem.