Effect of oocyte donor stimulation on recipient outcomes: data from a US national donor oocyte bank.

STUDY QUESTION: How does ovarian stimulation in an oocyte donor affect the IVF cycle and obstetric outcomes in recipients? SUMMARY ANSWER: Higher donor oocyte yields may affect the proportion of usable embryos but do not affect live birth delivery rate or obstetric outcomes in oocyte recipients. WHAT IS KNOWN ALREADY: In autologous oocyte fresh IVF cycles, the highest live birth delivery rates occur when ~15-25 oocytes are retrieved, with a decline thereafter, perhaps due to the hormone milieu, with super-physiologic estrogen levels. There are scant data in donor oocyte cycles, wherein the oocyte environment is separated from the uterine environment. STUDY DESIGN, SIZE, DURATION: This was a retrospective cohort study from 2008 to 2015 of 350 oocyte donors who underwent a total of 553 ovarian stimulations and oocyte retrievals. The oocytes were vitrified and then distributed to 989 recipients who had 1745 embryo transfers. The primary outcome was live birth delivery rate, defined as the number of deliveries that resulted in at least one live birth per embryo transfer cycle. PARTICIPANTS/MATERIALS, SETTING, METHODS: The study included oocyte donors and recipients at a donor oocyte bank, in collaboration with an academic reproductive endocrinology division. Donors with polycystic ovary syndrome and recipients who used gestational carriers were excluded. The donors all underwent conventional ovarian stimulation using antagonist protocols. None of the embryos underwent pre-implantation genetic testing. The average (mean) number of embryos transferred to recipients was 1.4 (range 1-3). MAIN RESULTS AND THE ROLE OF CHANCE: Per ovarian stimulation cycle, the median number of oocytes retrieved was 30 (range: 9-95). Among the 1745 embryo transfer cycles, 856 of the cycles resulted in a live birth (49.1%). There were no associations between donor oocyte yield and probability of live birth, adjusting for donor age, BMI, race/ethnicity and retrieval year. The results were similar when analyzing by mature oocytes. Although donors with more oocytes retrieved had a higher number of developed embryos overall, there was a relatively lower percentage of usable embryos per oocyte warmed following fertilization and culture. In our model for the average donor in the data set, holding all variables constant, for each additional five oocytes retrieved, there was a 4% (95% CI 1%, 7%) lower odds of fertilization and 5% (95% CI 2%, 7%) lower odds of having a usable embryo per oocyte warmed. There were no associations between donor oocyte yield and risk of preterm delivery (<37 weeks gestation) and low birthweight (<2500 g) among singleton infants. LIMITATIONS, REASONS FOR CAUTION: Ovarian stimulation was exclusively performed in oocyte donors. This was a retrospective study design, and we were therefore unable to ensure proportional exposure groups. These findings may not generalizable to older or less healthy women who may be vitrifying oocytes for planned fertility delay. There remain significant risks to aggressive ovarian stimulation, including ovarian hyperstimulation. In addition, long-term health outcomes of extreme ovarian stimulation are lacking. Lastly, we did not collect progesterone levels and are unable to evaluate the impact of rising progesterone on outcomes. WIDER IMPLICATIONS OF THE FINDINGS: Live birth delivery rates remain high with varying amounts of oocytes retrieved in this donor oocyte model. In a vitrified oocyte bank setting, where oocytes are typically sent as a limited number cohort, recipients are not affected by oocyte yields. STUDY FUNDING/COMPETING INTEREST(S): Additional REDCap grant support at Emory was provided through UL1 TR000424. Dr. Audrey Gaskins was supported in part by a career development award from the NIEHS (R00ES026648).

A review of the promises and pitfalls of oocyte and embryo metabolomics.

Embryo viability assessment is one of the most important and challenging tasks in IVF. Evaluation of embryo quality is critical when selecting the best embryo(s) to transfer or cryopreserve. Until recently, the only instrument used for embryo evaluation was the inverted light microscope, which provided information based on morphological characteristics. Developmental and morphological information gained from microscopic assessment have been positively associated with IVF outcomes, including pregnancy and implantation rates. However, based on general statistics, it is clear that IVF currently still results in relatively low pregnancy rates, while simultaneously being associated with relatively high multiple implantation rates. Only with novel embryo assessment and selection procedures would it be possible to improve these outcomes. Accordingly, it has been proposed that it is possible to test the culture environment of a developing embryo to gain valuable information regarding its viability. Different approaches have been used. These include the measurement of oxygen consumption by the embryo and testing of the soluble HLA-G in the environment, as it was proposed that secretion of HLA-G is associated with higher implantation rates. Amino acid turnover, which appears to be correlated to blastocyst development, can be measured as an indication of embryo viability. Other approaches, such as time-lapse video observation or cumulus cell gene expression analysis, may be used in the future to gain a broader understanding of embryo viability. Proteomics and metabolomics are also useful tools for assessment of embryo developmental potential. Results from recent studies on predicting embryo viability by analyzing the metabolome of different stage embryos are promising, as increases in pregnancy and implantation rates were obtained using the metabolomic profile for embryo selection. Several novel approaches are currently being developed to aid in viability assessment. These need to be evaluated in prospective clinical trials, while considering their practicality in the clinical laboratory.

Vitrification in assisted reproduction: myths, mistakes, disbeliefs and confusion.

The purpose of this work is to update embryologists and clinicians on different approaches in human oocyte and embryo cryopreservation, by clarifying some misunderstandings and explaining the underlying reasons for controversial opinions. The work is based on literature review and critical analysis of published papers or conference abstracts during the last 24 years, with special focus on the last 3 years. Due to the latest advancements in techniques, cryopreservation now offers new perspectives along with solutions to many demanding problems, and has developed from a backup procedure to a successful alternative that is an indispensable constituent of assisted reproductive techniques. However, this progress is not free from controversies, at some points is rather serendipitous, and many factors, including human ones, hamper the selection and widespread application of the most efficient technique for the given task. A better understanding of the basic features of the two rival approaches (slow-rate freezing and vitrification), a clarification of terms and technical details, and a balanced, pragmatic evaluation of possible risks and potential, or definite, gains are required to accelerate advancement. Alternatively, the increasing flow of patients to the few assisted reproduction clinics and countries that are highly successful in this field will enforce the required changes in methodology and mentality worldwide.

Co-transfer of embryos derived from cryopreserved and fresh natural cycle oocytes: a pilot study.

Italian legislation regarding reproductive medicine prohibits embryo storage while allowing cryopreservation of supernumerary oocytes. This study evaluated the effect of fresh oocytes obtained from natural unstimulated cycles on the clinical success rates derived from the use of frozen-thawed (FR-TH) oocytes obtained following ovarian stimulation. For 36 women, intracytoplasmic sperm injection was performed on FR-TH oocytes supplemented by a fresh oocyte, if available, derived from a natural cycle in which gonadotrophin-releasing hormone-antagonist was used for premature LH surge control. The retrieval rate of fresh oocytes was 61.1% and survival rate of FR-TH oocytes was 43.6%. The fertilization rate of fresh and FR-TH oocytes was 70% and 52.5%, respectively. Fifty embryos were transferred, 14 of them developed from fresh oocytes and 36 from FR-TH oocytes. Six pregnancies occurred in 10 cycles in which the embryos developed from fresh and FR-TH oocytes (pregnancy rate 60.0%) and two in 12 patients in whom the embryos were obtained from only FR-TH oocytes (pregnancy rate 16.7%) (P < 0.05). In summary, the data demonstrate that the transfer of embryos derived from oocytes cryopreserved following a previous ovarian stimulation and an embryo developed from a fresh one retrieved in natural cycle ensures an excellent clinical outcome.

A highly immunogenic trivalent T cell receptor peptide vaccine for multiple sclerosis.

BACKGROUND: T cell receptor (TCR) peptide vaccination is a novel approach to treating multiple sclerosis (MS). The low immunogenicity of previous vaccines has hindered the development of TCR peptide vaccination for MS. OBJECTIVE: To compare the immunogenicity of intramuscular injections of TCR BV5S2, BV6S5 and BV13S1 CDR2 peptides in incomplete Freunds adjuvant (IFA) with intradermal injections of the same peptides without IFA. METHODS: MS subjects were randomized to receive TCR peptides/IFA, TCR peptides/saline or IFA alone. Subjects were on study for 24 weeks. RESULTS: The TCR peptides/IFA vaccine induced vigorous T cell responses in 100% of subjects completing the 24-week study (9/9) compared with only 20% (2/10) of those receiving the TCR peptides/saline vaccine (P =0.001). IFA alone induced a weak response in only one of five subjects. Aside from injection site reactions, there were no significant adverse events attributable to the treatment. CONCLUSIONS: The trivalent TCR peptide in IFA vaccine represents a significant improvement in immunogenicity over previous TCR peptide vaccines and warrants investigation of its ability to treat MS.

Laser-assisted ICSI: a novel approach to obtain higher oocyte survival and embryo quality rates.

BACKGROUND: Degeneration of oocytes occurs even when maximum care is exercised during ICSI, especially when the oolemma is very fragile and/or the zona pellucida is resistant. In order to be able to minimize the risk of degeneration associated with microinjection this study applied a new method: a microhole on the zona pellucida of the oocyte was drilled by laser beam just prior to ICSI to permit the penetration of the microneedle without any trauma. METHODS: A total of 32 patients (32 cycles) who had one or more previously failed ICSI cycles with a high degeneration rate of oocytes (>20%) were included in the study. Oocytes of the same patients were randomly divided into the study group [laser-assisted ICSI (LA-ICSI)] and the control group [conventional ICSI (C-ICSI)]. The outcomes of the cycles were compared and analysed. RESULTS: After LA-ICSI compared with C-ICSI, survival rates of oocytes were 99.6 and 84% (P < 0.0001), fertilization rates were 76.6 and 68.6% (not significant) and embryo development rates ( vertical line 6 cells on day 3) were 76.5 and 57.3% (P = 0.0024) respectively. CONCLUSIONS: Creating a microhole on the zona pellucida of the oocyte by laser beam prior to ICSI provides a less traumatic penetration of the injection needle into the ooplasm and results in lower degeneration and higher embryo development rates than C-ICSI in patients with fragile oocytes.

Monozygotic twins and transfer at the blastocyst stage after ICSI.

The incidence of monozygotic twinning (MZT) is higher in pregnancies conceived after assisted reproduction than after natural conception. Alterations, produced by ovarian stimulation, in-vitro culture conditions and specifically alterations of zona pellucida are mentioned as possible causes of this phenomenon. A retrospective review was performed of the incidence of MZT in pregnancies generated in our centre during the period of January 1996 to December 1999. This variable was compared in 129 gestations that resulted from blastocyst transfer (occurring from September 1998 to August 1999) with 814 pregnancies produced by transfers of 4- to 8-cell embryos. Follicular development was induced with human menopausal gonadotrophin and urinary FSH during 1996 and 1997 and with recombinant FSH during 1998 and 1999. Blastocysts were cultured in sequential media using S1 or G1 up to 72 h and S2 or G2 to day 5. Five of the 129 pregnancies generated by blastocyst transfers were complicated by MZT gestation (3.9%). In comparison, only six of 814 pregnancies occurred from 4- to 8-cell transfers (0.7%), a difference that is statistically significant (P< 0.001 with Yates correction). The results confirm an increase of MZT in pregnancies from intracytoplasmic sperm injection as compared to the natural incidence. Moreover, the frequency of MZT was significantly higher when transfers were performed at the blastocyst stage, suggesting that extended in-vitro culture of embryos may be associated with alterations of the zona pellucida and the hatching process.

Fertilizable oocytes reconstructed from patient's somatic cell nuclei and donor ooplasts.

The only assisted reproduction treatment now available for women with ovarian failure or irreparable oocyte defects is oocyte donation. However, some women experience psychological barriers to the recourse to donor oocytes, related to the lack of contribution of their proper genes to the progeny. A pilot study in humans suggests that this problem may be overcome by the development of techniques for haploidization of somatic cell nuclei, allowing the formation of new oocytes bearing the complete nuclear genome of the patient. Somatic cell nuclei were obtained from cumulus cells of a patient who failed to produce fertilizable oocytes and were transferred into enucleated oocytes (ooplasts) from a donor. Out of six ooplasts injected with the somatic cell nuclei and fertilized with spermatozoa from the patient's husband, signs of haploidization were detected in three oocytes, two of which subsequently started embryonic development and were cryopreserved for eventual future transfer to the genetic mother. These data show that human oocytes can be used for both reprogramming and haploidization of somatic cell nuclei, allowing reconstruction of genetically own oocytes for patients without, or with seriously disturbed, ovarian function.

Sperm centriole disfunction and sperm immotility.

The two main causes of complete or nearly complete asthenozoospermia are necrozoospermia (presence of only non-viable spermatozoa) and the different ultrastructural abnormalities of spermatozoa. Ultrastructural alterations may affect also the function of the sperm centrosome, which can result in impaired motility. Because in human the inheritance of the centrosome is paternal and thus linked to the sperm, morphological or functional alterations of it can also be associated with fertilization abnormalities of the oocyte and cleavage irregularities of the embryo. Most of the cases of asthenozoospermia can be treated efficiently by intracytoplasmic sperm injection (ICSI) using ejaculated sperm (from repeated ejaculation) in combination with hypo-osmotic swelling test (HOST) or using testicular sperm depending on the etiology of the impairment. Replacement of abnormal centriole using donor sperm is a theoretical possibility, but at present it is not an efficient method.

Chemically and mechanically induced membrane fusion: non-activating methods for nuclear transfer in mature human oocytes.

Most current studies of nuclear transfer in mammalian oocytes have used electrofusion to incorporate donor cell nuclei into enucleated oocyte cytoplasts. However, the application of electrofusion to human oocytes is hampered by the relative ease with which this procedure induces oocyte activation. Here we tested a previously described chemical fusion technique and an original mechanical fusion procedure in this application. Enucleated metaphase II oocytes were first agglutinated with karyoplasts originating from other metaphase II oocytes and then induced to fuse with the use of polyethylene glycol or by micromanipulation with an intracytoplasmic sperm injection (ICSI) micropipette. Both techniques yielded a high frequency of fusion and did not cause oocyte activation. Moreover, the reconstructed oocytes were easily activated by subsequent treatment with ionophore A23187 and 6-dimethylaminopurine. These techniques may be used in attempts to alleviate female infertility due to insufficiency of ooplasmic factors by nuclear transfer from patients' oocytes to enucleated donor oocyte cytoplasts. For eventual future use in human cloning, they would ensure prolonged exposure of transferred nuclei to metaphase promoting factor, which appears to be required for optimal nuclear reprogramming.

[Dynamic MR examination of the breast--histological correlations]. / Az emló dinamikus MR-vizsgálata--szövettani korreláció.

The authors outline dynamic MR mammography (dMRM) as a highly sensitive diagnostic method for the examination of the breast. In a retrospective study relating to 84 processed cases, in the knowledge of the cytological-histological findings the diagnostic accuracy of the examinations was determined. The role of the method in detecting benign and malignant changes of the breast has been estimated. Misdiagnosed cases have been analysed and recommendations for the application of the method are included. The MR proved to be positive in 32 cases and negative in 3 cases of the analysed 35 malignant tumors. Benign lesions were found at microscopy in 49 cases, of which MR correctly diagnosed 40. The sensitivity and the specificity of dynamic MR mammography were 91% and 82%.

Reproductive capacity of spermatozoa from men with testicular failure.

Controversial reports have been published about the influence of sperm source and of the underlying testicular pathology on success rates of intracytoplasmic sperm injection (ICSI). In this controlled study, ICSI treatment cycles with testicular spermatozoa from men with obstructive and non-obstructive azoospermia were compared with ICSI ejaculated sperm cycles with semen parameters < or = 5 x 10(6)/ml and < or = 10% progressive motility. The control cases were matched for female age, rank of trial, female basal follicle-stimulating hormone serum concentrations and close proximity to the study group's procedure. The fertilization, cleavage, pregnancy and abortion rates were similar in matched groups irrespective of the type of azoospermia. However, the implantation rate in the non-obstructive azoospermic patient group was significantly lower than that in the matched ejaculated sperm group (13.4% versus 26%, P = 0.05). On the other hand, no impairment of the implantation rate was observed in the obstructive azoospermic patient group. These data show that testicular pathology has a negative impact on reproductive performance of testicular spermatozoa, resulting in a decreased implantation potential without any apparent effect on fertilization and early preimplantation development.

Effect of timing of oocyte denudation and micro-injection on survival, fertilization and embryo quality after intracytoplasmic sperm injection.

In human in-vitro fertilization (IVF), the oocytes are surrounded by cumulus and corona cells at the time of insemination so that their maturity cannot easily be evaluated. The best IVF results are obtained if the oocytes are inseminated 2-6 h after retrieval. In the intracytoplasmic sperm injection (ICSI) procedure, the oocytes are denuded by enzymatic and mechanical treatment in order to be able to perform the injection. As a consequence, the nuclear maturity of the oocytes can be evaluated and only those that have extruded the first polar body are injected. However, metaphase-II oocytes that have not yet reached cytoplasmic maturity cannot be recognized. The purpose of this study was to investigate the effect of different timing of cumulus-corona cell removal and injection on the outcome of ICSI. For this we allowed the oocytes to complete in-vitro cytoplasmic maturation in two different culture conditions: (i) surrounded by their cumulus and corona cells or (ii) totally denuded. We performed three different studies on sibling oocytes obtained after a standardized buserelin/human menopausal gonadotrophin (HMG) protocol. We investigated the effect of early (1-2 h after retrieval) and late (5-6 h after retrieval) oocyte denudation and injection on the survival and fertilization of the injected oocytes and on embryo cleavage after fertilization. We found no statistically significant differences between early and late injection, indicating that after a standardized buserelin/HMG protocol the metaphase-II oocytes do not need time for further cytoplasmic maturation. Furthermore, a different timing of cumulus-corona cell removal has no effect on the outcome of ICSI, suggesting that the surrounding cells are not necessary for survival, fertilization and cleavage after ICSI.

Timing of oocyte activation, pronucleus formation and cleavage in humans after intracytoplasmic sperm injection (ICSI) with testicular spermatozoa and after ICSI or in-vitro fertilization on sibling oocytes with ejaculated spermatozoa.

In the first study, we evaluated 101 oocytes [2, 4, 6, 8, 16, 18 and 20 h after intracytoplasmic sperm injection (ICSI)] that had been microinjected with testicular spermatozoa. Of the 70 normally fertilized oocytes (69%) 30 (43%) had two pronuclei by 6 h after ICSI. Fifty-one (73%) by 8 h, 69 (99%) by 16 h and four of them by 20 h cleaved to the 2-cell stage. In the second study, 95 cumulus-corona-oocyte complexes (CCOC) were divided into two groups. Forty-seven CCOC were inseminated by conventional in-vitro fertilization (IVF) and 40 metaphase-II oocytes by ICSI. Oocytes were evaluated at 2, 4, 6 (only after ICSI), 8, 10, 12, 18, 20, 22, 24, 26, 28 and 30 h after both ICSI and IVF. After IVF, 35 oocytes were fertilized normally (75%), four of which (11%) had two pronuclei by 8 h, 11 (31%) by 10 h, 27 (77%) by 12 h and 35 (100%) by 14 h. The first cleavages had occurred by 24 h after insemination (four oocytes, 11%). After ICSI, 34 oocytes were fertilized normally (79%), 13 of which (38%) had two pronuclei by 6 h, 27 (79%) by 8 h and 32 (94%) by 10 h. Three oocytes cleaved by 20 h after microinjection (9%) and 19 by 24 h (56%). Pronuclei developed asynchronously in six oocytes after ICSI (18%) as opposed to 16 oocytes after IVF (46%). The results of this study suggest that the timing of pronuclear formation is no different when a testicular spermatozoon is microinjected into the oocytes from when an ejaculated spermatozoon is injected. Secondly, pronuclear development and first cleavage generally take place 4 h sooner after ICSI than after IVF. On the other hand, a higher proportion of oocytes develop two pronuclei asynchronously after IVF than after ICSI.

Ultrastructure of gametes after intracytoplasmic sperm injection.

The ultrastructure of 18 metaphase-II oocytes after intracytoplasmic sperm injection (ICSI) was analysed from 30 min to 8 h following the microinjection. Three control metaphase-II oocytes were not injected. The 21 oocytes were embedded individually and examined in serial pole-to-pole sections. Seventeen oocytes had been matured in vitro from the germinal vesicle stage and showed ultrastructural alterations due to long-term culture. In microinjected oocytes, membrane-bound vacuoles and oolemma inclusions were found at the injection site. The ICSI oocytes showed evidence of plasma-membrane damage and increased oocyte exchange processes with residual and multivesiculated bodies. Gamete membrane fusion was not observed. Acrosome reaction was observed within the ooplasm 15 min after ICSI. Sperm elements were not incorporated into any oocyte vacuole. Onset of oocyte activation was associated with sperm chromatin decondensation. Cortical granules were released at 60 min and two pronuclei were formed at 6 h after ICSI. The male and female pronucleus formation was asynchronous. The findings suggest specific ICSI-related morphological features of early fertilization.

Special applications of intracytoplasmic sperm injection: the influence of sperm count, motility, morphology, source and sperm antibody on the outcome of ICSI.

The relationship between the three basic parameters of ejaculated spermatozoa, i.e. concentration, motility and morphology, and the results of intracytoplasmic sperm injection (ICSI) were investigated in 838 microinjection cycles. A further 123 ICSI treatment cycles in which testicular spermatozoa were used for microinjection were also evaluated. The influence of anti-sperm antibodies (ASA) on the outcome of ICSI was investigated by analysing 55 cycles where the proportion of ASA-bound spermatozoa was > or =80%. After microinjection, oocyte intactness, fertilization, embryo cleavage, transfer and pregnancy rates were recorded and compared. The results showed that neither the type nor the extent of sperm impairment had an important influence on the outcome of ICSI when ejaculated spermatozoa were used. Only two very rare conditions had a strongly negative influence on the result of ICSI, i.e. where immotile (presumably dead) spermatozoa or where round-headed spermatozoa were injected into the oocyte. Neither the proportion of ASA-bound spermatozoa, the type of dominantly present ASA, nor the location of ASA on the spermatozoa had an important influence on fertilization, embryo development or pregnancy rates after ICSI. In most of the cycles combined with testicular biopsy (79%), there were enough motile spermatozoa present in the wet preparation for injection of all the oocytes. Injection of motile testicular spermatozoa led to a higher normal fertilization rate than did injection of non-motile spermatozoa (65 versus 21%). It can be concluded that injection of motile (living) spermatozoa into oocytes is the most important factor in determining good results with ICSI and that other sperm parameters do not have a strong influence on the outcome of ICSI.

Correlation between motility of testicular spermatozoa, testicular histology and the outcome of intracytoplasmic sperm injection.

The objective of the present study was to analyse the influence of motility on the results of intracytoplasmic sperm injection (ICSI) when testicular spermatozoa are used for microinjection and to correlate this with testicular histology. A total of 197 ICSI treatment cycles (167 couples) was analysed retrospectively in which testicular spermatozoa were used, because of complete azoospermia, for microinjection. Fertilization, embryo cleavage, transfer and pregnancy rates were evaluated and compared in relation to motility of testicular spermatozoa. In 170 cycles, histological diagnoses were compared with findings on motility. Injection of motile testicular spermatozoa (in 159 cycles) provided a higher normal fertilization rate than did injection of non-motile spermatozoa (in 14 cycles; 65 versus 45% respectively). Normal spermatogenesis was diagnosed in a significantly higher proportion and incomplete maturation arrest in a significantly lower proportion in the group of patients in which only motile spermatozoa were used for microinjection (65 and 10%), as compared to the group where exclusively non-motile spermatozoa were used (36 and 36%). Fertilization rate after ICSI was relatively high when non-motile testicular spermatozoa were used for microinjection, but use of motile testicular spermatozoa was associated with a still higher fertilization rate (except when histology of the testicular biopsy showed normal spermatogenesis), and therefore selection of motile testicular spermatozoa is always preferable for ICSI. Normal spermatogenesis predicts a greater probability, and maturation arrest a lower probability of recovering motile testicular spermatozoa.

The predictive value of uterine artery blood flow measurements for uterine receptivity in an intracytoplasmic sperm injection program.

OBJECTIVE: To assess whether uterine artery blood flow impedance, measured as the pulsatility index on the day of ET in patients undergoing IVF-ET with microinjection, can predict the likelihood of pregnancy. DESIGN: Prospective clinical study. SETTING: A tertiary referral center for assisted reproduction. PATIENT(S): Seventy patients undergoing intracytoplasmic sperm injection (ICSI) for andrologic indications. INTERVENTION(S): Transvaginal color Doppler examination performed on the day of ET. MAIN OUTCOME MEASURE(S): Mean (+/- SD) pulsatility index value of the left and right uterine arteries, serum E2 levels, implantation rates, and ongoing pregnancy rates (PRs). RESULT(S): The patients were divided into pregnant and nonpregnant groups and were separated according to whether the pulsatility index was low (1.00-1.99), medium (2.00-2.99), or high (> or = 3.00). The pulsatility index values did not change statistically in the pregnant and nonpregnant groups. The implantation rates were 19.5%, 15.4%, and 25% for the low-, medium-, and high-pulsatility index groups, respectively. The ongoing PRs for the same groups were 35.3%, 26.7%, and 37.5%, respectively. CONCLUSION(S): The study suggests that blood flow, measured as the pulsatility index on the day of ET, cannot predict the likelihood of pregnancy in stimulated cycles of ICSI.

Effects of different hyaluronidase concentrations and mechanical procedures for cumulus cell removal on the outcome of intracytoplasmic sperm injection.

The aim of this study was to compare concentrations of hyaluronidase and mechanical methods used to denude human oocytes from surrounding cumulus and corona cells prior to intracytoplasmic sperm injection (ICSI). Cumulus and corona cells were removed in two pipetting steps: first in a medium containing hyaluronidase, and then in a medium without enzyme. The first step in the procedure was investigated. Different hyaluronidase concentrations (78, 39 or 10 IU/ml) and pipettes of different size (inner diameter 250 or > 1000 microm) were used to remove the cumulus cells. The time required to denude the oocytes was recorded. Metaphase II oocytes were injected, and the survival, fertilization, embryo development and pregnancy rates were evaluated. We found that by using a pipette with an inner diameter of at least 1000 microm we were able to decrease significantly the time an oocyte is exposed to hyaluronidase, even if the concentration of enzyme is very low (10 IU/ml). For the different conditions there was no statistically significant effect on the outcome in terms of survival, normal fertilization [two pronuclear (2PN)], parthenogenetic activation (1PN), abnormal fertilization (3PN), embryo development and pregnancy rates after ICSI. In conclusion, a concentration as low as 10 IU/ml hyaluronidase in combination with a pipette of at least 1000 microm inner diameter can be used successfully to denude oocytes for microinjection.