Carlo Flamigni

Birth of a healthy female after intracytoplasmic sperm injection of cryopreserved human oocytes

OBJECTIVE To describe the first birth achieved after intracytoplasmic sperm injection (ICSI) of cryopreserved human oocytes. DESIGN Case report. SETTING University of Bologna Hospital, Department of Obstetrics and Gynecology, Reproductive Endocrinology Unit, IVF and Infertility Center. PATIENT(S) One patient undergoing IVF. INTERVENTION(S) Transvaginal ultrasound-guided oocyte retrieval followed by oocyte freezing. Artificial preparation of the endometrium with E2 and P, oocyte thawing, and ICSI. RESULT(S) Four of 12 cryopreserved oocytes survived; using ICSI, 2 underwent normal fertilization but only 1 cleaved. One good-quality 4-cell embryo was transferred. A single gestation was confirmed by ultrasound at the 7th week. Amniocentesis was performed at the 16th week and demonstrated a normal female karyotype of 46,XX. After a normal pregnancy, a healthy female infant was born at the 38th week of gestation. CONCLUSION(S) The combination of ICSI and oocyte cryopreservation is a new tool in assisted reproductive technology.

Clinical experience and applications of oocyte cryopreservation

Oocyte cryopreservation is a viable solution for the ethical problems related to embryo storage, and the only available technique for preservation of fertility in women who have to undergo chemo- or radiotherapy. The main problems with oocyte cryopreservation are concerned with the survival rate and the fertilization rate. Recently the introduction of the intracytoplasmic sperm injection (ICSI) led to an increase in the fertilization rate. The success achieved with the first case treated encouraged us to set up a clinical trial on human oocyte cryopreservation. In the first stage of the study, 23 women with tubal infertility were enrolled. Superovulation was induced and 375 oocytes were retrieved; of these 338 oocytes were frozen. The survival rate was 59.5% and was independant of the duration of cryopreservation or the presence of cumulus. The normal fertilization rate was 64.4%, and only 7.5% of fertilizations were abnormal. A total of 90.8% of fertilized oocytes cleaved. A mean of 3.1+/-1.3 embryos per patient were transferred. Three pregnancies were achieved. In the second stage of our investigation, more patients were enrolled and similar results were observed. Sixteen pregnancies were achieved. A further stage of the investigation involved the fertilization of frozen oocytes with frozen sperm and even these resulted in a pregnancy. Our study demonstrated that pregnancies can also be achieved when frozen eggs are fertilized by testicular and epididymal sperm. As a consequence of the success of our investigations, a program of oocyte cryopreservation for oncological patients has been initiated in our centre. In our opinion, oocyte cryopreservation is, at present, a safe and efficient technique as documented by the birth of several healthy children.

Differential sucrose concentration during dehydration (0.2 mol/l) and rehydration (0.3 mol/l) increases the implantation rate of frozen human oocytes

Novel protocols have increased survival and fertilization rates of cryopreserved oocytes. Nevertheless, in most cases clinical experiences have been disappointing or contradictory. Human oocytes of 141 patients were cryopreserved using a modified slow-cooling protocol involving 1.5 mol/l propane-1,2-diol (PrOH) and 0.2 mol/l sucrose during dehydration, while rehydration was conducted applying decreasing concentrations of PrOH and 0.3 mol/l sucrose. One thousand and eighty-three oocytes were frozen and 403 were thawed, with a survival rate of 75.9%. Among the 306 surviving oocytes, 252 were microinjected and 192 (76.2%) showed two pronuclei. One hundred and eighty zygotes (93.8%) cleaved. The proportion of good quality embryos (grade I and II) was 86.2%. All embryos were transferred and 17 clinical pregnancies were obtained. Pregnancy rates were 21.3% per transfer, 21.8% per patient, and 18.9% per thawing cycle. The implantation rate was 13.5% while the miscarriage rate was 11.8%. To date, four babies have been delivered, while the remaining pregnancies are ongoing. Increased oocyte survival rates can be achieved by moderately high sucrose concentrations in the freezing and thawing solutions. This also ensures elevated success rates in terms of fertilization, embryo development and clinical outcome.

The role of leiomyomas in infertility

STUDY OBJECTIVE To assess the role of leiomyomas and their surgical removal on pregnancy rates. DESIGN (Canadian Task Force classification II-1). Setting. Academic center. PATIENTS Two hundred twelve women who were investigated for infertility. INTERVENTION Laparoscopic myomectomy. MEASUREMENTS AND MAIN RESULTS Patients were divided according to case control criteria as those who underwent laparoscopic removal of myomas (106) and those who did not (106); both groups were compared with 106 women with unexplained infertility without myomas. Of the 318 women, 83 (26%) became pregnant and delivered live infants. The 44 (42%) who underwent surgical removal of leiomyomas had higher delivery rates than 12 (11%) who did not undergo surgery (p <0.001) and 27 (25%) who did not have myomas (p <0.001). Patients whose myomas were not surgically treated had fewer deliveries than women who did not have myomas (12 vs 27, p <0.002). Fifteen women had spontaneous abortions before week 12: 3 (3%) who had surgery, 10 (9%) who did not have surgery, and 2 (2%) who did not have myomas. CONCLUSION Laparoscopic myomectomy improved pregnancy rates over nonsurgical management of myomas.

Adhesion formation after laparoscopic myomectomy

STUDY OBJECTIVE To determine the frequency of adhesion formation after myomectomy performed by operative laparoscopy or laparotomy. DESIGN Case-control study. Setting. Academic womens hospital. PATIENTS Thirty-two premenopausal women scheduled for myomectomy by one of two techniques. INTERVENTIONS Surgical removal of myomata. MEASUREMENTS AND MAIN RESULTS Of the 32 women, 16 underwent laparotomy and 16 laparoscopy. Second-look laparoscopy was performed in 28 patients, at which time adhesions were lysed. Compared with laparotomy, laparoscopy resulted in adhesions in significantly fewer patients, and in significantly lower scores when adhesions were detected. CONCLUSION Laparoscopic removal of uterine myomata is associated with fewer adhesions than removal by laparotomy.

Characteristics of uterine contractility during menses in women with mild to moderate endometriosis

OBJECTIVE To establish the role of uterine contractions in retrograde menstruation with subsequent abdominal implantation of endometrial tissue. DESIGN Controlled prospective study. SETTING University hospital-based study. PATIENT(S) Infertile women with (n = 22) and without (n = 22) endometriosis. MAIN OUTCOME MEASURE(S) Frequency, amplitude, and basal pressure tone of uterine contractions; correlation of contractions with retrograde bleeding and presence of viable endometrial cells; and dysmenorrhea before and 3 and 24 months after surgery. RESULT(S) Compared with controls, patients with endometriosis had uterine contractions with higher frequency (22.73 +/- 5.66 osc/10 min vs. 11.09 +/- 3.26 osc/10 min), amplitude (20.83 +/- 3.94 mm Hg vs. 6.77 +/- 2.83 mm Hg), and basal pressure tone (50.14 +/- 16.30 mm Hg vs. 24.68 +/- 6.14 mm Hg). Dysmenorrhea was scored as 4.09 +/- 1.44 in patients with endometriosis and 0.86 +/- 1.42 in controls. Retrograde bleeding was found in 73% of patients with endometriosis vs. 9% of controls, and only 45% of patients with endometriosis had viable endometrial cells in the cul-de-sac. CONCLUSION(S) Endometriosis may result from abnormal myometrial contractility through tubal transportation, dissemination, and implantation of endometrial viable cells into the abdomen.

Cumulative pregnancy rates resulting from the use of fresh and frozen oocytes: 7 years’ experience

Storing supernumerary embryos and transferring them later fully utilizes the reproductive potential of retrieved oocytes, allowing a significant increase in the overall number of pregnancies achieved from a single cycle of ovarian stimulation treatment. As an alternative to embryo cryopreservation, preservation of unfertilized oocytes has been proposed to maximize clinical outcome. This paper presents data concerning the cumulative pregnancy rate after use of fresh and cryopreserved oocytes. In 80 treatment cycles in which patients chose to have only a few fresh oocytes inseminated, 24 pregnancies were obtained (30.0%), with an implantation rate of 22.6%. After cryopreservation with the standard slow-cooling protocol, the survival, fertilization and cleavage rates of 918 frozen oocytes were 43.4, 51.5 and 86.0% respectively. A total of 14 frozen pregnancies were achieved, with pregnancy rate 19.2% per transfer and implantation rate 12.3%. The cumulative pregnancy rate was 47.5% per patient. Therefore, despite a low rate of oocyte post-thaw survival, it appears that oocyte storage appreciably improves the number of pregnancies per treatment cycle in cases in which only a minority of oocytes are destined for the fresh treatment. This outcome provides valuable information for appraising the chances of clinical success when the option of embryo cryopreservation is not available.

Longitudinal Change of Sonographic Ovarian Aspects and Endocrine Parameters in Irregular Cycles of Adolescence

ABSTRACT: We longitudinally studied clinical endocrine and ultrasound parameters of the ovaries in 73 healthy adolescents having persistent menstrual irregularities. After the first examination, they were reexamined after a variable period ranging from 2 to 7 y. During the first examination, three basic features of the ovaries were observed: homogeneous (36%), multifollicular (23%), and polycystic (41%). Polycystic ovaries were most frequent, and they generally exceeded the normal adult range. During the last examination, in the entire group of irregular adolescents, homogeneous ovaries decreased (–14%), polycystic ovaries increased (+ 18%), and a further higher number of subjects exceeded the normal adult range (+10%). The subjects with enlarged ovaries had the highest values of LH, testosterone, and androstenedione. Fourteen subjects out of 46 (30%), with normal ovarian volume in the first examination, registered an ovarian enlargement in the last examination, exceeding the normal range. Moreover, a change from the homogeneous or multifollicular structure to the polycystic one was observed. Twenty-one subjects out of 27 (78%) with enlarged ovaries in the first examination confirmed the high ovarian volume and the unchanged structure in the last examination, whereas six subjects (22%) showed ovaries within the normal adult range; the polycystic structure was substantially confirmed. These results indicate the following. 1) Homogeneous, multifollicular, and polycystic ovaries can usually be found in the postmenarcheal period. 2) Enlarged ovaries, polycystic structure, hyperandrogenemia, and high LH values are strongly linked, and they are frequent in irregular cycles even in the absence of signs of hyperandrogenism. These characteristics may all persist or in various aggregations become a permanent feature. 3) Only a few subjects may lose ovarian enlargement and show a change in the polycystic structure; however, they frequently maintain hyperandrogenemia. 4) During the postmenarcheal period, normal ovarian characteristics may suddenly change, and the ovaries may take on a polycystic structure and increase in volume. Moreover, some endocrine parameters may reach pathologic levels.

Predictive factors for embryo implantation potential

In spite of recent improvements in IVF, pregnancy rates have not increased significantly and one of the major problems remains the high multiple pregnancy rate. Better criteria are therefore necessary to establish the viability of a transferable embryo. Early prognosis of the developmental fate of the oocyte would help in selecting the best embryos to transfer, but non-invasive selection at the oocyte stage (extracytoplasmic and intracytoplasmic morphology) has proved to be of little prognostic value. Recently, it has been shown that follicular vascularization appears to be predictive of oocyte developmental fate, making it a good first-step approach for selection. Observation of pronuclei patterns at the zygote stage appears to offer an additional prognostic tool, correlating well with IVF outcome. Morphological evaluation of the embryo at days 2-3 remains the most used and valid method of selection, even though it is not sufficient to select embryos with the higher implantation potential. Blastocyst culture is another possible strategy for selecting the best embryos with reduced risk of aneuploidies, though not all major chromosomal aberrations are excluded by prolonged in-vitro culture. In summary, selecting the best embryo for transfer is a decision that should be based on choices made during the different stages of assisted reproductive technologies.

Oocyte donation program: pregnancy and implantation rates in women of different ages sharing oocytes from single donor*

OBJECTIVE To investigate the importance of uterus age as regards of pregnancy, implantation, and abortion rates using the oocyte donation model. DESIGN Retrospective data analysis of cases where recipients of different ages shared oocytes from single donor. SETTING A tertiary infertility center. PATIENTS One hundred fourteen women (21 to 49 years of age) undergoing a total of 114 cycles of oocyte donation were divided into two groups according to age (group A < = or 39 years: group B between 40 and 49 years). INTERVENTIONS Hormonal replacement therapy was given using increasing doses of 17 beta-E2 (2, 4, and 6 or 8 mg) and either 100 mg of P in oil or 600 mg of micronized P through the vaginal route. MAIN OUTCOME MEASURES Pregnancy, abortion, and implantation rates. RESULTS Fifty-seven transfer cycles were performed per age group. Twenty-seven clinical pregnancies were achieved in Group A and 14 in group B, with pregnancy rates (PRs) of 47.3% and 24.5%, respectively. There were four abortions in group A and one in group B, resulting in abortion rates of 14.8% and 7%, respectively. Thirty-four of 137 transferred embryos in group A and 20 of 134 in group B implanted, resulting in implantation rates of 24.8% and 14.9%, respectively. CONCLUSION This study seems to suggest that there are differences in pregnancy and implantation rates in recipients of different ages because of uterine receptivity. Fertility therefore does not depend merely on oocyte age and quality but also on uterine age.