Francesco Fusi

Myo-inositol in patients with polycystic ovary syndrome: A novel method for ovulation induction

Background. Polycystic ovary syndrome (PCOS) is often characterized by chronic oligo- or anovulation (usually manifested as oligo- or amenorrhea), and hyperandrogenism. In addition, 30–40% of PCOS women have impaired glucose tolerance, and a defect in the insulin signaling pathway (inositol-containing phosphoglycan mediators) seems to be implicated in the pathogenesis of insulin resistance. PCOS patients are subfertile as a consequence of such ovulatory disorders and often need drugs, such as clomiphene citrate or follicle-stimulating hormone, for ovulation induction, which increases the risk of multiple pregnancy and ovarian hyperstimulation syndrome. We hypothesized that the administration of an isoform of inositol (myo-inositol), belonging to the vitamin B complex, would improve the insulin-receptor activity, restoring normal ovulatory function. Materials and methods. Twenty-five PCOS women of childbearing age with oligo- or amenorrhea were enrolled in the study. Ovulatory disorder due to PCOS was apparently the only cause of infertility; no tubal defect or deficiency of male semen parameters was found. Myo-inositol combined with folic acid (Inofolic®) 2 g twice a day was administered continuously. During an observation period of 6 months, ovulatory activity was monitored with ultrasound scan and hormonal profile, and the numbers of spontaneous menstrual cycles and eventually pregnancies were assessed. Results. Twenty-two out of the 25 (88%) patients restored at least one spontaneous menstrual cycle during treatment, of whom 18 (72%) maintained normal ovulatory activity during the follow-up period. A total of 10 singleton pregnancies (40% of patients) were obtained. Nine clinical pregnancies were assessed with fetal heart beat at ultrasound scan. Two pregnancies evolved in spontaneous abortion. Conclusion. Myo-inositol is a simple and safe treatment that is capable of restoring spontaneous ovarian activity and consequently fertility in most patients with PCOS. This therapy did not cause multiple pregnancy.

Myo-inositol may improve oocyte quality in intracytoplasmic sperm injection cycles. A prospective, controlled, randomized trial.

OBJECTIVE To determine the effects of myo-inositol on oocyte quality in polycystic ovary syndrome (PCOS) patients undergoing intracytoplasmic sperm injection (ICSI) cycles. DESIGN A prospective, controlled, randomized trial. SETTING Assisted reproduction centers. PATIENT(S) Sixty infertile PCO patients undergoing ovulation induction for ICSI. INTERVENTION(S) All participants underwent standard long protocol. Starting on the day of GnRH administration, 30 participants received myo-inositol combined with folic acid (Inofolic) 2 g twice a day and 30 control women received folic acid alone, administrated continuously. MAIN OUTCOME MEASURE(S) Primary end points were number of morphologically mature oocytes retrieved, embryo quality, and pregnancy and implantation rates. Secondary end points were total number of days of FSH stimulation, total dose of gonadotropin administered, E(2) level on the day of hCG administration, fertilization rate per number of retrieved oocytes, embryo cleavage rate, live birth and miscarriage rates, cancellation rate, and incidence of moderate or severe ovarian hyperstimulation syndrome. RESULT(S) Total r-FSH units (1,958 +/- 695 vs. 2,383 +/- 578) and number of days of stimulation (11.4 +/- 0.9 vs. 12.4 +/- 1.4) were significantly reduced in the myo-inositol group. Furthermore, peak E(2) levels (2,232 +/- 510 vs. 2,713 +/- 595 pg/mL) at hCG administration were significantly lower in patients receiving myo-inositol. The mean number of oocytes retrieved did not differ in the two groups, whereas in the group cotreated with myo-inositol the mean number of germinal vesicles and degenerated oocytes was significantly reduced (1.0 +/- 0.9 vs. 1.6 +/- 1.0), with a trend for increased percentage of oocytes in metaphase II (0.82 +/- 0.11% vs. 0.75 +/- 0.15%). CONCLUSION(S) These data show that in patients with PCOS, treatment with myo-inositol and folic acid, but not folic acid alone, reduces germinal vesicles and degenerated oocytes at ovum pick-up without compromising total number of retrieved oocytes. This approach, reducing E(2) levels at hGC administration, could be adopted to decrease the risk of hyperstimulation in such patients.

Oocyte cryopreservation: clinical outcome of slow-cooling protocols differing in sucrose concentration

Oocyte cryopreservation represents an important option for management of female fertility, avoiding the ethical concerns associated with embryo storage. This retrospective study evaluated the clinical outcome of two alternative slow freezing protocols involving different sucrose concentrations. From January 2004 to March 2006, spare oocytes from selected couples undergoing IVF or intracytoplasmic sperm injection were frozen using a slow-cooling protocol and thawed at a later stage. Patients were divided into two groups: group A (n = 65), whose oocytes were frozen with propane-1,2-diol (PrOH) and 0.1 mol/l sucrose; and group B (n = 66) whose oocytes were frozen with 0.3 mol/l sucrose. A total of 543 oocytes were thawed in group A and 601 in group B, achieving a survival rate of 24.3 and 71.2% respectively. Whilst fertilization rate (53.5 and 80.4% respectively) was higher in group B, enhanced results for group A were achieved over all (implantation rate per transferred embryos 12.2 versus 5.7%; pregnancy rate per transfer 16.7 versus 9.5%). Normal births and ongoing pregnancies have occurred in both groups. Although in slow-cooling methods higher sucrose concentration in the freezing mixture allows higher post-thaw survival and fertilization rates, overall this did not coincide with an improved clinical outcome.

Pathologic findings in hysteroscopy before in vitro fertilization-embryo transfer (IVF-ET)

Background. The aim of this study was to evaluate hysteroscopy routinely performed prior to in vitro fertilization-embryo transfer (IVF-ET). Methods. We analyzed in a prospective study 300 patients who underwent hysteroscopy before the first IVF-ET cycle. We analyzed then in a retrospective manner 300 patients who did not perform hysteroscopy. Results. One-hundred-and-eighty (60%) hysteroscopies were normal but 120 (40%) revealed an unsuspected intrauterine abnormality. We did not find statistically significant differences between patients with normal or abnormal hysteroscopy in any characteristic. We found a statistically significant difference in pregnancy rate between women who performed hysteroscopy before IVF-ET cycle and in women who did not perform it. Conclusions. Hysteroscopy, as a routine examination, should be performed before the first IVF-ET cycle in all patients.

Objective evaluation of the viability of cryopreserved oocytes

Recent studies of fundamental cryobiology, empirical observations and more systematic clinical experiences have generated a renewed interest in oocyte cryopreservation. Poor survival rate has long been the limiting factor which has prevented widespread adoption of oocyte storage. Slow-cooling and vitrification protocols developed in the last few years have apparently solved this problem, ensuring high recovery of viable oocytes from liquid nitrogen storage. However, the definition of oocyte viability appears rather vague. In fact, post-storage survival as assessed on morphological criteria, indicated by the absence of overt cell degeneration, is not necessarily synonymous with viability. Despite its sensitivity to low temperatures, the meiotic spindle can be preserved after cryopreservation and its constitution after thawing can be monitored non-invasively through polarized light microscopy. Assessment of oocyte cryopreservation via clinical parameters is a daunting task. Most studies are small and difficult to interpret because of confounding factors, such as age, patient selection and quality and strategy of use of the cryopreserved material. Some progress has been made, however, as suggested by recent experiences in which the implantation efficiency of embryos produced from thawed oocytes approaches that reported using cryopreserved embryos directly.

Epigenetic analysis of the critical region I for premature ovarian failure: demonstration of a highly heterochromatic domain on the long arm of the mammalian X chromosome

Background: X chromosome rearrangements defined a critical region for premature ovarian failure (POF) that extended for >15 Mb in Xq. It has been shown previously that the region could be divided into two functionally distinct portions and suggested that balanced translocations interrupting its proximal part, critical region 1 (CR1), could be responsible for POF through downregulation of ovary expressed autosomal genes translocated to the X chromosome. Results and conclusion: This study reports that such position effect can indeed be demonstrated by analysis of breakpoint regions in somatic cells of POF patients and by the finding that CR1 has a highly heterochromatic organisation, very different from that of the euchromatic autosomal regions involved in the rearrangements. The chromatin organisation of the POF CR1 is likely to be responsible for the epigenetic modifications observed in POF patients. The characteristics of CR1 and its downregulation in oocytes may very well explain its role in POF and the frequency of the POF phenotype in chromosomal rearrangements involving Xq. This study also demonstrates a large and evolutionary conserved domain of the long arm of the X chromosome, largely corresponding to CR1, that may have structural or functional roles, in oocyte maturation or in X chromosome inactivation.

Natural cycle as first approach in aged patients with elevated follicle-stimulating hormone undergoing intracytoplasmic sperm injection: a pilot study.

Background. Poor ovarian response to standard in vitro fertilization–embryo transfer (IVF-ET) protocols or different regimens of treatment, as consequence of a diminished ovarian reserve, correlates strictly with patient age, elevated follicle-stimulating hormone (FSH) and reduced antral follicle count. The aim of the present pilot study was to evaluate the outcome of patients with poor prognostic features undergoing IVF-ET with natural cycles as a first approach and not as a consequence of a previous failure treatment. Materials and methods. Eighteen aged patients (mean ± standard deviation 40.2 ± 0.7 years, range 37–43 years) with elevated serum FSH and reduced antral follicle count underwent intracytoplasmic sperm injection (ICSI) after spontaneous ovulation. Results. A total of 26 natural cycles with ICSI were analyzed. Pregnancy was observed in three patients, of which two were ongoing as assessed by fetal heart beat at ultrasound scan performed 4–5 weeks after ET. Conclusion. The overall pregnancy rates achieved (11.5% per cycle, 20.0% per ET) are comparable with those of conventional IVF-ET in aged patients, and not impaired by a single embryo transferred. Better embryo quality, as a consequence of natural selection of oocytes, better endometrium receptivity and monthly repeatability of the procedure, can balance the relatively low chance to perform ET.

Production of Mouse Monoclonal Antibodies Directed Against the Oolemma of Human and Hamster Oocytes by Intra‐splenic Injection of Oocytes

PROBLEM: To develop an additional approach for the study of oolemmal surface moieties involved in gamete interactions, we decided to obtain monoclonal antibodies by intrasplenic injection of human and hamster oocytes in Balb/c mice.

Open epididymal sperm aspiration (OESA): minimally invasive surgical technique for sperm retrieval.

OBJECTIVES Male infertility caused by irreparable obstructive azoospermia is widely treated with MESA (Microsurgical Epididymal Sperm Aspiration) or TESE (Testicular Sperm Extraction) or PESA (Percutaneous Epididymal Sperm Aspiration) followed by an Intracytoplasmatic Spermatozoa Injection (ICSI). For each procedures are claimed advantages and disadvantages. Aim of this work is to describe our technique (OESA) for sperm retrieval reporting the results. METHODS From 1998 to 2003 153 sperm retrieval procedures in azoospermic patients were performed. The technique consists in a small (1 cm.) surgical incision of scrotal wall under local or spinal anaesthesia. After inspecting the intrascrotal component to record the status of the epididymis and of the testicle, epididymal sperm aspiration was performed in the most appropriate site with 18 gauge needle. The aspirated spermatozoa were used immediately after aspiration not being permitted in our Institute cryopreservation. When no sperms were found TESE was performed. RESULTS In 78 cases aspiration from epididymis was successful (OESA). In the other patients (75) TESE was performed. Higher count of spermatozoa were retrieved in all cases of OESA. Pregnancy rate was 26.6% for OESA and 6.4% for TESE. No complications occurred. CONCLUSIONS This technique is very simple and easy allowing to obtain the advantages of microsurgical techniques (mainly choice of the most appropriate site of aspiration and immediate haemostasis if needed) with minimal invasivity (avoiding epididymal surgical incision). Moreover if OESA does not allow sperm extraction is possible to perform immediately TESE.