Inositols: From Established Knowledge to Novel Approaches.

Myo-inositol (myo-Ins) and D-chiro-inositol (D-chiro-Ins) are natural compounds involved in many biological pathways. Since the discovery of their involvement in endocrine signal transduction, myo-Ins and D-chiro-Ins supplementation has contributed to clinical approaches in ameliorating many gynecological and endocrinological diseases. Currently both myo-Ins and D-chiro-Ins are well-tolerated, effective alternative candidates to the classical insulin sensitizers, and are useful treatments in preventing and treating metabolic and reproductive disorders such as polycystic ovary syndrome (PCOS), gestational diabetes mellitus (GDM), and male fertility disturbances, like sperm abnormalities. Moreover, besides metabolic activity, myo-Ins and D-chiro-Ins deeply influence steroidogenesis, regulating the pools of androgens and estrogens, likely in opposite ways. Given the complexity of inositol-related mechanisms of action, many of their beneficial effects are still under scrutiny. Therefore, continuing research aims to discover new emerging roles and mechanisms that can allow clinicians to tailor inositol therapy and to use it in other medical areas, hitherto unexplored. The present paper outlines the established evidence on inositols and updates on recent research, namely concerning D-chiro-Ins involvement into steroidogenesis. In particular, D-chiro-Ins mediates insulin-induced testosterone biosynthesis from ovarian thecal cells and directly affects synthesis of estrogens by modulating the expression of the aromatase enzyme. Ovaries, as well as other organs and tissues, are characterized by a specific ratio of myo-Ins to D-chiro-Ins, which ensures their healthy state and proper functionality. Altered inositol ratios may account for pathological conditions, causing an imbalance in sex hormones. Such situations usually occur in association with medical conditions, such as PCOS, or as a consequence of some pharmacological treatments. Based on the physiological role of inositols and the pathological implications of altered myo-Ins to D-chiro-Ins ratios, inositol therapy may be designed with two different aims: (1) restoring the inositol physiological ratio; (2) altering the ratio in a controlled way to achieve specific effects.

Breakthroughs in the Use of Inositols for Assisted Reproductive Treatment (ART).

It is well known that myo-inositol (MI) and D-chiro-inositol (DCI) are insulin-sensitizing agents, and MI is of proven utility in polycystic ovary syndrome (PCOS). In addition, MI plays a pivotal role in the physiology of reproduction, and has beneficial effects on the development of oocytes, spermatozoa, and embryos. By contrast, DCI has little effect on spermatozoa, but high concentrations in the ovary can negatively affect the quality of oocytes and the blastocyst. Overall, the evidence in the literature supports the beneficial effects of MI in both female and male reproduction, warranting clinical use of MI in assisted reproductive treatment (ART).

Single layer suturing in intracapsular myomectomy of intramural myomas is sufficient for a normal wound healing.

STUDY OBJECTIVE: To evaluate surgical outcomes of intracapsular single-layer myomectomy in terms of efficacy and safety as well as examine potential alterations based on kind of surgical approach. METHODS: A prospective observational study was performed between January 2010 and December 2018. Women in reproductive age, affected by intramural or subserous myomas (FIGO type 3-6) of 4-14 cm diameter were enrolled. Primary outcomes included initial and final uterine incision length, time to wound healing and uterine rupture in subsequent pregnancies. Furthermore, a sub-analysis was also performed regarding surgical approach, namely laparoscopical or laparoscopically-assisted myomectomy, in order to confirm whether overall observations are similar for both potential surgical approaches. RESULTS: There were finally 273 patients included in the present study. Overall mean uterine incision was initially 3.1 cm and was shortened to 2.2 cm at the end of operation, indicating a reduction of 29.1 %. Mean estimated blood loss was 154.2 mL and mean operative time was 82.1 min. No severe intraoperative and postoperative complications were presented. 121 of the studied women had pregnancy 3-36 months after myomectomy, without reporting any uterine rupture. When comparing LIM vs. LAIM, all outcomes were also favorable in the total of patients. CONCLUSION: Intracapsular myomectomy either by LIM or LAIM is a safe and attractive alternative to abdominal myomectomy in setting of premenopausal patients with myomas up to 14 cm. A single-layer continuous suturing in intracapsular myomectomies is enough for a successful wound healing.

Experts' opinion on inositols in treating polycystic ovary syndrome and non-insulin dependent diabetes mellitus: a further help for human reproduction and beyond.

Introduction: This Experts' opinion provides an updated scientific support to gynecologists, obstetricians, endocrinologists, nutritionists, neurologists and general practitioners on the use of Inositols in the therapy of Polycystic Ovary Syndrome (PCOS) and non-insulin dependent (type 2) diabetes mellitus (NIDDM).Areas covered: This paper summarizes the physiology of Myo-Inositol (MI) and D-Chiro-Inositol (DCI), two important molecules present in human organisms, and their therapeutic role, also for treating infertility. Some deep differences between the physiological functions of MI and DCI, as well as their safety and intestinal absorption are discussed. Updates include new evidence on the efficacy exerted in PCOS by the 40:1 MI/DCI ratio, and the innovative approach based on alpha-lactalbumin to overcome the decreased therapeutic efficacy of Inositols in some patients.Expert opinion: The evidence suggests that MI, alone or with DCI in the 40:1 ratio, offers a promising treatment for PCOS and NIDDM. However, additional studies need to evaluate some still unresolved issues, such as the best MI/DCI ratio for treating NIDDM, the potential cost-effectiveness of reduced gonadotropins administration in IVF due to MI treatment, or the benefit of MI supplementation in ovulation induction with clomiphene citrate in PCOS patients.

Spindle and chromosome configuration analysis of human biopsied versus non-biopsied embryos by confocal laser scanning microscopy following vitrification.

SummaryThe aim of this study was to investigate the effects of zona drilling and biopsy on day 3 followed by vitrification on day 5 on the cytoskeleton and development of human embryos, by analysing survival rates and spindle and chromosome configurations by fluorescence and confocal laser scanning microscopy in human biopsied and non-biopsied embryos. In total, 98 human blastocysts (50 non-biopsied and 48 following biopsy on day 3) were vitrified on day 5 using either a commercial dimethyl sulphoxide (DMSO)-free vitrification kit or increasing concentrations of DMSO/EG (5%/5-10%/10-20%/20%). Following warming, the blastocysts were allowed to recover in culture for 24 h and were immunostained with α-tubulin, acetylated tubulin, and/or γ-tubulin antibodies in combination with 4',6-diamidino-2-phenylindole (DAPI). Labelled embryos were examined by both fluorescence and confocal laser scanning microscopy. The survival rates following warming (92% non-biopsied vs 83.3% biopsied) and the incidence of normal spindle chromosome configurations was not statistically different between the two groups (65.2% non-biopsied vs 59.2% biopsied, P>0.05). The incidence of spindle abnormalities including multipolarity, chromosome lagging, congression failure and chromosome bridging were also similar between the two groups (P>0.05). This study is the first to compare the incidence of cytoskeletal abnormalities in biopsied and non-biopsied human embryos following vitrification. We conclude that there was no significant difference in the survival rates and the incidence of spindle abnormalities between the two groups.

GnRH antagonist administered twice the day before hCG trigger combined with a step-down protocol may prevent OHSS in IVF/ICSI antagonist cycles at risk for OHSS without affecting the reproductive outcomes: a prospective randomized control trial.

PURPOSE: The purpose this study is to investigate whether a double antagonist dose (0.25 mg/12 h) administered the day before hCG trigger is effective in preventing ovarian hyperstimulation syndrome (OHSS) in GnRH antagonist IVF/intracytoplasmic sperm injection (ICSI) cycles at risk for OHSS. METHODS: This is a prospective randomized control study, conducted from November 2012 to January 2016. A total of 194 patients undergoing a IVF/ICSI GnRH antagonist cycle that were at risk of OHSS and chose to proceed with embryo transfer and avoid cycle cancellation or embryo cryopreservation were allocated into two groups. The inclusion criteria consisted of a rapid rise of oestradiol ≥ 3500 pg/ml combined with ≥ 18 follicles > 11 mm in diameter without any mature follicle > 16 mm, in any day of stimulation. Overall, 97 patients (intervention group A) received a double dose of GnRH antagonist (0.25 mg/12 h) the day before hCG while 97 patients (control group B) did not. Recombinant FSH administration was tapered to 100 IU/24 h the day of the allocation in both groups. RESULTS: Incidence of early-onset moderate/severe OHSS was significantly lower in intervention group A compared to control group B (0 vs 12.37%, P < 0.001). Clinical pregnancy rate per cycle (50.52 vs 42.27%, P = 0.249) was not significantly different between the two groups. Oestradiol (3263.471 ± 1271.53 vs 5233 ± 1425.17, P < 0.001), progesterone (0.93 ± 0.12 vs 1.29 ± 0.14, P < 0.001) and luteinizing hormone (1.42 ± 0.31 vs 1.91 ± 0.33, P < 0.001) were significantly lower in group A the day of the hCG triggering. CONCLUSION: The administration of a rescue double GnRH antagonist dose the day before hCG trigger may represent a safe alternative preventive strategy for early OHSS without affecting the reproductive outcomes. TRIAL REGISTRATION NUMBER: ISRCTN02750360.

GnRH antagonist versus long GnRH agonist protocol in poor IVF responders: a randomized clinical trial.

OBJECTIVE: To compare the efficacy of the long GnRH agonist and the fixed GnRH antagonist protocols in IVF poor responders. STUDY DESIGN: This was a randomized controlled trial performed in the Iakentro IVF centre, Thessaloniki, from January 2007 to December 2011, concerning women characterised as poor responders after having 0-4 oocytes retrieved at a previous IVF cycle. They were assigned at random, using sealed envelopes, to either a long GnRH agonist protocol (group I) or a GnRH antagonist protocol (group II). RESULTS: Overall 364 women fulfilled the inclusion criteria and were allocated to the two groups: finally 330 participated in our trial. Of these, 162 were treated with the long GnRH agonist protocol (group I), and 168 with the fixed GnRH antagonist protocol (group II). Numbers of embryos transferred and implantation rates were similar between the two groups (P=NS). The overall cancellation rate was higher in the antagonist group compared to the agonist group, but the difference was not significant (22.15% vs. 15.2%, P=NS). Although clinical pregnancy rates per transfer cycle were not different between the two groups (42.3% vs. 33.1%, P=NS), the clinical pregnancy rate per cycle initiated was significantly higher in the agonist compared to the antagonist group (35.8% vs. 25.6%, P=0.03). CONCLUSIONS: Although long GnRH agonist and fixed GnRH antagonist protocols seem to have comparable pregnancy rates per transfer in poor responders undergoing IVF, the higher cancellation rate observed in the antagonist group suggests the long GnRH agonist protocol as the first choice for ovarian stimulation in these patients.

Improved recovery using multimodal perioperative analgesia in minimally invasive myomectomy: a randomised study.

OBJECTIVE: To evaluate postoperative pain using a multimodal analgesic protocol in women with uterine fibroids managed with minimally invasive myomectomy. MATERIALS AND METHODS: A prospective randomised trial was designed to evaluate the postoperative pain of women treated with minimally invasive myomectomy, using a multimodal analgesic protocol (consisting of perioperative pharmaceutical agents of local and systemic action adjuvant to the classic anaesthesia protocol). Ninety-five premenopausal women were assessed for minimally invasive myomectomy (laparoscopic myomectomy and laparoscopically assisted myomectomy). Ninety-two women were included in the final analysis and were randomly allocated in two groups using sealed envelopes: group I (n=47) consisted of women who received the multimodal analgesic protocol and was compared with group II (n=45) who did not receive the protocol. The main outcome measure was the postoperative pain score at 2 and 8 h after surgery, according to the Visual Analog Scale (VAS). Additionally, time for bowel peristalsis return, duration of hospitalisation and full recuperation to normal activity were also measured. RESULTS: Significantly lower VAS scores for postoperative pain, earlier return of bowel peristalsis and fewer hours of hospitalisation were observed in the group in which multimodal analgesia was used. The days for the full recuperation to normal activity were similar between the two groups. CONCLUSION: In the setting of minimally invasive myomectomy, the use of a multimodal analgesic protocol improved postoperative recovery, resulting in earlier hospital discharge.

Double GnRH-antagonist dose before HCG administration may prevent OHSS in oocyte-donor cycles: a pilot study.

This pilot study evaluated the possibility of preventing early ovarian hyperstimulation syndrome (OHSS) by increasing the daily dose of gonadotrophin-releasing hormone (GnRH) antagonist administration (to twice a day) in oocyte-donor cycles stimulated with the antagonist protocol. The study included 72 oocyte donors who underwent ovarian stimulation using the GnRH antagonist protocol and might have had their cycle cancelled because of ovarian hyper-response. All women were donors presenting a rapid rise of oestradiol > or = 3000 pg/ml early in the stimulation period with more than 15 follicles of < or = 15 mm in diameter. By decreasing the rFSH dose to 75 IU a day with an additional daily dose of GnRH antagonist (0.25 mg twice a day), the oestradiol concentrations were lowered or reached a plateau before human chorionic gonadotrophin was given. A marked decrease in oestradiol concentrations and ovarian volume was observed on the day of oocyte retrieval and 3 days post retrieval. None of the donors needed coasting, were cancelled or developed OHSS. In over-responding oocyte donors, by increasing the usual GnRH-antagonist dose to twice a day during ovarian stimulation, the oestradiol rise can be blocked while a minimal follicular stimulation may continue without the risk of developing OHSS or affecting the outcome.

Laparoscopically assisted myomectomy versus abdominal myomectomy in short-term outcomes: a prospective study.

PURPOSE: Comparison between laparoscopically assisted myomectomy (LAM) and abdominal myomectomy (laparotomy), used in the management of women with intramural or subserous uterine fibroids up to 90 mm of maximum diameter. METHODS: Seventy-five premenopausal women were prospectively enrolled in the study, managed by LAM (n=48) or by laparotomy (n=27) approach. The short-term outcomes were compared between the two groups. The patient characteristics were also analyzed. RESULTS: The mean (+/-SD) estimated blood loss was significantly less in the LAM procedure compared with laparotomy (246+/-161 vs. 351+/-219 ml, respectively, P=0.03). Similarly, the operative time was shorter in the LAM modality compared with laparotomy (68+/-21 vs. 83+/-24 min, respectively, P=0.01). Intraoperative and postoperative complications were not different between the two groups. The mean days of the bowel reactivity (1.04+/-0.2) was faster (P<0.0001), while the duration of hospitalization (1.2+/-0.6) was shorter (P<0.0001) in the LAM technique, when compared with abdominal myomectomy (1.8+/-0.5 and 4.2+/-0.8, respectively). CONCLUSIONS: In selected group of patients, LAM as minimally invasive approach is an attractive alternative to conventional laparotomic myomectomy, offering significant advantages.

GnRH antagonists and endometrial receptivity in oocyte recipients: a prospective randomized trial.

The effect that gonadotrophin-releasing hormone (GnRH) antagonists exert on endometrial receptivity has not yet been elucidated. GnRH antagonists might directly affect oocytes, the embryo and/or the endometrium. The aim of this study was to investigate the direct effect of GnRH antagonists on the endometrium in oocyte donation cycles. In an oocyte donation programme, oocytes from each donor (n = 49), stimulated with gonadotrophins and a GnRH antagonist, were equally shared between two different matched recipients. Recipients were randomly allocated to either receive a GnRH antagonist concomitant to donor during their endometrial priming with oestradiol (group I, n = 49) or to solely continue with their endometrial preparation (group II, n = 49). Pregnancy rate was 55.1% in group I and 59.1% in group II. Implantation rate was 26.1% in group I and 24.4% in group II. Endometrial thickness was also similar between the two groups on the day of human chorionic gonadotrophin injection to the donor. In conclusion, GnRH antagonist administration during the proliferative phase at a dose of 0.25 mg per day does not appear to adversely affect endometrial receptivity in oocyte recipients.

Low-dose human chorionic gonadotropin during the proliferative phase may adversely affect endometrial receptivity in oocyte recipients.

The effect of low-dose human chorionic gonadotropin (hCG) administration in the proliferative phase of oocyte recipients was investigated in a prospective randomized trial. Sibling oocytes from the same donor were shared at random among two different recipients. In group I oocyte recipients received 750 IU of hCG every three days concomitant to endometrial preparation with estradiol until hCG injection to the donor, whereas in group II recipients received no hCG during endometrial priming with estradiol. Endometrial thickness was significantly lower in group I compared with group II, although similar endometrial thickness was detected during the mock cycle. Pregnancy rates were significantly lower in group I than in group II (13.6% vs. 45.4%, p<0.05). Implantation rates were also significantly lower in group I (1.7% vs. 22.4%, p<0.01). The study was discontinued prematurely for ethical reasons when 22 cycles were completed, as pregnancy rates were very low in group I. In conclusion, hCG administration in the proliferative phase might directly affect endometrial proliferation and receptivity.

Laparoscopy vs laparoscopically assisted myomectomy in the management of uterine myomas: a prospective study.

OBJECTIVE: This study was undertaken to compare the intraoperative and short-term outcomes between the 2 modalities of minimally invasive surgery for the management of uterine fibroids. STUDY DESIGN: A total of 116 patients with inclusion criteria were prospectively collected in a study period from March 1997 through 2007. Laparoscopic (n = 40) vs laparoscopically assisted myomectomy (n = 76) were compared for the management of no more than 3 intramural or subserous uterine myomas, of a maximum diameter of 90 mm. RESULTS: The patients' characteristics by age, parity, body mass index, number and location of myomas were well balanced between the 2 study groups. The mean diameter of the myomas was the only characteristic significantly higher in the laparoscopically assisted myomectomy group. The operative time in the laparoscopically assisted myomectomy was significantly shorter compared with the laparoscopic myomectomy (mean +/- standard deviation: 66 +/- 19 minutes vs 94 +/- 18 minutes, P < .0001). A shorter uterine incision was found in the laparoscopically assisted myomectomy technique compared with the laparoscopic myomectomy (2.9 +/- 0.6 vs 4.3 +/- 1.2, P < .0001). Estimated blood loss was significantly higher in the laparoscopically assisted myomectomy group (P = .002). Intraoperative, early postoperative complications, hospitalization days, and fully returned activity were similar between the 2 study groups. CONCLUSION: The present data suggest that the laparoscopically assisted myomectomy is a valid alternative to laparoscopy in a setting of minimally invasive surgery for the management of uterine fibroids.

Spindle abnormalities in normally developing and arrested human preimplantation embryos in vitro identified by confocal laser scanning microscopy.

BACKGROUND: Despite recent technical improvements, many human preimplantation embryos fail to develop to the blastocyst stage or implant after transfer to the uterus. A possible cause for this developmental arrest is the high incidence of nuclear and postzygotic chromosomal abnormalities observed during cleavage, including chaotic chromosome complements, suggestive of defects in mitotic chromosomal segregation. The underlying mechanisms are largely unknown, but similarities with chromosome instability in human cancers led to the proposal that cell cycle checkpoints may not operate at these early stages. METHODS: To investigate this and to examine whether spindle abnormalities contribute to chromosome malsegregation, we have used fluorescence and confocal laser scanning microscopy, following immunolabelling with antibodies specific for alpha-tubulin, gamma-tubulin, or acetylated tubulin, combined with a DNA fluorochrome to visualize nuclei, spindle and chromosome configurations in normal and arrested human embryos, from cleavage to blastocyst stages. RESULTS: In addition to frequent interphase nuclear abnormalities, we identify for the first time various spindle abnormalities including abnormal shape and chromosome loss and multipolar spindles at cleavage and blastocyst stages. CONCLUSIONS: We propose that a major pathway leading to postzygotic chromosomal abnormalities is the formation of binucleate blastomeres with two centrosomes which result either in a bipolar spindle and division to two tetraploid blastomeres, or in a multipolar spindle, chromosome malsegregation and chromosomal chaos.

Comparison of effects of zona drilling by non-contact infrared laser or acid Tyrode's on the development of human biopsied embryos as revealed by blastomere viability, cytoskeletal analysis and molecular cytogenetics.

Use of a non-contact infrared laser (IRL) or acid Tyrode's for zona drilling before embryo biopsy was compared by assessing blastomere viability using various fluorescent markers or culture of the single biopsied blastomere, and, by cytoskeletal and molecular cytogenetic analysis of the biopsied embryos following culture to the blastocyst stage. There was no significant difference in the proportion of biopsied embryos that showed no damage in both the biopsied blastomere and in the remaining embryo (acid Tyrode's: 75% versus IRL: 68%), or in the proportion of single biopsied blastomeres that divided in culture (P > 0.05). However, single biopsied blastomeres from laser drilled embryos showed a greater tendency to form miniblastocysts. The proportion of laser or acid Tyrode's biopsied embryos that reached the blastocyst stage by day 6 was similar, although evident earlier (day 5) in the laser biopsied embryos. Spindle abnormalities at the blastocyst stage included tripolar and tetrapolar spindles, but their incidence was not significantly different from controls. In addition, no significant difference was observed in the incidence of chromosomal abnormalities and mosaicism between the two groups. It is concluded that using an IRL at a safe working distance does not cause adverse immediate or longer term effects on the development of human biopsied embryos, although damage can occur if drilling within this distance is unavoidable. Acid Tyrode's drilling can also cause damage, and tended to retard blastocyst development.