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It is well defined that ovarian steroids play a role in the cardiovascular system, acting as vasoactive compounds. The aim of this study is to assess the endothelial function during the menstrual cycle using flow-mediated dilation of the brachial artery.A total of 21 healthy premenopausal women, with regular menstrual cycles, were included in this observational, longitudinal, and prospective study. The endothelium function was assessed by ultrasound during four phases of the menstrual cycle: early follicular phase (EFP), late follicular phase (LFP), early luteal phase (ELP) and late luteal phase (LLP).We observed a significant variation among the vasodilatation response during the menstrual cycle phases (P0.001). The result was higher during LFP than during ELP (P0.001) or LLP (P0.001). Late luteal phase had higher response than observed during ELP (P=0.003) and EFP was higher than LLP (P=0.003). There were no significant differences between the results observed during EFP and LFP (P=1.0), or EFP and ELP (P=0.137).Our results suggest that the ovarian steroids may play an important role in modulating endothelial function.

The objective of our study was to compare the osmolality in sequential and single step culture media, used for in vitro human embryo culture, covered with mineral oil and paraffin, in dry and humid incubators.We performed a prospective observational study. A total of 120 Petri dishes, with 960 droplets of culture media, were evaluated. Each dish was prepared with 4 droplets of single step medium and sequential medium. Sixty dishes were covered with mineral oil and 60 with paraffin oil. Half were incubated in a dry incubator and half in a humid. Osmolality was measured on days 1, 3, 5, 7. ANOVA test was performed for statistical analysis.Osmolality results for single step and sequential medium, that were covered with both mineral and paraffin oil and placed in the dry incubator, significantly increased throughout the study time (D7D5D3). In the humid incubator, the results were similar for all periods. Osmolality was significantly lower in humid incubator, in all periods, when droplets were covered with both oils. When both culture media were placed in the humid incubator, no variation was detected, using both oils. However, when single step medium was placed in the dry incubator, covered with mineral oil, we observed a higher osmolality than the covered with paraffin oil.TWe can conclude that humid incubator is better for maintaining osmolality and paraffin oil protect single step media from evaporation in dry incubator.

Objective Lyophilization is potentially more practical and cost-effective alternative for sperm preservation. However, there are no studies that evaluate the ultrastructure of human spermatozoa after lyophilization. Therefore, the aim of our study was to evaluate the ultrasctructure of lyophilized spermatozoa using Transmission Electron Microscopy. Methods From a total of 21 donated seminal samples, 30 aliquots were originated and divided into two aliquots so that one could have been submitted to cryopreservation/thaw and the other for lyophilization/rehydration. The liquefied aliquots were homogenized at room temperature. Samples assigned for cryopreservation were placed in straws and samples assigned for lyophilization were placed in the appropriate vials. Cryopreservation samples were placed at -30oC for 30 minutes subsequently for 30 minutes at vapour phase and then plunged into liquid nitrogen. Lately, were warmed in water bath at 37oC for 10 minutes followed by 10 minutes centrifugation. The pellet was resuspended and analysed in a Makler chamber. The semen vials assigned for lyophilization were loaded into a pre-fixed freeze-drying chamber. Following lyophilization, vials were removed from the freeze-drying chamber and kept at 4oC until rehydration. TEM was performed after rehydration and thawing. Sperm samples were fixed, rinsed in buffer, post fixed and dehydration was carried out in escalating concentrations of alcohol solution, acetone and then, embedding in Epon resin. Ultrathin sections were stained and examined in a Transmission Electron Microscope. Results Analysis of sperm after freezing/thawing using Transmission Electron Microscopy showed lesions to the midpiece, with some mitochondria degeneration and random rupture of plasma membrane. In the head, we identified intact plasma membrane, nucleus and acrosome, as in the flagellum all main structures remained intact including the plasma membrane, the longitudinal columns of dense fibers and the semicircular fibers. Analysis by Transmission Electron Microscopy showed that spermatozoa heads had ruptured plasma membranes, absence of acrosomes, nuclei with heterogeneous and decompressed chromatin. Mitochondria were deteriorated in the midpiece. Longitudinal columns of dense fibers were absent in the flagellum. Axonemes, in cross-sections, were disrupted with disorganized structures. Conclusions To our knowledge, our study demonstrated, for the first time, the structure of the human spermatozoa after lyophilization using Transmission Electron Microscopy. The use of a fixed lyophilization protocol with media containing cryoprotectants might explain the damage to the structures. More studies are necessary to improve the results of sperm lyophilization. In the future, the use of lyophilization of spermatozoa might reduce the costs of fertility preservation, since there will be no need for storage space and transportation is simpler.

Objective To evaluate COS and oocyte retrieval results in ART treatment cycles initiated at any stage of the menstrual cycle (random start) in cancer patients, who could not postpone the onset of cancer treatment. Methods Prospective observational study of 26 women with cancer, with an indication to start cancer treatment within the next 20 days and wishing to preserve their fertility. Ovarian stimulation started immediately with FSH followed by GnRH antagonist for pituitary suppression and GnRH agonist for oocyte maturation. Treatment started from day 1 to day 14 of the menstrual cycle was considered to be in the follicular phase, and that started from day 15 to day 28 was considered to be in the luteal phase. Oocyte retrieval was performed 34 h after GnRH agonist administration. After identification and maturity classification, metaphase II oocytes were cryopreserved using vitrification. Results A total of 13 women had breast cancer, 4 ovarian cancer, 3 Central Nervous System cancer, 3 endometrial cancer, 2 cervical cancer and one bowel cancer. Thirteen patients started treatment during follicular phase and 13 during luteal phase. We found similar results for the duration of treatment, total dose of follicle stimulating hormone, number of ampoules of gonadotropin releasing hormone antagonist, mean number of follicles identified at ultrasound on the day of trigger and retrieval, number of aspirated oocytes and Metaphase II oocytes. Conclusion Random-start controlled ovarian stimulation for emergency fertility preservation for minimizing delay in oncologic treatment for cancer patients does not interfere with the number of metaphase II oocytes, and therefore can be routinely used for stimulation followed by cryopreservation.

Pre-implantation genetic diagnosis (PGD) or screening (PGS) technology, has emerged and developed in the past few years, benefiting couples as it allows the selection and transfer of healthy embryos during IVF treatments. These techniques can be performed in oocytes (polar-body biopsy) or embryos (blastomere or trophectoderm biopsy). In this case report, we describe the first two live births to be published in Brazil after a polar-body (PB) biopsy. In case 1, a 42-year-old was submitted to PB biopsy with PGS due to advanced maternal age and poor ovarian reserve. Five MII oocytes underwent first and second polar body biopsy and four cleavage embryos were cryopreserved. The PGS analysis resulted in two euploid embryos (next generation sequence). A frozen-thawed embryo transfer (FET) was performed after endometrial priming and a healthy baby was delivered after a cesarean section (37 weeks, female, 3390g, 47.5 cm). In case 2, a 40-year old patient with balanced translocation and poor ovarian response was submitted to PB biopsy. Two MII oocytes underwent first and second polar body biopsy and two embryos were cryopreserved in cleavage stage. The analysis resulted in one euploid embryo that was transferred after endometrial priming. A preterm healthy baby (34 weeks, female, 2100g, 40 cm) was delivered via cesarean section. In conclusion, although the blastocyst biopsy is the norm when performing PGS/PGD during IVF treatments, other alternatives (as PB biopsy) should be considered in some specific situations.

Objective To evaluate if there are differences in the risks of obstetric outcomes in IVF/ICSI singleton pregnancies when compared fresh to frozen-thawed embryo transfers (FET). Methods This was a systematic review and meta-analysis evaluating the obstetric outcomes in singleton pregnancies after FET and fresh embryo transfer. The outcomes included in this study were pregnancy-induced hypertension (PIH), pre-eclampsia, placenta previa, and placenta accreta. Results The search yielded 654 papers, 6 of which met the inclusion criteria and reported on obstetric outcomes. When comparing pregnancies that arose from FET or fresh embryo transfer, there was an increase in the risk of obstetric complications in pregnancies resulting from FET when compared to those emerging from fresh embryo transfers in PIH (aOR 1.82; 95% CI 1.24-2.68), pre-eclampsia (aOR 1.32, 95% CI 1.07, 1.63), and placenta accreta (aOR 3.51, 95% CI 2.04-6.05). There were no significant differences in the risk between the FET and fresh embryo transfer groups when evaluating placenta previa (aOR 0.70; 95% CI 0.46-1.08). Conclusion The obstetric outcomes observed in pregnancies arising from ART may differ among fresh and FET cycles. Thus, when evaluating to perform a fresh embryo transfer or a freeze-all cycle, these differences found in obstetric outcomes between fresh and FET should be taken into account. The adverse obstetric outcomes after FET found in this study emphasize that the freeze-all policy should not be offered to all the patients, but should be offered to those with a clear indication of the benefit of this strategy.

The freeze-all strategy has emerged as an alternative to fresh embryo transfer (ET) during in vitro fertilization (IVF) cycles. Although fresh ET is the norm during assisted reproductive therapies (ART), there are many concerns about the possible adverse effects of controlled ovarian stimulation (COS) over the endometrium. The supra-physiologic hormonal levels that occur during a conventional COS are associated with modifications in the peri-implantation endometrium, which may be related to a decrease in pregnancy rates and poorer obstetric and perinatal outcomes when comparing fresh to frozen-thawed embryo transfers. The main objective of this study was to assess the available literature regarding the freeze-all strategy in IVF cycles, in regards to effectiveness and safety. Although there are many potential advantages in performing a freeze-all cycle over a fresh ET, it seems that the freeze-all strategy is not designed for all IVF patients. There is a need to develop a non-invasive clinical tool to evaluate the endometrial receptivity during a fresh cycle, which enables the selection of patients that would benefit from this strategy. Today, it is reasonable to perform elective cryopreservation of all oocytes/embryos in cases with a risk of OHSS development, and in patients with supra-physiologic hormonal levels during the follicular phase of COS. It is not clear if all normal responders and poor responders may benefit from this strategy.

Objective To evaluate the cost-effectiveness of freeze-all cycles when compared to fresh embryo transfer. Methods This was an observational study with a cost-effectiveness analysis. The analysis consisted of 530 intracytoplasmic sperm injection (ICSI) cycles in a private center in Brazil between January 2012 and December 2013. A total of 530 intracytoplasmic sperm injection (ICSI) cycles - 351 fresh embryo transfers and 179 freeze-all cycles - with a gonadotropin-releasing hormone (GnRH) antagonist protocol and day 3 embryo transfers. Results The pregnancy rate was 31.1% in the fresh group and 39.7% in the freeze-all group. We performed two scenario analyses for costs. In scenario 1, we included those costs associated with the ICSI cycle (monitoring during controlled ovarian stimulation [COS], oocyte retrieval, embryo transfer, IVF laboratory, and medical costs), embryo cryopreservation of supernumerary embryos, hormone measurements during COS and endometrial priming, medication use (during COS, endometrial priming, and luteal phase support), ultrasound scan for frozen- thawed embryo transfer (FET), obstetric ultrasounds, and miscarriage. The total cost (in USD) per pregnancy was statistically lower in the freeze-all cycles (19,156.73 ± 1,732.99) when compared to the fresh cycles (23,059.72 ± 2,347.02). Even in Scenario 2, when charging all of the patients in the freeze-all group for cryopreservation (regardless of supernumerary embryos) and for FET, the fresh cycles had a statistically significant increase in treatment costs per ongoing pregnancy. Conclusions The results presented in this study suggest that the freeze-all policy is a cost-effective strategy when compared to fresh embryo transfer.