JBRA Assist. Reprod. 2019;23(3):301-320
POSTER PRESENTATIONS
doi: 10.5935/1518-0557.20190052
Abstracts of the 23rd Annual Congress of the SBRA, Curitba/PR, 2019
1Hospital de Clínicas de Porto Alegre
2Universidade Federal do Rio Grande do Sul
Obejective: Despite recent advances in assisted reproduction techniques, implantation and birth rates remain low. Although embryo quality is considered the main determinant of implantation success, maturity and endometrial receptivity are also important factors to consider. However, what precisely constitutes a receptive endometrium remains uncertain. The objective of this study is to investigate if clomiphene citrate alters the endometrial maturation in infertile patients.
Methods: In a prospective self-matched cohort study, the ovulation of women was evaluated in spontaneous and stimulated cycles (with clomiphene citrate). Subjects were submitted to transvaginal ultrasounds to follow the cycle and to determine the day of ovulation. In both cycles, four blood samples (BS1 - at early proliferative phase, BS2 - at mid proliferative phase, BS3 - after ovulation and BS4 - at mid luteal phase) were taken to determine the serum concentrations of FSH, LH, estradiol and progesterone. An endometrial biopsy was performed five days after ovulation in both cycles and sent to Hospital de Clínicas de Porto Alegre pathology lab for analysis and classification according to Noyes criteria.
Results: Twenty-two participants completed the study protocol. There were significant differences in FSH BS3 (p = 0.001), in LH BS3 and BS4 (p<0.001 and p=0.049, respectively), in estradiol BS2, BS3 and BS4 (p<0.001, p=0.024 and p<0.001, respectively) and in progesterone BS3 and BS4 (p = 0.028 and p<0.001, respectively). Considering Noyes criteria, there was a one-day delay when comparing the stimulated cycle with the spontaneous cycle (p=0.004), and a two-day delay when comparing the stimulated cycle with the biopsy day.
Discussion: This study indicates that the ovarian stimulation with clomiphene citrate delays the endometrial maturity, and could possibly impair the implantation process. The antiestrogenic effects of clomiphene citrate in the endometrium may be responsible for this difference and, to avoid this effect, it is suggested to consider the freeze-all strategy when using clomiphene citrate for controlled ovarian stimulation, in order to minimize the endometrial asynchrony.
1Huntington Centro de Medicina Reprodutiva - São Paulo - SP - Brazil
2Department of Gynecology, Paulista School of Medicine, Federal University of São Paulo, São Paulo, SP, Brazil
3Department of Gynecology, School of Medicine, University of São Paulo, São Paulo, SP, Brazil
4Endometriosis Center - Hospital e Maternidade Santa Joana
Introduction: We report an extremely rare case of spontaneous single pregnancy followed by twin pregnancy in a woman with uterus bicornis unicollis after Deep infiltrating endometriosis (DIE) surgery.
Case report: A 26-year-old patient was admitted in our center with one year of infertility history after a miscarriage. She was diagnosed with uterus bicornis unicollis and deep infiltrating endometriosis (DIE); therefore, she underwent endometriosis focus removal surgery. After six-months, she conceived spontaneously and delivered one healthy baby. One year after the first pregnancy delivery, she conceived spontaneously and delivered twins in an extremely rare condition of uterus bicornis unicollis, of which there are only fifteen cases reported worldwide. Both pregnancies were monitored every two or three weeks using ultrasonography to assess fetal growth, and cervical length was measured to assess the risk of premature delivery.
Comments: The management of endometriosis-related infertility remains controversial, and optimal care should be individualized. A lack of consensus remains because different types of treatment strategies lead to similar pregnancy outcomes. The most commonly used treatments are videolaparoscopy and in vitro fertilization, and generally the woman's age, time of infertility, and semen analysis helps choose the treatment that will likely lead to better results. However, recent studies demonstrated that surgery improved pregnancy rates in infertile women diagnosed with endometriosis, mostly in younger women. A twin pregnancy with one gestational sac located in each horn of a uterus bicornis unicollis is an extremely rare condition, especially if the pregnancy is conceived spontaneously. We understood that it is a potentially high-risk pregnancy, and a literature review showed only fifteen cases of twin pregnancies associated with a bicornuate uterus Optimal twin pregnancy follow up is controversial in patients with Mullerian fusion anomalies, and treatment must be individualized because of the risks and rare occurrence of these cases. Serial ultrasound every two or three weeks is a worthy option to assess fetal growth and measure cervical length to assess the risk of premature delivery.
1Faculdade IPEMED de Ciências Médicas e Centro In Vitro Reprodução
2Curso de Pós-Graduação Lato Sensu em Dermatologia
Introduction: The objective is to share with the medical community knowledges about Darier-White Disease and to present a new genetic variant that was never described in the literature, as well as demonstrate the application of innovative genetic studies together with assisted reproduction techniques to avoid proliferation of the Darier-White disease in future generations.
Case report: First case in the world of assisted reproduction having prevented the progression of Darier-White disease in its decedents, associated the identification of mutation in splicing never before described in the literature. Complete sequencing of the ATP2A2 gene for Darier-White's disease was performed, which identified a splicing variant never before described in the medical literature, described as NM170665.3: _c.2607 + 1_2607 + 2insT, in heterozygosis, classified as VOUS. In order to elucidate whether this variant was also responsible for Darier-White's disease, in addition to those already described in the medical literature, a segregation study was performed on affected and healthy relatives of the patient using the PCR technique, and genotyping of variant NM170665.3 : _c.2607 + 1_2607 + 2insT in the ATP2A2 gene in relatives of the central patient, in this case were mapped: mother, maternal aunt and maternal cousin. With the identification and confirmation of the presence of the variant, we confirmed that it is possible to prevent offspring from being affected by the same disease through the pre-implantation genetic diagnosis for monogenic diseases (PGD).
Comments: Darier-White disease is a rare genodermatosis, with varying degrees of aggressiveness and clinical manifestations that surpass the limits of the skin, causing considerable social losses, is a genetic disorder of autosomal dominant inheritance, treatable, but incurable in those who already carry the disease. disease, however avoidable for future generations through the genetic isolation of diseased embryos, and thereby eliminating the disease in the offspring.
1Fertipraxis Centro de Reprodução Humana, RJ, Brazil
2Universidade Federal do Rio de Janeiro -UFRJ - Brazil
3Clinica Damian - RJ - Brazil
4Pontifícia Universidade Católica - Rio de Janeiro - Brazil
Objective: A group was created to gather different professionals whose work is related to reproductive techniques in order to share information and increase both theoretical and clinical insights about them.
Methods: Since September 20,2017 one psychoanalist and one clinician started a monthly meeting group of specialists that would be interested in discussing ART from an open view of their practices and the evolution of new developments on mentality and atitudes not only influencing their own practice as well as society changes. As time went by 04 other colleagues were invited and today the group is composed by 13 people: 04 psychonalysts, 01 psychologist, 03 reproductive clinicians,01 nurse, 01 anthropologist ,01 psychiatrist, 01 urologist and 01 anesthetist. The first members sign this paper.
Results: The meetings discuss the biomedicalisation of reproduction as well as the causes and consequences of the reproductive work, which is increasingly performed, its paradoxes and intersectional dimension. Thus came the demand for children, the sterility and absence of shadow depicted in literature and also in a Strauss opera, the notion of Don in different cultures, unborn children and the donation of gametes. psychic birth and the interface with assisted reproduction, male sexual dysfunction and infertility, visited in the movie "One More Chance", the discussion about Freud and his text "The Strange", Bioethics and ART, the role of nursing in listening, and lately, the parental myth.
Conclusion: We will continue to expand this forum of discussions and we intend to produce printed documents from these experiences.
1Hospital Pérola Byington - Centro de Referência da Saúde da Mulher- SP- Brazil
Introduction: During controlled ovarian stimulation, a precision between the LH peak and the oocyte recovery is essential for subsequent manipulation of the gametes. The human chorionic gonadotropin (hCG) is the gold standard to achieve this goal.
Objective: This study aims to evaluate the effect of the administration of hCG gonadotropin for final oocyte maturation with different intervals between its use and oocyte retrieval (36 and 38h), analyzing its impact on the oocyte recovery rate and its degree of maturation.
Method: A total of 78 patients were submitted to treatment with controlled ovarian stimulation and oocyte recovered between March 2017 and April 2018, in the Pérola Byington Hospital. The patients were randomly divided to be submitted to oocyte recovery at distinct intervals after hCG trigger: after 36 or 38 hours. A comparison was performed about number of follicles predicted on the day of the trigger, number of oocytes collected and number of mature oocytes collected.
Results: There was not statistically significant differences in a number of oocytes captured and maturation rate as a function of the different intervals - 36h or 38h - between the application of the hCG dose and oocyte retrieval. We also did not identify any case of ovulation that occurred before oocyte retrieval after 36h or 38h of hCG application.
Conclusion: The interval between the application of hCG and of oocyte retrieval may vary between 36 and 38 hours without impairing follicle maturation and without risk of ovulation.
1Fertipraxis Centro de Reprodução Humana - RJ, Brazil
2Huntington Centro de Medicina Reprodutiva - SP, Brazil
3Nilo Frantz Medicina Reprodutiva - RS, Brazil
4Clínica Monteblanco - Chile
5Clínica Concebir - Perú
6Cergyr - Argentina
Objective: REDLARA, the Latin American network of assisted reproduction, is attentive to the growth of the cycles in its region and is aware of details that can make the difference in the care (reception and persistence) in the treatments.
Methods: A questionnaire was created to identify different professionals whose work is performed in assisted reproductin clinics throughout Latin America in order to share information and improve the performance profile. From October till December 2018 an electronic questionnaire was sent to 195 REDLARA's centres directors, asking them to design the best one in their staff to answer 9 questions: who was the support professional in charge of the centre (non-doctor, non-embryologist, non- administrative manager), how many were involved to get this function, what were their qualifications, difficulties and needs.
Results: There resulted 56 answers, from 36 Nurses, 11 Matrons and 7 Nursing technicians or auxiliaries, relating 81 Nurses, 40 Midwives, 17 Matrons , 32 Surgical Nurses and 139 Nursing technicians/auxiliaries. In Latin America we identified 4 different areas of universitary education (Nursing, Matron, Midwifery and Surgical Nurses), with training that is sometimes different in years and contents, but which manage the centers of reproduction, from clinical to surgical environments. As they "learned in-service", the most difficult and necessary needs are the specific training in assisted reproduction.
Conclusion: From then on we identified the importance of interconnecting these professionals, seeking mechanisms for their continued education in assisted reproduction. REDLARA will start such a project in the pursuit of excellence.
1Merck B.V., Global Patient Safety, Schiphol Rijk, The Netherlands
2EMD Serono, Research & Development Institute, Billerica, U.S.A
3Merck KGaA, Global Medical Affairs Fertility, Darmstadt, Germany
Objective: Recombinant-human follicle-stimulating hormone (r-hFSH) is used for ovarian stimulation as part of treatment with assisted reproductive technologies (ART), with dosing (both starting dose and during treatment) individualized to optimize safety and efficacy. Reporting rates of OHSS and thromboembolism demonstrate that the risk after ovarian stimulation treatment with GONAL-f is low.
Methods: Reports of OHSS and thromboembolism were obtained from the Merck KGaA Global Safety Database from 29-Nov-2000 to 19-Oct-2018. Number of cycles were estimated from sales data based on an average 1875 IU administered per treatment-cycle.
Results: The systematic review identified 45 studies, including 5186 patients exposed to GONAL-f and 5240 treatment-cycles. Overall 272 reported cases of OHSS (reporting rate: 5190 per 100,000 treatment cycles; 5.19%) and 10 cases of severe OHSS (reporting rate: 191 per 100,000 treatment cycles; 0.19%) were identified; no fatal cases. No reports of thromboembolism were identified. In the Global Safety Database, there were 1110 reported cases of OHSS (using MedDRA preferred term=OHSS) and 80 reported cases of thromboembolic events (MedDRA SMQ = Embolic and thromboembolic events). Overall, there have been an estimated 16,525,975 treatment cycles since 29Nov-2000. This resulted in reporting rates for OHSS and thromboembolism of 6.7 per 100,000 treatment cycles (0.007%) and 0.48 per 100,000 treatment cycles (0.0005%), respectively. Three fatal cases were recorded; two fatal cases of OHSS and one fatal case of thromboembolism.
Conclusion: GONAL-f has been used for more than 18 years, with low rates of OHSS and thromboembolism. This provides reassurance about using GONAL-f for ovarian stimulation.
1Fertipraxis Centro de Reprodução Humana, RJ, Brazil
Objective: Assisted reproductive treatments are moving to individualized FSH dosing. This study aims to check serum AMH predicting the number of oocytes retrieved in stimulation with delta follitropin, based on a dosing algorithm.
Methods: Prospective, observational, unicentre study, from October 2018 to February 2019. 69 first ICSI or Oocyte Cryopreservation cycles, no age restriction. Fixed daily follitropin delta SC dose determined by serum AMH (Elecsys immunoassay® (ng/mL) and weight. Patients were divided in groups according to AMH levels: I 0,1-0,7; II 0,71-2,0; III 2,1-4,9; IV ≥5. Primary outcome was number of MII oocytes, ≤3, 4 to 7, 8 to 14 or ≥15.
Results:Mean parameters for age, days of stimulation and AMH values were respectively: Group I: 38.8; 9.8; 0.38; Group II: 38.4; 9.4; 1.31; Group III: 35.9; 9.2; 3.2; Group IV: 35.2; 9.2; 6.8. Primary outcome for number of MII oocytes retrieved were: Group I: ≤3 =9 and 4 to 7 = 8. Group II: ≤3=7; 4 to 7 =6 ; 8 to 14= 6. Group III: ≤3= 3, 4 to 7 =11 ; 8 to 14=10. Group IV:8 to 14= 2 and ≥15= 3. In groups I and II mean age was significantly higher than in Groups III and IV (p=0,004). There were no cancelled cycles or moderate/severe OHSS.
Conclusion: AMH can predict the number of oocyte MII retrieved by individualized dosing but groups II and III could benefit from higher follitropin doses. Besides, age remains the main barrier to overcome.
1Huntington Medicina Reprodutiva, São Paulo - Brasil
Objective: To identify male prognostic factors in obstructive azoospermia patients prior to PESA. We analyzed if male age, vasectomy time and sexual hormone levels could affect the quality of sperm from PESA and assisted reproduction outcomes.
Methods:Retrospective cohort study of institutional database, between November/2017 to January/2019. Fifty-three patients diagnosed with obstructive azoospermia were included. Patients were submitted to PESA and ICSI were performed if motile sperm were obtained. Sexual hormone data was available from 46 patients. Embryo parameters were also analyzed.
Results: Median testosterone and FSH levels were higher in patients which PESA yielded motile sperm (478 vs. 340 ng/dL, p=0.0353 and 5 vs. 3.5 IU/L, p=0.0337, respectively). Male age, BMI, time from vasectomy, estradiol, FSH, LH, and testosterone/estradiol ratio were not different between groups. Patients with testosterone above 350 ng/dL had a higher chance of providing motile sperm (88.9% vs. 50%, p=0.0149), with a relative risk (RR) of 4.1 (95% CI 1.51-11.21) and more likely to result in total sperm count higher than 0.001 millions/mL (p=0.0152, RR 3.87, 95% CI 1.32-11.36). Fertilization, blastocyst and top-quality blastocyst rates were not different between groups.
Conclusions: Testosterone levels above 350 ng/dL yields higher sperm concentration and motility in obstructive azoospermia patients after PESA. Men with lower testosterone may be better counselled on hormone managements or more complex procedures, in order to avoid unnecessary interventions.
1Department of Gynecology and Obstetrics, University of Sao Paulo, Ribeirao Preto, Brazil
2Department of Surgery, University of Utah School of Medicine, Salt Lake City, USA
3Drug Research and Development Center, Department of Surgery, Federal University of Ceara, Fortaleza, Brazil
4ANDROFERT, Andrology and Human Reproduction Clinic, Campinas, Brazil
Objective: To evaluate the global DNA methylation profile in spermatozoa of patients with varicocele, relating to seminal quality and sperm chromatin integrity.
Methods: A prospective observational case-control study, that included 26 men with varicocele and 26 fertile men who did not have the disease. Seminal quality was evaluated by spermogram and DNA fragmentation using the chromatin dispersion technique. The incidence of abnormal protamine expression was assessed by quantitative Western blot analysis. The sperm DNA methylation pattern was investigated using the Infinium MethylationEPIC BeadChip Kit 450K platform (Illumina, USA).
Results: Men with varicocele displayed decreased sperm concentrations (p=0.0004) and vitality (p=0.0224), and altered sperm morphologies (p=0.0233), as compared with controls. The sperm DNA fragmentation and protamine ratio were similar between the varicocele and control groups. Whole DNA methylation analysis showed that men with varicocele exhibit hypomethylation of sperm DNA in comparison to the control group without the disease (p=0.0373). The total of 59 CpG sites were identified, out of five were hypermethylated and 54 were hypomethylated in the varicocele group. In regional analyzes, 1695 DNA regions were differentially methylated in men with varicocele, and they were related to gametogenesis, meiotic and meiosis cell cycle, and seminal quality.
Conclusion: Besides to decreasing seminal quality, men with varicocele have a differential methylation profile when compared to men without the disease, either in a global, locus- or region-specific analysis.
1Universidade Estadual Paulista "Júlio de Mesquita Filho" - UNESP FCL/Assis Curso de Engenharia Biotecnológica
2Universidade Estadual Paulista "Júlio de Mesquita Filho" - UNESP. Laboratório de Matemática Aplicada - FCL/Assis - SP/ Departamento de Ciências Biológicas
Objective: Apply the Artificial Neural Networks technique in conjunction with the Genetic Algorithm for the determination of success pregnancy from the physiological data of patients submitted to assisted fertilization.
Methods: A database of 807 patients was used, which contained the physiological parameters of the patient adopted as input for the Artificial Neural Network, being: age, number of attempts at fertilization, number of oocytes, number and size of ovarian follicles and mass index body. The output parameter adopted was positive or negative live birth. For optimization of the Artificial Neural Network, a Genetic Algorithm technique was used.
Results: The results were 77% of the correct for the cases of positive live birth and 76% for the negative live birth. The AUC values for the training data were 0.8743 for live births and 0.8729 for non-birth cases. Those that were for the test data were 0.4753 for success cases and 0.5021 for cases of gestational failure.
Conclusion: The results show that the technique still needs adjustments, since for the training data we obtained great results considering the AUC values, however we did not obtain the same good results for the test data, indicating, thus, the necessity of change or insertion of some parameters from the patients, as input variable for the Artificial Neural Network.
Support: FAPESP (Processes 2017 / 19323-5) and CNPQ bolsa PIBIC / UNESP.
1Universidade Estadual Paulista "Júlio de Mesquita Filho", FLC - Assis; Curso de Engenharia Biotecnológica
2Universidade Estadual Paulista "Júlio de Mesquita Filho", FLC - Assis; Departamento de Ciências Biológicas
Objective: To use the Artificial Intelligence technique and the cleavage times for the prediction of the formation of the blastocyst stage in human embryos.
Methods: With the time-lapse monitoring performed by the EmbryoScope® devices, 5400 embryo cleavage samples were extracted. Based on these kinetic data, twelve variables were selected, presented according to the nomenclature usually found in the time-lapse system, t2, t3, t4, t5, t6, t7, t8, s2, s3, CC2, CC3, and RelCC2.For the prediction of the blastocyst formation, an artificial intelligence algorithm was developed, using the artificial neural network technique associated with the genetic algorithm, in order to find the optimal network for the proposed problem.
Results: The results point to an artificial neural network with 94.74% accuracy and AUC for training and test, respectively, of 0.8110 and 0.8305 for the prediction of blastocyst formation. While for the prediction of non-formation of the blastocyst the accuracy was 62.59% with an AUC for training and testing, respectively, of 0.8119 and 0.8318.
Conclusion: The technique presented excellent accuracy for the prediction of the blastocyst formation and less accuracy for non-formation of the blastocyst, despite good AUC values. The discrepancy between the results may be explained by the imbalance of the data distribution, since the database consisted of 4473 samples for the embryos that reached the blastocyst stage and 927 samples for non-formation of the blastocyst.
Support: FAPESP (Processes 2018/24252-2 and 2017/19323-5).
1Universidade Estadual Paulista "Júlio de Mesquita Filho" - UNESP. Laboratório de Matemática Aplicada - FCL/Assis - SP/ Departamento de Ciências Biológicas
2Imperial College London, UK
3Cornell University, New York, U.S.A.
4Instituto Valenciano de Infertilidad, Valência, Spain
Objective: This research used the technique of artificial intelligence - AI together with the technique of digital image processing to classify human embryos according to Gardner.
Methods: The database of 225 blastocyst images was obtained from the time-lapse imaging software (EmbryoViewerTMSoftware-Vitrolife-Sweden), and divided into 70%, 15% and 15% for training, testing and validation respectively. In order to determine the input variables of AI, digital image processing was used to standardize the images on their characteristics, resolution and illumination, using a computational algorithm on the MatLab® platform. At the end we obtained 33 variables that mathematically define the human embryo.
Results: For the classification of the expanded blastocyst there was an overall accuracy of 72.7% for the test data and 83.4% for the training data, for the classification of the internal cell mass there was a success of 97% and 86% respectively and for the trofectoderm classification we obtained 78.8% and 84.7% of correct answers. The mean AUCs for the expanded blastocyst were 0.8129 and 0.9165 for the test and training data respectively, for the internal cell mass we obtained 0.9926 and 0.9719 and for the trofectoderm data we obtained mean values of 0.8071 and 0.9409.
Conclusion: Based on the results obtained, we observed that the AI technique applied to the classification of human embryos obtained excellent indexes of correctness, which makes it promising as a noninvasive and objective technique, that is, without subjectivity to be used daily in the clinics of assisted reproduction.
Support: FAPESP (Process 2017 / 19323-5).
1Fertilitat - Centro de Medicina Reprodutiva
2Pontifícia Universidade Católica do Rio Grande do Sul - PUCRS
Objective: Oocyte freezing is the gold standard technique for fertility preservation. It is believed that the number of vitrified mature oocytes in the oncological population is lower than in patients submitted to elective freezing. The objective of this study was to verify this hypothesis and compare it with clinical and laboratory data.
Method: A cross-sectional study, carried out at a reproductive medicine center between 2009 and 2018. The sample consisted of 341 patients divided into two groups: neoplasia (n=39) and elective (n=302). Data were presented as mean±standard deviation or median (IIQ) and percentage, according to the distribution of the variables. Data were analyzed using Student's t-test, Mann-Whitney test or Chi-square test, considering p<0.05.
Results: The mean age in the neoplasia group was 36.5±2.4 and in the elective group 31.0±5.5 years (p<0.001). The basal FSH of the neoplasia group was significantly lower when compared to the elective group: 3.8mUI / mL (3.3; 5.5) and 6.8 (5.4; 9.6) mIU / mL groups (p<0.001 ). However, the median of mature oocytes in the neoplasia 7 (3; 12) group was not inferior to the elective group 6 (3; 11) (p=0.75), as the rate of mature oocytes / total oocytes did not show significance statistic: group neoplasia 0.68 ± 0.26 vs. elective group 0.72±0.24 (p=0.48).
Conclusion: The presence of neoplasias does not seem to influence the number of viable mature oocytes for vitrification when compared to the elective group. This result strengthens the continuity of indication for fertility preservation in this population. It should be noted, however, that studies with larger cases are needed to strengthen this evidence.
1Fertility Medical Group - São Paulo -Brazil
2Instituto Sapientiae - Centro de Estudos e Pesquisa em Reprodução Assistida - São Paulo -Brazil
Objective: To compare ovarian response to controlled ovarian stimulation (COS) and intracytoplasmic sperm injection (ICSI) outcomes between patients in which the trigger of final follicular maturation was achieved with dual trigger or recombinant human chorionic gonadotropin (r-hCG) only.
Methods:A case-control within-subject analysis was performed to compare COS and ICSI outcomes in patients which had the first ICSI cycle triggered with r-hCG (r-hCG Group, n=18) and the following ICSI cycle with dual trigger (Dual Trigger Group, n=18). The outcomes that were significantly different were used as cut-offs to select patients that had two ICSI cycles triggered with r-hCG (r-hCG 1st Cycle Group, n=18, and r-hCG 2nd Cycle Group, n=18), in order to verify if patients presenting with similar characteristics would also benefit from a following cycle triggered with r-hCG.
Results: When repeated cycles (r-hCG only vs. dual trigger) were investigated, it was observed higher oocyte yield, and mature oocyte rate, lower immature oocyte rate, higher fertilization rate, and higher blastocyst development rate in Dual Trigger Group. Higher rates of cycles with embryo transferred and implantation were observed in Dual Trigger Group. When repeated cycles (r-hCG vs. r-hCG) were investigated, lower immature oocyte rate and higher implantation rate were observed in r-hCG Trigger 2nd Cycle Group.
Conclusion: Dual trigger regimen is a more effective approach than r-hCG trigger in patients with maternal age > 37 years-old, and previous low rates of oocyte retrieval, mature oocyte and blastocyst development, yielding improved response to COS, and laboratorial and clinical outcomes.
1Nilo Frantz Medicina Reprodutiva, Porto Alegre, RS, Brazil
Objective: This study aims to evaluate cycles of patients who performed day four embryo transfer and compare them to blastocyst or cleavage stage embryo transfers.
Methods: The study selected patients from January to July 2018 who underwent fresh embryo transfer at day four (D4), three (D3) or five (D5) after oocyte collection. Group D4 was compared to groups D3 and D5. Student's t-test was used to compare age, body mass index, anti-Müllerian hormone, fertilization rate and good-quality cleavage stage embryos rate. Clinical pregnancy and implantation rates were assessed using Fisher's exact test.
Results: Ninety-six cycles were selected: 13 from D4, 23 from D3 and 60 from D5. The groups were similar to age (34.5, 36.0 and 33.6 years), body mass index (23.9, 24.7 and 23.1 Kg/m2), anti-Müllerian hormone (2.3, 1.5 and 1.8 ng/ml), fertilization rate (78.4%, 67.2% and 76.2%) and good-quality cleavage stage embryo rate (51.2%, 50.0% and 47.5%). Clinical pregnancy (46.2%, 34.8% and 55.0%) and implantation (34.6%, 25.0% and 39.2%) rates had no statistical difference. Clinical pregnancy and implantation rates were comparable between the groups.
Conclusion: Data suggests a trend of better rates on blastocyst embryo transfer, followed by D4 embryo transfer and, finally, cleavage stage embryo transfer, although data is not significant. Embryos in D4 were expected to undergo embryonic genome activation process, helping in selection of embryos to transfer when extended culture to day five is not possible. Our data suggests embryo transfer at day four is a viable alternative and can be recommended.
1Universidade Estadual Paulista, UNESP FCL-Assis, Departamento de Ciências Biológicas
2Imperial College London, UK
Objective: The use of human blastocyst digital images processing for the extraction of predictive mathematical variables of morphological quality that, associated with embryonic morphokinetic variables, were applied in a software that predicts with high accuracy the potential for live birth.
Methods: About one thousand digital images, provided by the IVI Valencia (Spain) and Weill Cornell Medicine (USA) clinics obtained by time-lapse system (EmbryoScope), were standardized and segmented for variable extraction, using algorithms developed on the MatLab® platform. Morphokinetic information was used in isolation or together relating cleavage times. These morphological and morphokinetic parameters were applied to the input variables of the evaluated software.
Results: Eight variables related to morphology were obtained: contrast, correlation, energy, homogeneity, area and eccentricity of the inner cell mass, blastocoel area and blastocyst expansion. The most promising morphokinetic variables in prediction of live birth were: t2, t3, t4, t5, t6, t7 and t8 (time, in hours, from fertilization/insemination to division into two, three, four, five, six, seven and eight cells to be complete, respectively); tSB (time, in hours, onset of blastocyst formation); tB (time, in hours, when the blastocyst is evident); (t3 - t2), CC3 (t5 - t3), RelCC2 (t3 - t2 / t5 - t2 - x100), t4 - t3, tSB - t8 and tB - tSB values.
Conclusion: Predictive variables of the morphological quality of the blastocyst associated with variables originating from embryonic morphokinetic resulted in a promising prediction of live birth.
Support: FAPESP (Processes 2018/19053-0 and 2017/19323-5).
1Valencian Institute of Infertility - IVI Salvador, Bahia, Brazil
Objective: To evaluate clinical results (clinical pregnancy and implantation rates) of the egg recipient patients in a shared egg donation program.
Methods: A retrospective cross-sectional study was carried out in which 70 patients were followed up in the IVI Salvador shared egg donation program, where IVF/ICSI cycles were performed from February 2018 to May 2019. Data were analyzed regarding sperm quality, age group of patients, mean number of donated eggs, fertilization rate, embryo quality, number of transferred embryos correlating with clinical pregnancy rate and implantation.
Results: Seminal quality was classified as normal in 17.14% of cases, regular in 51.4% and abnormal in 31.4%. The mean age of donors and egg recipients was 31.5 years and 43.9 years, respectively. The mean number of donated eggs was 7.6 ovules and the mean fertilization rate was 75.2%, and 78.5% of the transferred blastocyst embryos were classified as of good quality. Most patients (61.4%) received 2 embryos and the other (38.6%) received 1 embryo. The clinical pregnancy rate was 55.7% and the implantation rate was 42.1%.
Conclusion: A good rate of fertilization and embryo quality was obtained, which resulted positively in the clinical gestation rate.
1Universidade Estadual Paulista "Júlio de Mesquita Filho" (UNESP) - Faculdade de Ciências e Letras (FCL) / Assis. Curso de Engenharia Biotecnológica
2Imperial College London, UK
3Universidade Estadual Paulista "Júlio de Mesquita Filho" (UNESP) - Faculdade de Ciências e Letras (FCL) / Assis. Departamento de Ciências Biológicas
Objective: Through the concepts and patterns of human embryonic morfocinetic using the technique of artificial neural networks, sought to find the prediction of the most suitable embryos for transference.
Methods: The dataset counted with 9568 information of embryonic divisions, defined as: t2, t3, t4, t5, t6, t7 and t8, times, in hours, from fertilization to division in two, three, four, five, six, seven and eight cells are complete, respectively. We also use the time, the beginning of formation of the blastocyst and the one in which the blastocyst is in evidence. The relations CC2 = t3 - t2; CC3 = t5 - t3; RelCC2 = [(t3 - t2/t5 - t2)]x100; t4 - t3; tSB - t8 and tB - tSB, were also used. These variables present relationships of cleavage times and embryo development, seeking to represent their evolution mathematically. In the same dataset we have the information of birth or not of the transferred embryo, defined as positive or negative liverbirth, respectively. An artificial neural network model was constructed using the information mentioned, seeking an optimal network for the classification of blastocysts in livebirth positive/negative.
Results: The results obtained for the training data were 93% for livebirth positive classification and 85% for livebirth negative. For the blind test the result for livebirth negative was 82%, and for livebirth positive 59%.
Conclusion: The results show that the livebirth positive score for the blind test is still quite challenging, however, for the others, we obtained a very promising assertiveness.
Support: FAPESP (Processes 2018/19371-2 and 2017/19323-5).
1Department of Gynecology and Obstetrics, University of Sao Paulo, Ribeirao Preto, Brazil
2Drug Research and Development Center, Department of Surgery, Federal University of Ceara, Fortaleza, Brazil
Objective: Analysis of differential gene expression in men with varicocele using RNA sequencing data.
Methods: Sperm RNA sequencing data was obtained from a previous study of our research group, a case-control study that included semen samples from men with varicocele (n=6) and fertile men without disease (n=3). Quantification of the number of transcripts was performed by large-scale RNA sequencing (RNA-seq) using the TruSeq SBS kit v5 (Illumina, USA) on Illumina Genome Analyzer IIx equipment (Illumina, USA). The results of this study were using an R programming language, more specific the DESeq2 package and cluster Profiler. For the determination of a differentially expressed gene, the filters of p<0.05 and logFC>1.5 (| logFC |>1.5).
Results: Our findings showed that 494 genes were differently expressed between men with varicocele and fertile men without the disease. After data mining, HDCA2 (p=0,0417) and EHMT2 (p=0.0153) genes were less expressed in men with varicocele (logFC = -0.401) when compared to controls (logFC = 0.802). These genes are responsible for important functions associated with self-renewal and germ cell conservation. The alteration in the testicular microenvironment associated with varicocele, caused by thermal and oxidative stress, may be responsible for the genes expression alteration.
Conclusions: The HDCA2 and EHMT2 genes, essentials for spermatogenesis process, were less expressed in men with varicocele. Decreased or inhibited expression of these genes may lead to changes in seminal quality in patients with the disease.
1Huntington Medicina Reprodutiva, São Paulo - Brazil
Objective: To analyze paternal factors that could have impact on embryo aneuploidy rate in an oocyte donation program, including male age, semen parameters (sperm concentration, morphology) and sperm DNA fragmentation.
Materials and methods: Retrospective analysis of Preimplantation Genetic Testing for Aneuploidy data of biopsied embryos (blastocyst stage, NGS platform) between January 2017 and March 2019. Two-hundred twenty-nine biopsied embryos from 75 cycles were allocated according to paternal age: <= 41 years (n=26) and >= 42 years (n=49); sperm concentration in normozoospermic (n=67) and oligospermic (n=8); morphology according to Kruger's strict criteria >=4 (n=17) and <4 (n=58). DNA sperm fragmentation has been assessed in 29 cases.
Results: On average, 6.2 MII oocyte were fertilized and 1.9 blastocysts were formed. From 229 biopsied embryos, 143 were normal and 86 altered (37.5%, 79 numerical and 7 structural abnormalities). Advanced paternal age - median of 44.7 years old - and sperm concentration were not related to an increase in the absolute number of embryo aneuploidy (p=0.15 and 0.70, respectively). According to strict morphology, Kruger < 4% had 36% of aneuploidy comparing with 42.5% in Kruger >= 4 (p=0.38). Sperm DNA fragmentation in a 15% cut-off did not show statistic difference between the groups (p=0.08).
Conclusions: Therefore, these results suggested that the paternal factors, including age, sperm concentration, strict morphology and DNA sperm fragmentation were not related to the aneuploidy rate in preimplantation embryos in an oocyte donation program.
1Faculty of Medicine. University Center of João Pessoa-UNIPÊ.
2Faculty of Medicine Nova Esperança.
Objective: Show the endocrine-disruptive properties present in daily chemicals and how this interferes with biosynthesis and hormonal actions. Among them, bisphenol A (BPA), phthalates and perfluoroalkyl substances (PFAs) directly affect the female reproductive system.
Methods: Integrative literature review on the subject using the Scielo and Pubmed platforms as a database.
Results: DNA methylation is the main mechanism that can relate the use of phthalates and BPA to female reproductive disorders. Studies have shown that high levels of BPA were associated with lower numbers of recovered oocytes, mature oocytes from metaphase II, normally fertilized oocytes, low serum levels of E2 and a tendency for less blastocyst formation. Other studies correlate BPA and androgen levels, implying that BPA may play a role in the pathophysiology of PCOS. Phthalates are associated with less ovarian reserve in infertile women, and the literature indicates that it may inhibit the size of the growing array of antral follicles, impairing fertility and fecundity. PFAs appear to be related to parity and may affect steroidogenesis, leading to abnormally high androgens and contributing to the pathogenesis of PCOS.
Conclusions: Endocrine disruptors that are present in common substances can affect the females fertility. Therefore, it is necessary for gynecologists to know such properties in order to inform their patients about harmful products and their risks, especially in the area of reproduction.
1Faculdade de Medicina de Ribeirão Preto da Universidade de São Paulo (FMRP-USP)
2MicroControl LTDA - Supera Parque/USP Ribeirão Preto
3Invitra Tecnologia da Reprodução Assistida LTDA
Objective: To evaluate how microbiological monitoring practices are performed in Brazilian fertility clinics.
Methods: This cross-sectional study was carried out using a semi-structured electronic questionnaire containing questions elaborated based on the requirements for the manipulation of human gametes and embryos in an ISO Class 5 cleanroom.
Results: Of the 46 participating laboratories, 33 (71.7%) reported that they perform microbiological monitoring every six months, while 15.2% performed the tests once a year. Regarding the types of microbiological tests, the laboratories reported the use of contact plates (69.5%), swabs (63%), and settle plates (34.8%). The monitored environments most frequently cited were the interior of the vertical CO2 incubator (97.8%), the laminar flow cabinet (93.5%) and the air from the embryology laboratory (91.3%). Only 67.4% of the respondents (31/46) sampled the micromanipulator, considered a critical area. No consensus was observed regarding the number and location of sampling sites or ideal period to perform microbiological tests (activity 41.3%, rest 37%, both 21.7%). The occurrence of microbiological contamination of IVF cultures was reported by 26.1% of the laboratories.
Conclusion: Taken together, the data obtained from the responses by the embryologists points to the existence of many gaps in the practice of microbiological monitoring, including: lack of consistency in the sampling procedures (critical areas to be monitored, representative sampling points, ideal methods and period of time) among different clinics, and the lack of clear and definitive guidelines that address the different sources of microbiological contamination, the need for routine monitoring and specifications for its execution.
1Faculty of Medicine. University Center of João Pessoa-UNIPÊ.
2Faculty of Medicine Nova Esperança.
Objectives: The aim of this study is to investigate how cancer in the female genital tract can influence female fertility and seek to know about the most used methods for preserving fertility.
Methods: Integrative review of the literature on the subject, based on articles from the last fourteen years. It was used as a database: Scielo and PubMed. Researched during the period from January 2019 to May 2019.
Results: The therapies used to treat oncological diseases, chemotherapy and radiotherapy, involve cytotoxic drugs with a high germ cell lesion, especially ovules. Prior to the definitive treatment of the disease, embryo cryopreservation, oocytes, ovarian tissue, suppression of ovarian function and ovarian transposition were used in order to preserve fertility. One of the non-surgical options, hormone therapy, may be considered for women with low-grade endometrial cancer at an early stage. In addition, one should always try, when the treatment is surgical, to perform the fertility-sparing surgery, such as conization.
Conclusion: Treatment for cancer in the female reproductive system has consequences for the woman, but in some cases it is possible to perform the control, always aiming at the care of the patient as a whole, respecting their wishes and their dreams. A considerable number of patients who are affected by a neoplastic disease will survive it and may manifest reproductive desire in the future, so the importance of this subject will be studied more and more deeply.
1Nilo Frantz Medicina Reprodutiva, Porto Alegre, RS, Brazil
Objective: To describe the epidemiological profile of men from Rio Grande do Sul who seek for clinical investigation in a center of reproductive medicine in Porto Alegre.
Methods: Descriptive epidemiological study of 53 men who searched for medical assistance at a reproductive medicine center between December 2018 and June 2019. The Information was collected through the application of a questionnaire about main comorbities and diseases that may interfere with the reproductive status of the male participants.
Results: Men studied had between 30 and 54 years old (average 39), with the majority of Causasian participants (94%). Among the analyzed comorbities, high cholesterol levels are what most affects this group of patients (13.21%), followed by dermatitis (11.32%) and tabagism (9.43%). In relation to the comorbities recognized and handled by the patients stand out heart and lung diseases, diabetes and epilepsy. When considering reproductive health, mumps (16.98%) and varicocele (15.09%) were prevalent, although these were the diseases with lowest treatment indexes (22.22% and 25%, respectively).
Conclusion: The profile analyzed through this study brings together factors that are known to interfere with sperm concentration and quality, which are often disregarded in the evaluation of male fertility. Research and identification of these comorbities may be relevant when it is intended to improve the conditions of male reproductive health. Comorbities identification and further treatment prior to performing assisted reproductive cycles can result in higher success rates, optimizing in vitro fertilization treatments.
1Faculty of Medicine. University Center of João Pessoa-UNIPÊ.
2Faculty of Medicine Nova Esperança.
Objective: To present the judicial updates, in the tangent of post-mortem fertilization, of Brazil in relation to developed countries regarding the characteristics and the ethical aspects of the subject.
Methods: Bibliographic review of the topic, using a sample of 9 articles published between 2009-2017, indexed in the SCIELO, LILACS, BVS and PubMed databases, in English, Portuguese and Spanish, and selected according to the "classification of relevance "of these bases.
Results: Judicial authorization for sperm acquisition with intentional post-mortem fertilization is still a debatable issue, since each country has its own specific legislation. Several ethical factors are considered, such as the express authorization of the deceased party. In the Netherlands, Spain, Canada and Brazil (through Law nº 1.957 / 2010 of the Resolution of the Federal Medical Council), this technique is only allowed with this consent of the donor. In Belgium and the UK this is not necessary. In addition, in some countries such as Canada, fertilization is only allowed after a period of 6-12 months, allowing the family to consider all the implications of that decision. However, the Brazilian legislation does not define a period for carrying out the procedure.
Conclusion: Post-mortem fertilization is still considered taboo in Brazilian legislation, and it is difficult to defer the plausible ethical limits of this theme. However, it is necessary to define more criteria to judge this issue, such as the adequate time after the death to occur fertilization and a suitable psychiatric analysis of the individual who will use the genetic material to avoid precipitated measures.
1Igenomix Brazil - Sao Paulo - Brazil
2Igenomix SL - Valencia - Spain
Objective: Conventional karyotyping is the gold standard for chromosomal investigations of products of conception (POC). However, it is vulnerable to selective overgrowth of maternally derived cells, falsely producing a finding of normal female karyotype at rates of 29% to 89.7%. Our group has shown POC by next generation sequencing (NGS) combined with Short Tandem Repeat (STRs) for fingerprinting analysis of maternal cell contamination (MCC) is faster and can detect some abnormalities not possible with conventional karyotyping because of contamination, number of cells and resolution. AIM: To evaluate and describe chromosomal abnormalities in POC by NGS + STRs techniques.
Methods: 225 first and 10 second trimester miscarriage specimens plus patient´s blood were received between January 2017 and December 2018 at our laboratory. We performed NGS (Thermo Fisher®) plus STRs analysis to detect chromosomal abnormalities and discard MCC.
Results: Given the specimens from first trimester, 58.2% had abnormal results (42.6% with trisomies and 5.3% with monosomies; 8% were polyploid or triploid and 2.2% contained structural chromosome abnormalities) and 23% were normal (XX or XY). Only 17.3% had MCC and 1.3% were not evaluated because of their quality. Regarding the second trimester specimens, 40% had abnormal results (trisomy 18, triploidy and partial deletion of the chromosome 4), 50% were normal (XX or XY) and one sample presented MCC.
Conclusion: With NGS and STRs tissue origin could be determined when the result was normal so maternal contamination results were notably reduced in first and second trimester (< 20%) when compared to the traditional analysis.
1Fertilitat - Centro de Medicina Reprodutiva - Porto Alegre
2Departamento de Ginecologia e Obstetrícia da Faculdade de Medicina da PUCRS - Porto Alegre
Objective: To determine if the transfer of only one euploide embryo after thawing generates higher pregnancy rate when compared to transfer of 2 non analyzed blastocysts after thawing, in 2 different age groups.
Methods: Pregnancy and abortion rates were analyzed in 164 cycles where one euploid embryo was transferred following thaw and 317 cycles where two blastocysts embryos were transferred following thaw without biopsy. The patients were divided in two groups: <38 years and >38 years, chi-square test was used and the Fisher exact test (p< 0.05) using BioEstat 5.3 program.
Results: The group of patients <38 years were performed 132 cycles with a transfer of only one euploid embryo, it was observed 49% pregnancy rate and 6% abortion rate. In this same group, the patients that transferred two thawed blatocysts, the pregnancy rate and abortion rate were 55% and 12%, respectively (p=0,3108; p=0.274). When patients with > 38 years were compared the pregnancy rates for the transfer of one euploid embryo versus 2 non analyzed embryos were 72% and 47 % (p=0.0279) and the abortion rate were 10% and e 8% (p=0.995) respectively.
Conclusions: It was observed that patients with age >38 years, that transferred embryos biopsied increased significative the pregnancy rate/transfer. However for the group <38 years, the genetic analysis did not affect pregnancy and abortion rates, therefore the use of PGT for this group is not indicated, avoiding any risk of manipulating the embryos during the biopsy procedure.
1Disciplina de Ginecologia, Departamento de Obstetrícia e Ginecologia, Hospital das Clínicas, Faculdade de Medicina, Universidade de São Paulo, São Paulo, Brazil.
Objective: To determine risk factors for poor ovarian response (POR) to controlled stimulation in assisted reproduction cycles and to describe the outcomes of this specific interest group.
Methods:This is an observational retrospective study comparing 108 in vitro fertilization cycles from a POR group (< 4 oocytes retrieved) to 61 cycles from a control group (≥ 10 oocytes retrieved). All the cases came from a two-year period (2015-2016) and where conducted in a single quaternary center in Brazil.
Results: The POR group was older than the control group (mean age POR group 36.55±2.67 vs. 34.56 ±3.15 in the control group, p<0,001) and had a lower antral follicle count (mean 7.19±5.99 vs. 13.10±5.07, p<0,001). The POR group had shorter cycles, lower estradiol levels on the triggering day and thinner endometrium. They've had formed fewer embryos and most of the transfers of the POR group where done in cleavage stage compared to the control group, whose where mostly day-5 transfers. The POR group had lower clinical pregnancy rates from fresh transfers when compared to control group (p<0,001).
Conclusions: Age and ovarian reserve are associated with the response to ovarian stimulation. The oocyte recovery rate has a direct impact in IVF treatment success.
1Faculdade de Medicina de Ribeirão Preto da Universidade de São Paulo (FMRP-USP)
2Invitra Tecnologia da Reprodução Assistida LTDA - Supera Parque/USP
Objective: To investigate the influence of the supplementation of antioxidant (AX) coupled with different classes of lipids (LIPID) and/or phospholipids (PL) on the cryotolerance, survival, in vitro embryo development, and mitochondrial activity of oocytes vitrified in Tvitri-4 standard medium (T4 - Invitra Tecnologia da Reprodução Assistida Ltda.)
Methods: In this experimental study, in vivo matured oocytes of C57BL/6 mice were distributed in 5 groups: fresh control (CT) and vitrified: T4 (control), T4-AX, T4-LIPID and T4-PL. After vitrification and thaw, we analyzed oocyte survival. Viable oocytes were fertilized in vitro or used for analysis mitochondrial activity markers (15 oocytes per group): reactive oxygen species (ROS), oxidative metabolism, and mitochondrial membrane potential (JC-1). Oocytes from the 5 groups submitted to IVF were cultured for 96h and subsequently compared in terms of fertilization/blastocyst formation, total number of nuclei (NT) and internal cell mass (ICM)/trophectoderm cells (TC).
Results: The survival rate of oocytes vitrified in T4 was higher than the T4-AX, T4-LIPID and T4-PL (100%, 97.07%, 96.75% and 97.95%, respectively). The number of MCI cells in CT group embryos was higher than in T4 and T4-AX, and similar to T4-LIPID and T4-PL (p>0.05). Mitochondrial integrity and density (JC-1) was similar in oocytes from the CT, T4-LIPID and T4-PL groups.
Conclusion: Analysis of the MCI number and mitochondrial function outcomes suggests that the prospective lipid/antioxidant formulation improves oocyte cryotolerance and embryo quality. Mitochondrial integrity was preserved in oocytes treated with antioxidants, lipids and/or phospholipids during cryopreservation, and β-oxidation was stimulated in T4-LIPID oocytes.
1Humana Medicina Reprodutiva- Goiania- GO- Brazil
Objective: To evaluate the initial results of oocytes accumulation for patients with low ovarian reserve and patients with a low number of embryos to transfer in a previous cycle of in vitro fertilization without success.
Methods: The patients from Humana M.R. who opted for accumulation of eggs between April 2016 and May 2019 were evaluated. These patients were divided into two groups: Group 1 (26 patients aged 39 years) and Group 2 (17 patients between 40 and 42 years). It was evaluated the average of the antral follicles, the average number of collections per patient, the average number of ovules MII per patient, survival rate of MII eggs after devitrification, fertilization rate, percentage of embryos with ≥6 blastomeres in D3, chemical pregnancy rate when embryos was transferred with ≥6 blastomeres in D3, chemical pregnancy rate by ITT(treatment Intention = 2 egg collections with at least 1 frozen MII), evolution gestation for ITT rates and non-transfer rate.
Results: Initial Results: The average of antral follicles : Group 1 [5.15 (1-10)], Group 2 [4.35 (1-8)], the average number of collections per patient: Group 1 [3.22 (2- Group 2 [2.45 (2-4)], Group 2 [2.24 (2-4)], The average number of ovules in MII per patient: : Group 1 [ 2,45 (2-5)], Group 2 [2,41 (2-4)] ], survival rate of the ovules MII post devitrification: Group 1 [84,00%], Group 2 [73,00%], fertilization rate: Group 1 - [83,72%], Group 2 [68,20% ], percentage of embryos with ≥6 blastomers in D3: Group 1 [70.30%], Group 2 [60.77%]. Chemical pregnancy rate when was transferred embryos with ≥6 blastomers in D3: Group 1 [50,00%] and Group 2(44,00%). Chemical pregnancy for ITT: Group 1 [34.6%], Group 2 [23.52%] . Ongoing pregnancy rate for ITT: Group 1 [28, 90%], Group 2 [17.64%] and non-transfer rate: Group 1 [3.84%], Group 2 [17.60%].
Conclusion: These initial results have shown moderate evolving gestation rates in patients with worse prognosis, it motivates us to continue.
1ORIGEN, Rio de Janeiro
Objective: To quantify the effect on reproductive outcome of assisted hatching (AH) applied to overall in vitro fertilization (IVF) without any specific criteria concerning embryo and/or patients' characteristics.
Methods: This is a retrospective cohort study performed in the period between June - December 2017. The study included 598 embryo transfers in ICSI cycles, with fresh and frozen embryo transfer (FET) cycles (n=299 with AH; n=299 without AH), and with cleavage and blastocyst stage transfers. We included patients that performed the ICSI cycle with their own oocytes and also oocyte donation. Due to the heterogeneity of the group a multivariable analysis was performed using Logistic regression in which the effect of AH on ongoing pregnancy was calculated considering maternal age, oocyte origin (donated or own), embryo developmental stage, number of embryos transferred and the proportion of good quality embryos transferred as potential bias factors. The effect of AH was calculated with its corresponding odds ratio (OR) and the 95% Confidence Interval (CI) together with the OR of the potential bias variables. The statistical analysis was performed by using SPSS 26 software.
Results: We observed that the empirical use of AH is not independently associated with ongoing pregnancy rates in ICSI cycles neither in fresh nor FET cycles (OR=0.942; 95% CI: 0.645-1.376).
Conclusion: Our results suggest that there is still a lack of evidence to show in which group of patients AH would improve clinical outcomes. The overall use of AH does not improve clinical outcomes and may increase embryologist workload and embryo manipulation without any advantage.
1Center of Human Reproduction Insemine, Porto Alegre/ RS, Brazil.
Objectives: To evaluate the influence of blastomere symmetry at day 3 on blastocysts quality and biochemical pregnancy rate.
Methods: A cohort of 221 patients with blastocyst transfer from 2010 to 2019 at the Insemine Human Reproduction Center. Age, cause, AMH, FSH, number of recovered oocytes, number of MII, fertilization rate, embryo quality at day 3 and day 5 and blastomere symmetry between positive BhCG and negative BhCG groups were compared.
Results: The variables symmetry and embryonic quality at day 5 were statistically significant (p=0.027 and p=0.0001) between positive BhCG and negative BhCG groups. As well as the influence of symmetry in embryo quality at day 5 (p=0.032). A multivariate analysis controlling the pregnancy variable for symmetry and quality in D5 demonstrated independence and association with the outcome (p=0.032 and p=0.001). The analyzes of NNT between symmetry and pregnancy, quality at day 5 and pregnancy, symmetry and quality at day 5, respective values were obtained: 4.4 (95% CI 16.7 to 2.53), 3.4 (95% CI 6.36 to 2.32) and 6.67 ( 95% CI 3.26 to ∞).
Conclusion: The blastomere asymmetry at day 3 affects negatively the blastocysts quality, as well as the biochemical pregnancy rate. Patients with asymmetric embryos who developed good quality blastocysts are 6 times more likely to have negative biochemical gestation compared to patients who had good blastocysts from symmetric embryos.
1Igenomix Brazil - São Paulo - Brazil
2Igenomix SL - Valencia - Spain
Objective: Carrier screening test identifies autosomal recessive (AR) and X-linked mutations in individuals. Nowadays, it is becoming a standard practice for individuals with a positive family history of a recessive disease (RD) but few studies evaluated risks and the frequency for multiple single-gene disorders among Brazilian asymptomatic individuals. AIM: To evaluate the frequency of AR mutations obtained in our carrier screening tests (CGT600 by Igenomix®) for Brazilian patients (couples in reproduction treatments, egg donors, consanguineous).
Methods:Retrospective study. 268 adults with or without familiar history who underwent CGT600 tests between 2015-2019 at our centre were enrolled.
Results: 238 (88.05%) patients had no family history of rare disease and given the ethnicity, the majority was Caucasian (72.8%), followed by Latin (23.1%), Ashkenazi (2.2%), Arabian (0.75%), African (0.4%) and others (0.75%). About results, in the total 223 mutations were detected. Regarding the conditions: Caucasian were positive for Alpha-thalassemia (N=11) and Spinal muscular atrophy (N=8), Latin were Cystic fibrosis (N=5) and Alpha-thalassemia (N=4), Ashkenazi was Non-classic congenital adrenal hyperplasia (N=2), Arabian were Classic congenital adrenal hyperplasia (N=1) and Non-classic congenital adrenal hyperplasia (N=1), African was Retinitis pigmentosa type 43 (N=1).
Conclusion: The study shows the frequency of carriers of autosomal recessive disorders in Brazilian population is similar results to reported data using the same panel for different population. To prevent rare diseases prior to pregnancy, carrier testing may be suggested to patients and those with a positive may consider an IVF treatment plus PGD for monogenic diseases or gamete donation.
1Fertipraxis Centro de Reprodução Humana, RJ, Brazil
Objective: Follicular flushing with double-lumen retrieval needles attempts to increase oocyte yield. However, it takes more time to pick-up and higher costs to the procedure. Does follicular flushing improve the number of oocytes retrieved in poor responders? Methods: Prospective, comparative single centre study of 208 oocytes retrieval using follicular flushing (Wallace DNS1733) double lumen needle (n: 105) or direct aspiration (Wallace ONS1733) single lumen needle (n: 103), whenever ≤5follicles 15-17mm, ≤4 of 18mm on hCG day and ≤ 7 oocytes retrieved. Main outcome was number of Metaphase II.
Results: There were no differences in age 39.07±3.88 x 38.11±3.43, weight 61.73±17.53 x 65.96±15.44 or AMH 0.63±0.59 x 0.94±0.97. No difference on days of stimulation 9.57±1.87 x 10.29 ±2.82, Estradiol 788.94±670.82 x 940.16±694.69, Progesterone levels 617.29±319.76 x 561.18±486.78 and follicles ≥18mm 1.84±0.95 x 2.07±1.09. Although total of gonadotropins 1678.28±798.52 x 2080.45±852.36 (p=0.0008), number of oocytes 3.00±2.11 x 3.69±2.20 (p=0.02) and Metaphase II retrieved 2.20±1.64 x 2.99±1.88 (p=0.02) were significantly different, the ratio of total oocytes /mature follicle ≥15mm (0.93 x 0.98) and Metaphase II oocytes/mature follicle ≥15mm (0.68 x 0.79) were similar in both groups. Oocyte pick up failure occurred in 16% x 9.8%.
Conclusion: Considering that there were no differences in the oocyte per follicle ratio, the follicle flushing didn´t improve the number of oocytes retrieved in poor responders in our study.
1Centro de Reprodução Humana de São José do Preto (CRH Rio Preto), São José do Rio Preto, SP, Brasil
Objectives: To compare elective single embryo transfer (eSET) with non-elective single embryo transfer (n-eSET) of blastocyst and also compare single transfer of embryos (eSET e n-eSET) with double embryo transfer (DET) of blastocyst.
Methods: Prospective study, including 108 ICSI patients, (July/2017 to December/2018). Exclusion factors: azoospermia and age above 38 years. Patients were divided into 3 groups: Group 1: single embryo transfer: 10 patients; Group 2: single selective embryo transfer: 16 patients and Group 3: double embryo transfer: 82 patients.
Results: Mean age: group 1: 30.3 years; group 2: 31.4 years and group 3: 31.4 years (p(0.05). Mature oocytes: group 1: 70%; group 2: 69% and group 3: 69% (p = 0.05). ICSI fertilization rate: group 1: 81%; group 2: 70% and group 3: 71.8% (p>0.05). Rate of blastocysts: 53.3% group 1; 62.4% group 2 and 62.8% group 3 (p>0.05). Clinical pregnancy rate: 50% (5/10) group 1; 43.7% (7/16) group 2 and 59.7% (49/82) group 3 (p=0.32). Abortion rate: 0% in group 1; 14.3% group 2 and 22% group 3 (p=0.36). Multiple pregnancy rate: 0% in groups 1 and 2 and 25% (7/35) in group 3 (p=0.136). Conclusion: Blastocyst transfer in patients up to age 38 helps to select the embryo to transfer, minimizing risks of multiple gestation, without significantly affecting baby's home rate. Although N is not significant, these data help convince couples of single blastocyst transfer, since the difference in single to double transfer pregnancy rate is not significant, with negligible risk of multiples.
1Universidade do Vale do Itajaí (UNIVALI)
Objective: The objective of this study was to relate the morphological defects of spermatozoa heads with the flagellum edema class after the hyposmotic test (HOST).
Methods: Human seminal samples (N=11) were submitted to a hyposmotic solution in the proportion 1:1 during 5 minutes at 37°C. Posteriorly was performed smear and slides were stained with Quick Panoptic Kit (Laborclin®). In optical microscope (1000x) was performed and classified (N=200) the morphological defects of spermatozoa head and relationed with the types of flagellum edema (A-G) according to the World Health Organization.
Results: The spermatozoa with flagellum edema in the hyposmotic test of type 'D', followed of the type 'C', showed more normal morphological rate, being respectively 33% and 23%, and the spermatozoa with flagellum edema of type 'G' presented the less normal spermatozoa rate (12%), being the defect amorphous the biggest prevalence at the spermatozoa type 'G' (rate of 28%), while the spermatozoa type 'C' and 'D' showed respectively rates of the 20% and 15% amorphous.
Conclusion: It was observed that the spermatozoa with flagellum edema classified as 'C' and 'D' are the ones with the best head morphology in comparison with those of class 'G', which may be a selection criterion for intracytoplasmic sperm injection (ICSI), accomplished after the hyposmotic test (HOST).
1Instituto Ideia Fértil de Saúde Reprodutiva
Objective: Assisted Reproduction techniques have brought to the present scenario what was previously unthinkable. Today, indeed, new families are constituted through reception of semen. In Brazil there is no research that identifies motives for semen donation, so professionals involved in offering these technologies know little about the donors. Objective: To identify the main reasons referred by men to be donors of semen; knowing the psychological profile of donors; provide grants for staff to work more effectively and integrated with men who are candidates for donation.
Method: January 2008 to December 2013 men were approved to be semen donors. Along with the physical assessment questionnaire, the candidate answered some subjective questions.
Results: Eighty-three men were accepted for donation, the average age was 26 years old; 67% were single; attending university degree 40%; white people 66% and 70% without children until the moment of donation. According to the donors' perception about them who would receive his donation 22% thought they would be happier to receive their donation. Fifty-two percent consider themselves happy/kind/calm. Regarding the motivation to donate - help patient known (discount on treatment) were 22% and help woman/couple (unknown)55%, the others, 23%, interested in "perpetuating the species".
Conclusion: Narcissistic questions appeared as motivation to donate; in addition, the data indicate that altruism is the main motivation. When we consider the emotions, we know that "part of self donated to the other" involves many questions beyond pure altruism and that vehement dedication to the other carries within itself strong symbolic value.
1Center of Human Reproduction Insemine, Porto Alegre/ RS, Brazil.
Objective: To identify parameters that characterise the fertilization faillure occurence. Methods: The cases of fertilization faillure between january 2010 and april 2019 (n=139) and the cases without fertilization failure (n=424) composse the sample.
Results: Couples' ages were not diferente between groups: 37 years old [33-40] and 36 years old [33-40] for women and 38 years old [34-42] and 37 years old [33-42] for men in study group and control group, respectively, as well as serum levels of anti-Müllerian hormone. Serum levels of follicle stimulating hormone are reduced in the fertilization failure group, 6.40 UI/mL [4.23-9.50] in comparison to the control group, 7.20 UI/mL [5.90-9.27] (p=0.04). The Follicular Output RaTe (FORT) for the study group was 58.54%±6.38% and for the control group was 86.41%±3.72% (p = 0.001). The retrieved oocyte number, 2 [1-5] and 5 [3-8] (p=0.0001) and mature oocyte number, 2 [1-3] and 5 [2-7] (p=0.0001) for study and control group, respectively, presented the same pattern. Seminal volume, pre and post-processing spermatic concentration and pre-processing spermatic motility did not revealed statistical diferences between groups. However, post-processing spermatic motility for study group was 80 million/mL [70-90] and for control group was 90 million/mL [80-95] (p=0.0001). Linear regression resulted in a statisticallly significant model [F(6,67) = 3.45; p<0.005; R² = 0.236].
Conclusion: FORT value and post-processing spermatic motility are predictors of fertilization failure.
1Departamento de Ginecologia e Obstetrícia - Faculdade de Medicina- Universidade de São Paulo -Ribeirão Preto- Brazil
Objective: This study aimed to evaluate the effects of in vitro follicular development on oxidative metabolism, on the generation of cellular oxidative stress and levels of genes. Methods: Three-dimensional in vitro culture of bovine secondary follicles was carried out for 16 days in supplemented α-MEM medium, in an atmosphere of 5% CO2 and 20% O2. As controls two groups of fresh follicles were included. Were performed morphological, analysis RT-qPCR expression of target genes, quantification of intracellular reactive oxygen species and the redox state evaluation of the mitochondria and the oocyte cytoplasm.
Results:In this study a reduced production of reactive oxygen species was observed in oocytes from follicles cultured in vitro in relation to oocytes obtained from the control groups: fresh secondary follicles (p<0.0065) and in vivo partially matured pre-antral follicles (p<0.001). However, it was observed that the oxidative metabolismo redox status of the mitochondria and the cytoplasm of oocytes obtained from in vitro culture was higher when compared to controls (p>0.05). Analysis of mRNA expression in oocyte and in follicular somatic cells showed a significant decrease in the relative quantification of Kit Ligand in the cultured follicle when compared to controls (p=0.03).
Conclusion: Although there was an increase in redox status in oocytes, the system was efficient in controlling the production of the reactive oxygen species during cell metabolism. The Kit Ligand gene expression indicated damage in the communication betweenfollicular somatic cells and the oocyte.
1Centro de Reprodução Humana Insemine
2Universidade Federal do Rio Grande do Sul
Objective: to correlate serum thyroid stimulating hormone (TSH) level with ovarian reserve and pregnancy rate.
Methods: A prospective study was conducted with 242 infertile patients undergoing in vitro fertilisation, without previous diagnosis of hypothyroidism. Thyroid function was evaluated through serum TSH. Patients were selected from January 2017 to December 2018, in a single human reproduction center. Serum antimullerian hormone (AMH) and antral follicle count (AFC) were performed to evaluate ovarian reserve. Serum dosage of human chorionic gonadotropin (beta-HCG) was used as a reproductive outcome, characterizing biochemical pregnancy if positive.
Results: The mean age was 35.9 years old (SD±4.44) and serum TSH mean was of 2.07 mU/L (SD±1.08). Biochemical pregnancy rate was 35.8%. There were no correlation between TSH and AMH levels (r= -0.13, p=0.866) and AFC (r=0.02, p=0.981), even as no association between biochemical pregnancy rate and TSH levels (Student's t-test, p=0.763).
Conclusion: in this particular population of women with good thyroid function control measured by TSH, there was no correlation between serum TSH levels with the ovarian reserve measured by AMH or AFC, as with the reproductive outcome evaluated by the biochemical pregnancy rate.
1Universidade do Vale do Itajaí (UNIVALI).
Objective: To investigate the effect of perfume on embryo development in vitro.
Methods: A total of 162 murine embryos from B6D2F1 females (C57BL/6 X DBA/2) were used. The 2-cell embryos were cultured (20µL) in GV-Blast medium (Ingámed®) supplemented with 10% Soro Ingámed (Ingámed®) and divided into 6 groups: Control 1 (C1)- cultured without exposure to the perfume and kept inside the incubator (n=21); Control 2 (C2)- cultured without exposure to the perfume and kept daily outside the incubator (n=21); T1- cultured on plates with scent brushed on the internal part of lids (10µL) 1 hour before exposure (n=30); T2- cultured on plates with scent brushed on the internal part of lids (10µL) 6 hours before exposure (n = 30); T3- cultured on plates with scent brushed on the internal part of lids (10 µL) 24 hours before exposure (n=30); T4- grown on fragrance-free plates daily handled by a person using perfume (20µL) applied one hour earlier (n=30). Groups C2 and T4 remained outside the incubator daily for 10 minutes under heated plate laminar flow (37°C).
Results: Groups C1, C2, T1, T2, T3 and T4 presented blastocyst rates of 95%, 100%, 100%, 97%, 97%, 97%, respectively, and hatching rates 60%, 57%, 57%, 41%, 59% and 62%, respectively.
Conclusion: The application of perfume (10µL) in the culture plates and used by a person (20µL) that manipulates the embryos outside the incubator daily did not interfere in the rates of blastocyst formation, although the T2 group showed a decrease in the hatching.
1Igenomix Brazil - Sao Paulo - Brazil
2Igenomix SL - Valencia - Spain
Objective: One of the possible mechanisms involved in Recurrent implantation failure (RIF) is the change in endometrial receptivity. Despite extensive research on underlying causes for it, our understanding is still limited. AIM: To evaluate endometrium receptivity profile based on endometrial receptivity analysis (ERA) test among women with RIF in Brazil.
Methods: 235 patients with RIF who underwent ERA test and submitted to hormone replacement therapy (HRT) (average progesterone= 119 hours) between 2015-2018 at our centre in Brazil were enrolled. Patients were stratified by age in five groups and their endometrial receptivity status was assessed by ERA computational predictor. Results were divided in 2 main groups: receptive ® (including early and late receptive) or non‐receptive (NR) (pre and post receptive). Data were analyzed comparing groups with Kruskal-Wallis test and considered statically significant when p≤0.05.
Results: Regarding the receptive results, a slight decrease in percentage rate was observed with increasing maternal age (58.8% in women <35 years; followed by 52.6%; 50.9%, 56.8% and 37.9% in ≥44years). In the non-receptive (NR) category, a little increase in percentage rate with advancing maternal age was detected (41,18 % in women <35 years, followed by 42.27%, 49.1 %, 43.18% and 62.07% in women ≥44years), although results were not statically significant (p=0,15).
Conclusion: Our results showed women with a history of RIF presented a displacement in the window of implantation and consequently a non-receptive endometrium, showing no relationship with maternal age and demonstrating the importance of the study of endometrial factor in these cases.
1Centro de Reprodução Humana Insemine
2Universidade Federal do Rio Grande do Sul
Objective: Endometriosis is associated with infertility; however, the exact mechanism is unclear and embryo development could be one of the factors involved. Our objective is to evaluate the embryo quality on Day 3 and 5 and symmetry on Day 3 in women with and without endometriosis undergoing in vitro fertilization.
Methods: It was a retrospective study with 221 patients and 1308 embryos. Inclusion criteria were women with infertility and indication of IVF/ ICSI who had at least 1 embryo Day 5 for fresh transfer. They were divided in two groups: with and without endometriosis. Statistic analysis was performed with SPSS, T-student test was used for parametric variables and qui-square for categorical variables. The significance threshold was set at 0.05 for P-value.
Results: The group with endometriosis had 47 patients (21.3%), and the group without endometriosis had 174 patients (78.7%). There was no diference of age (media 33.89 vs 35.05, P 0.138), oocyte number (media 9.02 vs 9.35, P 0.605), MII number (media 8.36 vs 8.08, P 0.613) or number of embryo transfer (1.96 vs 1.99, P 0.251) between the groups with and without the disease, respectively. In relation to embryo quality on Day 3, symmetry on Day 3 and quality on Day 5, also there was no statistical difference (P 0.470, P 1.000, P 0.431, respectively).
Conclusion: There was no difference in quality and symmetry of embryos in women with and without endometriosis undergoing IVF, according most recent studies about the fertility impact of endometriosis.
1Universidade Estadual do Oeste do Paraná - Campus Francisco Beltrão - PR.
Introduction: The examination method of choice for the investigation of infertility caused by uterus and fallopian tube abnormalities is the histerosalpingrography. It's considered a safe and easy method with a low complication's rate. Although, when facing adverse events, serious and even fatal endings can occur, one of then is the uterine perforation. In the presente study, a case of a fallopian tube disruption due to the exam is analised.
Caso Report: A 40-year-old female patient underwent HSG for infertility investigation on May 9th, 2019 in the city of Francisco Beltrão, Paraná. Soon after the procedure, which was reported as painful, the patient started a severe pain in the hypogastric region, evolving with constant pain but with lower intensity on subsequent days. He used anti-inflammatories and other analgesics in order to ease the pain, without success. On May 18th, there was intensification of hypogastric pain associated with tachypnea. Patient said got worse during the course of the day and was reported as unbearable late in the afternoon, prompting her to seek medical help. At admission to the emergency room at the Hospital São Francisco, she had severe pain in the hypogastric area associated with dyspnea, asthenia, inability to remain in the orthostatic position, sweating and cold skin. Upon physical examination, the patient was hypocorate, desaturated, gout, hypotensive, tachypneic, apathetic, with chills, axillary temperature of 36.5oC and blood pressure of 90x60 mmHg. To investigate the cause of hypotension, a computed tomography (CT) scan of the abdomen was performed, which showed moderate to large amounts of fluid in the abdominal cavity, suggesting that the acute abdomen was hemoperitoneal due to rupture of viscera. She was sent to the surgical center for unilateral salpingectomy and exploratory laparotomy, where hemoperitoneum and pelvic abscess were observed. On May 19th, due to the maintenance of signs of hypotension and hyposaturation, she was transferred to the intensive care unit, where vasoactive drugs, blood transfusion and volume resuscitation were administered, as well as orotracheal intubation to correct the observed desaturation. On 05/20, the patient was hemodynamically stable. On subsequent days, she presented clinical improvement, pulmonary improvement and reduction of hemoperitoneum, demonstrated in abdominal CT .On 05/23, she developed cerebral edema, causing unresponsive ocular opening and global motor deficit. On May 25th, he presented pneumothorax associated with partial collapse of the lung. After management of the condition, the patient presented clinical improvement and was transferred to the hospital bed for motor and pulmonary recovery. Currently, on May 31st, the only symptom referred is dyspnea.
Comments: HSG is a widely used method for assessing infertility because of its low cost, ease and practicality. Despite their importance, the number of studies and reviews published in recent years that address their complications is too limited. The case report addresses a tubal rupture event due to HSG, in which immediate intervention was necessary to correct the alterations at the genital and systemic level. In this context, the clinical case reinforces the need for medical class knowledge about HSG. Therefore, it is essential that more studies are conducted addressing adverse events, even infrequent ones, as well as their management.
1Fertilitat Centro de Medicina Reprodutiva
2Pontifícia Universidade Católica do Rio Grande do Sul - PUCRS
3Universidade Federal de Ciências da Saúde de Porto Alegre - UFCSPA
Objective: To describe the profile of patients submitted egg sharing and the outcomes obtained in 15 years of experience.
Methods: A retrospective cohort study, carried out at a reproductive medicine center, between 2002 and 2017, by medical records; 591 cycles of in vitro fertilization with egg-sharing were included.
Results: 293 donation cycles and 298 receiving cycles were performed. The average donors age was 30.6±3.3 years and the recipients was 43.0±5.0 years, the youngest recipient was 26 years and the oldest 51 years. The mean number of oocytes aspirated was 16.4±8.6; among this 12.7±7.0 was mature. After the division, the recipients obtained on average 6.4±3.2 oocytes in M2 stage. Spermatozoa were obtained from ejaculate (82.59% vs. 74.5%), sperm bank (6.18% vs. 2.3%) and testicular extraction (11, 26% vs. 2. 3.15%). Gestation rates, in a single fresh cycle 39.6% for donors and 36.4% for recipients.
Conclusion: Adequate numbers of oocytes were aspirated for sharing as well as satisfactory pregnancy rates, when analyzing only single fresh transfer. Broadening the above investigation with the inclusion of frozen embryo transfers and cumulative analysis of pregnancy rates tends to contribute for elucidation of prognostic factors at this modality of treatment, that increasing substantially in the last years and has few published epidemiological data about the Brazilian population.
1Universidade do Vale do Itajaí (UNIVALI).
2Fertas Brasil
Objective: To compare the development in vitro of continuously cultivated murine embryos, with partial or total exchange of the culture medium.
Methods: 186 murine embryos from females B6D2F1 (C57BL/6 X DBA/2) were used. After euthanasia, the uterine tubes were collected and flushed with GV-Hepes (Ingámed®) for the recovery of embryos in the zygote stage. These were cultivated in drops of 40µL of medium GV-Blast (Ingámed®) and supplemented with 10% of serum (Ingámed®) and divided in groups as followed: Continuous Group (G1) - cultivated continuously until the day 5 (n=46); Partial Exchange Group (G2) - with exchange of the 50% of the culture medium on day 3, with medium equilibrated for 3 days (n=46); Total Exchange Group A (G3) - embryos were transferred on day 3 to another drop, equilibrated with CO2 for 3 days (n=47); Total Exchange Group B (G4) - the embryos were transferred at the day 3 to another drop, equilibrated overnight (n=47).
Results: Rates of blastocyst formation in groups G1, G2, G3 and G4 were 83%, 85%, 83% and 77% respectively, and the hatching rates were 61%, 64%, 62% and 69% respectively. The results didn't present statistical difference.
Conclusion: The culture conditions tested did not interfere significantly in the formation and hatching of blastocysts, suggesting that any of these culture forms can be used without interfering in embryonic development.
1ANDROFERT - Centro de Referência para Reprodução Masculina - Campinas - SP
Objective: Fabry disease (DF) is a rare condition associated to the X-chromosome. It is caused by mutations in the GLA gene, whose incidence varies from 1: 1,250 to 1: 117,000. This mutation results in deficit or absence of the enzyme alfagalactosidase-A, responsible for the degradation of the glycolipids in the lysosomes. The progressive accumulation of globuli-silyceramide (GL-3) and glycosphingolipids, triggers a cascade of events beginning with a dysfunction of basic metabolic processes at cellular level moving to a multiple organ progression. Azoospermia in men diagnosed with DF has not yet been elucidated but it may be related to deposit of GL-3 in Leydig cells, and is more likely to occur after 30-40 years old. The present study describes a case of azoospermia in a patient with FD submitted to IVF treatment.
Methods: Case report.
Results: Nine years of primary infertility, 33 years old woman and 40 years old azoospermic patient diagnosed with DF at the age of 27 and submitted to a testicular microdissection (microTESE). Obtained motile and viable spermatozoa, allowing its use for ICSI and cryopreservation. Ovarian stimulation followed by oocyte retrival, obtaining 8 MII, with 6 of them fertilized (75%, 6/8). Was chosen to transfer 2 embryos in Day-3 and culture the other embryos. Cryopreserved 2 blastocysts (66.7%, 2/3). Single pregnancy in progress.
Conclusion: Enzyme replacement therapies have not been effective for the reversal of azoospermia. Thus, the use of microTESe combined with cryopreservation techniques must be considered viable, with cryopreservation being an alternative for preserving fertility, before the reduction of seminal parameters.
1Federal University of São Paulo, Department of Gynecology, São Paulo-SP, Brazil;
2Rey Juan Carlos University, Department of Gynecology and Obstetrics, Madrid, Spain
3IVIRMA Global Madrid, Madrid, Spain.
4Mater Prime, São Paulo-SP, Brazil.
5Origen - Center for Reproductive Medicine, Rio de Janeiro-RJ, Brazil.
Objective: To evaluate the overall performance and oocyte quality of follicular (FPS) versus luteal phase stimulation (LPS) among patients undergoing double ovarian stimulation (DuoStim).
Methods:Observational retrospective two-center cohort study including 79 infertile women who underwent a total of 87 DuoStim cycles between January 2017 and May 2019. Besides assessing baseline characteristics in order to determine the patients' clinical profile, we analyzed the FPS and LPS regarding the total dose of gonadotropin received, the duration of stimulation, the number and maturity of oocytes, fertilization and blastocyst formation rates, and the number of blastocysts obtained.
Results: The patients' baseline characteristics were compatible with a diminished ovarian reserve and poor reproductive prognosis. While the luteal phase needed longer stimulation [12 days (5-19) vs. 11 (7-16), p<0.001] and slightly higher gonadotropin doses [2946 ± 890 IU vs. 2550±970 IU, p<0.001], no significant differences were detected in the oocyte maturity, fertilization and blastocyst formation rates. However, the number of oocytes retrieved [5 (0-16) vs. 4 (0-15), p=0.006], mature oocytes [4 (0-15) vs. 3 (0-11), p=0.032], and blastocysts obtained (70 vs. 53) were substantially greater after 4 LPS.
Conclusion: The DuoStim strategy in poor prognosis patients increases the number of oocytes and blastocysts available. Moreover, the number of oocytes and blastocysts obtained are higher after LPS when compared to FPS. Thus, it should be considered for selected patients in order to not only improve reproductive outcomes but also shorten the time to pregnancy.
1Faculdade de Medicina da PUC-RS (Pontifícia Universidade Católica do Rio Grande do Sul).
2Fertilitat Centro de Medicina Reprodutiva.
Objective: Compare ovarian stimulation outcomes between patients with normal body mass index (BMI), overweight and obese who underwent thru in vitro fertilization (IVF).
Methods: Retrospective case-control study with patients undergoing IVF between 2013 and 2018, performed at Fertilitat - Centro de Reprodução Humana, Brazil; divided into three groups: 1) body mass index (BMI) <25 Kg/m² (n=231); 2) BMI>25 Kg/m² <29,9 Kg/m² (n=228); 3) BMI ≥ 30 Kg/m² (n=66), with a total of 525 fresh embryos cycles, from 298 patients. Number of oocytes aspirated, mature oocytes and oocyte maturation rate were analysed.
Results: Comparing G1 vs. G2 vs. G3, there were: oocytes aspirated, (11[6-16] vs. 8[4-12] vs. 7[3-12.25], p=0,42), mature oocytes (8[5-12] vs. 5[3-9] vs. 5[2-8,25], p=0,40) and oocyte maturation rate (0.83[0.70-0.91] vs. 0.8[0,67-1] vs. 0,75[0.63-0.92], p=0.37). Comparing G1 vs. G3: oocytes aspirated (p<0,001), mature (p<0.001), oocyte maturation rate (p=0.04). A weak negative correlation was found between mature oocytes vs. BMI (r = -0,214; p<0.001) and oocyte maturation rate (r = -0.126; p=0.029).
Conclusion: Comparing normal BMI and obesity, there is a statistically significant decrease of aspirated and mature oocytes, suggesting lower response to ovarian stimulation in obesity. In addition, the negative correlation found corroborates the hypothesis that obesity has negative impact on ovarian stimulation after IVF.
1ANDROFERT - Centro de Referência para Reprodução Masculina - Campinas - SP
Objective: Pregnancy failure after euploid blastocysts transfer reinforces the involvement of multiple factors in embryo implantation. Recent studies suggest that blastocysts with a high number of mitochondrial DNA copies (mtDNA) have a lower implantation potential, but there is no consensus. This study aimed to determine the predictive value of mtDNA in euploid blastocysts implantation potential.
Methods: The mtDNA was quantified in 125 euploid blastocysts, previously submitted to new generation sequencing (NGS). A score based on mitochondrial DNA / genomic DNA ratio (Low-Intermediate-High) was applied. Only single euploid blastocyst transfer cycles were included.
Results: Among blastocysts with a High score (n=17), 4 (23.5%) resulted in ongoing pregnancy. On the other hand, the rates for euploid blastocysts with Low-score (n=82) and Intermediate (n=26) were 51.2% (42/82) and 50.0% (13/26), respectively (p=0.0399). Logistic regression showed that trophectoderm quality (Exp (B) = 0.425, p=0.023) and mtDNA (Exp (B) = 3.596, p=0.042) were predictors of implantation. The positive predictive value of the ongoing pregnancy rate was 64.2% and the negative predictive value was 72.4%, with sensitivity of 72.9% and specificity of 63.6%.
Conclusion: Regardless of the mechanism that explains the non-intuitive deleterious effects of excess mtDNA, a combination of high-accuracy measurement technique, adequate data adjustment and the establishment of a score seem critical in determining mtDNA role as a valid biomarker.
1Vida Bem Vinda Clinic, Department of Reproductive Medicine, São Paulo, Brazil.
Objective: State of the art vitrification techniques opened possibilities in IVF, allowing think ovarian stimulation and endometrium preparation separately. Two variables could be rethought: suppression of LH surge and phase of stimulation. It is known that synthetic progestins are effective suppressors of LH surge, but previously not used for this purpose due to endometrial effect. Recently, some authors proposed a progestin primed ovarian stimulation as an alternative to suppress the LH. Progestins have advantages of being oral, inexpensive and accessible, but concerns remain about quality/quantity of oocytes. As embryos will not be transferred, another advantage is starting stimulation according to follicular waves.
Methods: In this retrospective cohort study, we recruited all patients submitted to progestin primed ovarian stimulation (PPOS) between January and December 2017 and followed them until embryo transfer, if applicable. There was no loss of follow up for the main outcome. After providing informed consent, we obtained 100 cycles from 79 patients who were divided in two groups according to the phase of menstrual cycle that they initiated treatment. Matching criteria were patients who underwent PPOS treatments at VidaBemVinda Clinic in 2017. The follicular phase group had 32 subjects and 39 cycles and the luteal phase group had 47 subjects and 61 cycles. Both groups were homogenous for age, BMI, smoking habits, time and causes of infertility and ovarian reserve tests (FSH, AFC and AMH). The assays were performed using R software 3.5.1 and a 5% significance level was considered.
Results: The mean number of oocytes collected was 7.9 (SD:5.7) in follicular phase group versus 9.6 (SD:5.7) in luteal phase. Although the absolute numbers were higher there was no statistically significant difference between groups (p = 0.21). We also noticed that in the luteal phase group, the duration of stimulation (11.3 vs 12.2 days p=0.04) and dose of FSH were significantly higher (2718 U vs 2233 U p=0.001). The blastulation rate was also slightly better in the luteal phase group ( 24.2% x 35.7% p=0.046) , but the mean clinical pregnancy rate was similar between groups (54.2% x 51.7% p=0.9).
Conclusion: The use of follicular/luteal phase progestin primed ovarian stimulation is simple, inexpensive and flexible. This protocol could be used in almost all IVF/ICSI cycles, from poor responders to oocyte donation and fertility preservation. Luteal phase could possibly have better ovarian response, however the difference was not statistically significant in this study.
1Clinifert - Centro de reprodução Humana - Florianópolis- Brazil.
Objective: To assess the influence of Body Mass Index (BMI) and age in the ovarian response, fertilization rate, embryo development rate and pregnancy rate of patients who undergo IVF treatments.
Methods: Data from 966 patients who underwent IVF treatment from January 2013 to March 2019 were analyzed. Patients were categorized according to BMI in groups 1 to 4 (Underweight, Normal weight, Overweight and Obesity) and age in groups A, B and C (18-34 years old, 35-39 years old and over 40 years old). BMI and age impact were analyzed both separately and together.
Results: Group 2 presented higher number of oocytes retrieved and lower fertilization rate than group 4, and higher cleavage rate than group 3. Group A had more oocytes and better pregnancy rate than groups B and C, and lower cleavage and embryo development rates than group B. Group A2 had a higher number of oocytes than group A4 and higher cleavage rates than group A3. Group B showed no statistical difference amongst the BMI groups for the assessed parameters. Group C4 had less oocytes comparing to the other BMI groups, and lower pregnancy rate than groups C1 and C2.
Conclusion: Overall, a higher BMI is associated with a lower number of oocytes retrieved and cleavage rate. The fertilization rate remain comparable between the groups, when analyzing both BMI and age. Specially in women over 40 years old, weight influences the outcomes of number of oocytes retrieved and pregnancy rates.
1Clinica Conceber - Centro de Medicina Reprodutiva
Objective: Excessive use of gonadotropins for ovarian stimulation in IVF may be detrimental to the success of the procedure by increasing the rate of spontaneous abortion and decreasing the rate of pragnancy, further influencing embryonic aneuploidies. The objective of this study was to correlate the total dose of gonadotropins used with the number of euploid embryos after biopsy.
Methods: A total of 203 patients were analyzed between February 2012 and March 2019, who underwent PGT-A. Patients were divided into two groups: <3000 IU and >3000 IU, Was evaluating age, dose, mean of days of stimulation, number of biopsied embryos and average number of embryos biopsied in each group. Data were analyzed using the Chi-square, Fisher and Mann Whitney tests.
Results: The dose of gonadotrophin (2114.5±537.3UI and 3511.9±319.4UI) (p=0.0001), the days of stimulation (10.1±1.7 and 12.8±2.1) (p=0.001), the number of biopsied embryos (531 x 86), (p=0.05) and the mean number of euploid embryos (1.2±1.3 and 0.5±0.6) ( p=0.001) show a significant difference between the groups. However, no difference was observed when we analyzed the number of euploid embryos (p=0.18) and age of the patients (37.4±3.9) (p<0.05).
Conclusions: Based on the results obtained, we can conclude that the dose has no relation to the result of embryonic ploidy, however our work shows that patients receiving >3000 IU of gonadotropin is 2.3 times more likely to have no euploid embryo.
1Conceber Centro de Medicina Reprodutiva - Curitiba, PR
Objective: Evaluate if the dosage of AMH has a quantitative relationship with the number of eggs aspirated in different age groups.
Methods: A total of 733 patients were analyzed between 2014 and 2019, divided into 2 groups of AMH values: <1.2 and >1.2. and by age group: ≤35 years, 36 to 39 years and ≥40 years. The number of retrieved eggs in each group and the number of eggs per age group within each group were compared.
Results: The mean number of retrieved eggs in each group of AMH was, respectively, 3±3,1 and 10±6,8, (p=<0,0001). When comparing the number of retrieved eggs per age group, we found respectively for the group <1.2: 5±3.81, 3±2.78 and 2±2.4 and for the group >1.2: 11±7.7, 9±5.8 and 9±6.1. Showing significance among all age groups among the AMH groups. It was also significant when we compared the mean value of AMH dosage by age groups (p = <0.0001).
Conclusion: Collaborating with the literature, AMH can be used as an efficient biomarker to determine, in addition to the ovarian reserve, the expected number of eggs in IVF treatments, therefore, women with AMH >1.2 have a significantly higher number of eggs aspirated per procedure and that maternal age, directly linked to the value of AMH, has an influence on the number of aspirated eggs.
1Conceber Centro de Medicina Reprodutiva - Curitiba, PR
Objective: The objective of this study was to retrospectively evaluate blastocyst formation and pregnancy rate in different age groups.
Methods: Between 2012 and 2015, 363 fresh-cycle patients with oocytes and their own semen, without PGT-A, were divided into 3 groups (≤34a, 35 to 39a and ≥40a), comparing the results obtained when embryos were transferred at cleavage stage or blastocyst. The following rates were compared: Blastocyst formation x age group; blastocyst culture x age group and clinical pregnancy x embryonic development by age group. For the statistical analysis we used Fisher's exact test.
Results: The rate of blastocyst formation for the groups "≤34a", "35 to 39a" and "≥40a" was 30.5, 21.3 and 0% respectively. 71.4% of the patients who obtained blastocyst were up to 34 years old, while 28.6% were patients between the ages of 35 and 39 years. Blastocysts were not obtained in patients over 40 years of age. Blastocyst transfer results significantly in more clinical pregnancies when compared to embryos at the cleavage stage, regardless of the age range (61.4 and 35.3%) ≤34a (38.5 and 15.9%) from 35 to 39a and (0 and 10%) ≥40a, respectively.
Conclusion: Transfer of embryos at the blastocyst stage showed better pregnancy outcomes when compared to embryos at the cleavage stage. Collaborating with the literature, the younger the patient, the higher the rate of blastocyst formation.
1Clínica La Vitta - Centro de Reprodução Humana do Amazonas, Manaus-AM.
Objective: For the success of embrionary development in vitro, several parameters should be analysed and taken into consideration by the embryologist in charge. Establishing the culture media is one of the important aspects so that the well succeed development happens and positive results are obtained. Assuming that the importation of means does not guarantee the reliability concerning to their conservation due to severe changes of temperature during the transportation, this study aimed to realize a retrospective analysis of the rates of embrionary development from zigotto to blastocyst
Methods: Conducted at the Centro de Reprodução Humana do Amazonas - La Vitta from 2016 to 2018 with the exclusive use of the bicarbonate buffer system GVBLAST(tm) by the national company INGÁMED®.
Results: During this time 875 oocyts were aspirated, 716 oocytes (81.82%) in metaphase II were submitted to the technic of Sperm Intracytoplasmic Injection. The rate of fertilization was 76.11%, the rate of cleavage was 92.47% and the rate of formation of blastocyst was 53.21%.
Conclusion: It was aimed to prove that using national culture media it is possible to obtain successful embrionary development and also compatible with the data offered by literature. Thus, bringing quality products just like the international ones as well as better cost-benefit advantages to the routine of the laboratory and also valuing the technology developed in the country.
Objective: To develop and validate a protocol for the evaluation of sperm DNA fragmentation.
Methods: A protocol was developed using the Sperm Chromatin Dispersion (SCD) technique, consisting of a denaturation solution and two lysis solutions. For its validation, it was realized a test with four normozoospermic samples, as one aliquot of each sample was exposed to a solution of hydrogen peroxide (0.5%) for 1 hour to induce fragmentation (Test A - positive control). Submission to the fragmentation test was performed with a fraction of fresh samples concomitant with the positive control, laid up on pre-treated slides with agarose, and stained with Quick Panoptic Kit (Laborclin®). To validate the results, a comparison of this protocol with the Halosperm kit (Halotech®) (Test B) was performed, using two normozoospermic samples. For all the tests, the analyzes were done under optical microscope (1000x), counting 100 spermatozoa, determining the percentage of spermatozoa with fragmented DNA according to the size of the halo observed.
Results: In Test A, it was obtained an average of 20% spermatozoa with fragmented DNA in the fresh sample, and the positive control an average of 95%. In test B, the first and second samples tested with both kits obtained, respectively, 9.8% and 21% of sperm DNA fragmentation with Halosperm, and 9.5% and 24% with the proposed protocol.
Conclusion: The developed protocol presents equivalence with the commercial kit, and the result of the positive control shows its efficacy, therefore, it can be used in the laboratory routines.
1Purdue University, West Lafayette, IN
2Invitra, Assisted Reproductive Technologies Ltd., Ribeirão Preto, Brazil
Objective: Amount of lipids in oocyte/embryo are low and difficult to be detected. Multiple reaction monitoring profiling (MRM-Profiling) is a strategy for exploratory analysis of small molecules intended to improve the speed and efficiency of discovery and screening of metabolites and lipids based on functional group profiling. By using class-related fragments and fast MRM scans, MRM-profiling provides specific and sensitive lipid profiling. Here we report the feasibility of using this approach for quantitative analysis in embryology.
Methods: Bovine-derived oocytes and blastocysts produced in vitro were processed by the Bligh-Dyer method. Both polar and lipid phases were dried and then dissolved in acetonitrile/methanol/300 mM ammonium acetate (3:6.65:0.35, v/v/v) containing isotopically labeled lipid standards (Avanti SPLASH®_LIPIDOMIX®) for quantitative analysis. The lipid class screening methods in oocytes and embryos were taken as previously described (J Mass Spectrom .2018;53(12):1247-1252).
Results: Each sample was interrogated for 369 MRMs using two sample injections to obtain at least 10 scans per MRM. Classes of lipids quantified in the samples included glycerophosphocholine (PC), glycerophosphoethanolamine (PE), glycerophosphoglycerol (PG), sphingomyelin (SM), and triacylglycerol lipids (TAG). When the sum of ion intensity of transitions in each class was normalized to the spiked internal standards, coefficients of determination (R2) were 0.997, 0.999, 0.9790, 0.997, and 0.943 for PC, PE, PG, SM, and TAG,respectively. MRM-Profiling experiments, with or without adding internal standard, provided similar results on experimental group discrimination and most significant MRMs.
Conclusion: Relative and absolute quantification of lipids detected in oocytes and embryos provided similar outcomes for principal component analysis and ANOVA.