Cumulus cell microRNA expression when LH is added to the ovarian stimulation protocol: a pilot study.

RESEARCH QUESTION: Recombinant FSH administration in ovarian stimulation for IVF is a standard procedure, whereas the role of LH is controversial. MicroRNAs (mRNA) are small endogenous non-coding transcripts that are involved in the regulation of many cellular processes, including foliculogenesis and gonadotrophin function. The aim was to study the possible role of miRNA in ovarian follicular development in groups having different ovarian stimulation protocols. Are there different miRNA expression profiles in cumulus cells of infertile women undergoing IVF? What are the regulated pathways? DESIGN: This prospective observational study included 13 patients who fulfilled the following inclusion criteria: younger than 38 years of age; a tubal infertility factor; a male factor; or idiopathic infertility. This is a pilot study in which the patients were aleatory enrolled into two groups: seven in FSH group (recombinant FSH, 225 IU) and six in FSH plus LH group (recombinant FSH, 150 IU + recombinant LH, 75 IU). The granulosa cells obtained from the follicular ovarian retrieval were analysed using polyerase chain reaction. Results were analysed using DIANA Tools, an online bioinformatics tool. RESULTS: Among the 84 microRNAs evaluated, 11 were differentially expressed between the groups, all of which were upregulated in the FSH plus LH group, compared with the FSH group. Differentially expressed miRNA profiles are related to oestrogen signalling, oocyte meiosis and pluripotent cells regulation. CONCLUSION: miRNA overexpression in the FSH plus LH group is consistent with the independent and fundamental role of LH in folliculogenesis, leading to a distinct molecular response between groups.

A Single Dose of a Hybrid hAdV5-Based Anti-COVID-19 Vaccine Induces a Long-Lasting Immune Response and Broad Coverage against VOC.

Most approved vaccines against COVID-19 have to be administered in a prime/boost regimen. We engineered a novel vaccine based on a chimeric human adenovirus 5 (hAdV5) vector. The vaccine (named CoroVaxG.3) is based on three pillars: (i) high expression of Spike to enhance its immunodominance by using a potent promoter and an mRNA stabilizer; (ii) enhanced infection of muscle and dendritic cells by replacing the fiber knob domain of hAdV5 by hAdV3; (iii) use of Spike stabilized in a prefusion conformation. The transduction with CoroVaxG.3-expressing Spike (D614G) dramatically enhanced the Spike expression in human muscle cells, monocytes and dendritic cells compared to CoroVaxG.5 that expressed the native fiber knob domain. A single dose of CoroVaxG.3 induced a potent humoral immunity with a balanced Th1/Th2 ratio and potent T-cell immunity, both lasting for at least 5 months. Sera from CoroVaxG.3-vaccinated mice was able to neutralize pseudoviruses expressing B.1 (wild type D614G), B.1.117 (alpha), P.1 (gamma) and B.1.617.2 (delta) Spikes, as well as an authentic P.1 SARS-CoV-2 isolate. Neutralizing antibodies did not wane even after 5 months, making this kind of vaccine a likely candidate to enter clinical trials.

Delivering embryos following 10 years of cryopreservation, using unpaired freeze/thaw techniques: A case report.

Although frozen embryo transfer is a widely established route for assisted reproduction, successful frozen embryo transfer using embryos that have undergone long term cryopreservation remains relatively unexplored, and its efficacy remains a matter of some debate. This case report describes two successful frozen embryo transfer conceptions in the same patient, one after 3 months of cryopreservation and the second 10 years after cryopreservation. These embryos were cryopreserved using the slow freezing technique and were thawed using an unpaired technique (ultra-rapid warming) after 10 years of storage.

Extracellular matrix metalloproteinase expression in endometrial tissue after arterial embolization of myomas.

OBJECTIVES: Arterial embolization of myomas (AEM) is controversial because of the changes that occur in the extracellular matrix (ECM) of the endometrium and its effect on gestational success in infertile patients desiring reproductive capability. Therefore, we performed this study on the expression of genes in the ECM of the endometrium, such as those coding metalloproteinases (MMP), before and 6 months after embolization of the uterine arteries. METHODS: Seven women with leiomyomas were evaluated, and MMP3 and MMP10 levels were measured. The women underwent pelvic nuclear magnetic resonance (NMR), examination, and endometrial biopsy between the 20th and 24th day of the menstrual cycle, and pre- and post-AEM (after 6 months). For data analysis, the Cq comparative method, also known as the 2-ΔΔCT method, was used to calculate the relative quantities of MMP gene expression among the samples collected. RESULTS: There was a significant decrease by 9.52 times in the expression of MMP3 (p=0.007), and a non-significant change in the expression of MMP10 (p=0.22) in post-AEM-treated women than pre-AEM-treated women. CONCLUSIONS: The results suggest that ECM continues to undergo tissue remodeling 6 months after AEM, at least with regard to MMP3 expression, suggesting that AEM affects the ECM for at least 6 months after the procedure.

Extracellular matrix metalloproteinase expression in endometrial tissue after arterial embolization of myomas

OBJECTIVES: Arterial embolization of myomas (AEM) is controversial because of the changes that occur in the extracellular matrix (ECM) of the endometrium and its effect on gestational success in infertile patients desiring reproductive capability. Therefore, we performed this study on the expression of genes in the ECM of the endometrium, such as those coding metalloproteinases (MMP), before and 6 months after embolization of the uterine arteries. METHODS: Seven women with leiomyomas were evaluated, and MMP3 and MMP10 levels were measured. The women underwent pelvic nuclear magnetic resonance (NMR), examination, and endometrial biopsy between the 20th and 24th day of the menstrual cycle, and pre- and post-AEM (after 6 months). For data analysis, the Cq comparative method, also known as the 2-ΔΔCT method, was used to calculate the relative quantities of MMP gene expression among the samples collected. RESULTS: There was a significant decrease by 9.52 times in the expression of MMP3 (p=0.007), and a non-significant change in the expression of MMP10 (p=0.22) in post-AEM-treated women than pre-AEM-treated women. CONCLUSIONS: The results suggest that ECM continues to undergo tissue remodeling 6 months after AEM, at least with regard to MMP3 expression, suggesting that AEM affects the ECM for at least 6 months after the procedure.

The insulin signaling pathway is dysregulated in cumulus cells from obese, infertile women with polycystic ovarian syndrome with an absence of clinical insulin resistance.

METHODS: This is a cohort study, conducted at a university-based reproductive medicine center and private reproductive medicine center that aimed to evaluate granulosa cumulus cell gene expression in the insulin signaling pathway in Polycystic Ovary Syndrome (PCOS) patients undergoing in vitro fertilization (IVF) treatment and to compare the cumulus gene expression between normal weight and obese women without clinical insulin resistance. Fifteen PCOS patients, nine normal weight patients and six obese patients presenting normal HOMA IR (Homeostasis Model Assessment-Insulin Resistance), participated. Patients underwent oocyte retrieval for IVF and after the procedure, granulosa cumulus cells were removed from the oocytes for RNA extraction. Quantitative polymerase chain reaction (PCR) array analysis of 84 genes from insulin signaling pathway was conducted. The results were expressed as fold up- or fold down-expression in obese patients compared with normal weight patients. Any fold change ⩾3 or ⩽3 and any p ⩽ 0.05 were considered statistically significant. RESULTS: There were 10 genes that were overexpressed in obese compared with normal weight women, BCL2L1, BRAF, CBL, DOK1, FBP1, FRS2, MTOR, PCK2, RPS6KA1, and SORBS1, that had a fold change ⩾3 and p ⩽ 0.05. DISCUSSION: In the obese group, the overexpressed genes are mainly responsible for the proliferation and differentiation of cumulus cells during oocyte maturation, insulin resistance, apoptosis regulation, and glucose metabolism during early embryogenesis, suggesting that in the follicular environment, insulin resistance is present even in the absence of clinical signs. CONCLUSION: Together, our findings and the related literature suggest that those alterations may be associated with the worse prognosis of follicular development and oocyte maturation observed in PCOS obese women.

Progesterone's role in deep infiltrating endometriosis: Progesterone receptor and estrogen metabolism enzymes expression and physiological changes in primary endometrial stromal cell culture.

To study progesterone signaling activation, we measured changes in extracellular pH as a reflection of Na+/H+ exchange (NHE) using a cytosensor microphysiometer and assessed progesterone receptor (PR) and estrogen metabolism enzymes mRNA expression in cultured endometrial cells from women with deep infiltrating endometriosis and healthy controls using real-time quantitative PCR. This study was conducted at a University hospital and included patients with and without deep infiltrating endometriosis (DIE). Primary endometrial stromal cells (ECs) from women with DIE and controls were treated with 17ß-estradiol and progesterone prior to microphysiometer measurements and qPCR evaluations. Decreased progesterone responsiveness and decreased total nuclear PR and HSD17B1 mRNA expression were observed in cultured ECs from women with deep infiltrating endometriosis relative to those from control samples before and after hormone treatment. These cells also showed increased 17ß-hydroxysteroid dehydrogenases types 2 (HSD17B2) relative to control group and increased expression of aromatase (CYP19) after exposure to progesterone. These physiological and expression patterns observed in ECs cultures from women with DIE reinforces previous findings in the literature supporting the progesterone resistance hypothesis in the pathogenesis of endometriosis.

A review of initial data on pregnancy during the COVID-19 outbreak: implications for assisted reproductive treatments

The current outbreak of the novel 2019 coronavirus disease (COVID-19) started in China in December 2019 and has since spread to several other countries. On March 25, 2020, a total of 375,498 cases had been confirmed globally with 2,201 cases in Brazil, showing the urgency of reacting to this international public health emergency. While in most cases, mild symptoms are observed, in some cases the infection leads to serious pulmonary disease. As a result, the possible consequences of the COVID-19 outbreak for pregnant women and its potential effects on the management of assisted reproductive treatments, demand attention. In this review, we summarize the latest research progress related to COVID-19 epidemiology and the reported data of pregnant women, and discuss the current evidence of COVID-19 infections during pregnancy and its potential consequences for assisted reproductive treatments. Reported data suggest that symptoms in pregnant women are similar to those in other people, and that there is no evidence for higher maternal or fetal risks. However, considering the initial data and lack of comprehensive knowledge on the pathogenesis of SARS-CoV-2 during pregnancy, human reproduction societies have recommended postponing the embryo transfers and do not initiate new treatment cycles. New evidence must be considered carefully in order to adjust these recommendations accordingly at any time and to guide assisted reproductive treatments.(AU)

The impact of balanced reciprocal translocation - 46,XX,t(7;17) (p13;q24) probably involving the SOX9 gene in the in vitro fertilization with own oocytes evaluated by preimplantation genetic testing or donated oocytes.

Preimplantation genetic testing (PGT) for in vitro fertilization (IVF) - also known as PGT for Structural Rearrangements (PGT-SR) - has emerged as an option for at-risk couples carrying balanced translocations. The female in the couple featured in this case report is a carrier of a balanced reciprocal translocation who underwent IVF. PGT showed all her embryos were aneuploid. She subsequently had two cycles using donor oocytes, which ended in miscarriages.

Egg donation of vitrified oocytes bank produces similar pregnancy rates by blastocyst transfer when compared to fresh cycle.

PURPOSE: Advances in reproductive techniques, mainly the introduction of oocyte vitrification, have provided the opportunity to conceive from oocyte banks. The aim of this study was to compare the clinical outcomes of fresh and vitrified oocytes in an egg donation program following blastocyst transfer. METHODS: This retrospective observational study included 504 oocyte donation cycles. All donor women were younger than 30 years of age. The recipient cycles were divided into two groups: fresh oocytes (n = 78) or vitrified oocytes (n = 426). All oocytes were fertilized by ICSI using ejaculated sperm, followed by blastocyst transfer. Endometrium preparation was performed with estradiol valerate plus micronized progesterone according to standard protocols. RESULTS: Recipients were of similar age (fresh 42.0 ± 4.5 years vs vitrified 41.8 ± 4.8 years; p = 0.790). The fresh group received more mature oocytes for injection compared to the vitrified group (10.1 ± 2.8 vs 9.2 ± 2.2; p = 0.005). The two pronuclei (2PN) rate (74.5 vs 77.4%; p = 0.195) and blastocyst rate (48.8 vs 51.6%; 0.329) were similar between the fresh and vitrified groups, respectively. The rates of clinical pregnancy were 60.9% in the fresh and 59.0% in the vitrified groups (p = 0.771). CONCLUSIONS: Our findings suggest that vitrified oocytes result in similar pregnancy rates when compared to fresh oocytes with blastocyst transfer in an egg donation program. Moreover, vitrified oocytes may allow for a better cycle schedule, starting with a lower number of oocytes to be fertilized. Therefore, we hypothesize that egg banks with vitrified oocytes could be safely utilized in an egg donation program.

Does slow embryo development predict a high aneuploidy rate on trophectoderm biopsy?

The aneuploidy rates in expanded blastocysts biopsied on days 5 and 6 development were assessed in women undergoing IVF followed by array comparative genomic hybridization. This study included 1171 expanded blastocysts from 465 patients. Among the 465 patients, 215 and 141 underwent embryo biopsy on day 5 and day 6 (46.2% and 30.3%, respectively), and 109 underwent biopsy on both days 5 and 6 (23.4%). The cycles of 206 women were cancelled because only aneuploidy embryos were present (44.3%). The aneuploid embryos were classified according to the type as single, double or complex aneuploidy. No differences were observed in the distributions of these three categories according to the day of the biopsy. The aneuploidy rate was also evaluated according to maternal age, and was found to be higher in older patients; however, no differences in this rate were detected between embryos biopsied on days 5 and 6 according to maternal age. Biopsy was carried out when blastocysts reached the expanded stage. The embryos biopsied on day 6 had a higher rate of aneuploidy (69.9%) than those biopsied on day 5 (61.4%); however, the euploid embryos transferred had similar chances for successful and healthy gestation.

Analysis of Gene Expression in the Endocervical Epithelium of Women With Deep Endometriosis.

Endometriosis affects approximately 12% of reproductive-age women and is currently diagnosed using invasive laparoscopic surgery. Differences in gene expression in the eutopic endometrium between women with and without endometriosis have been reported, and determining the reproducibility of these genetic differences in the endocervical epithelium would represent an important step toward developing novel diagnostic strategies. In this study, we analyzed gene expression in the endocervical epithelium in women with and without moderate or severe endometriosis. Using RT2 Profiler PCR Arrays, we analyzed gene expression in endocervical epithelial cells from women with deep endometriosis (n = 4) and healthy women (n =6). Nine genes were identified as being upregulated: 5 cell cycle genes (cyclin B1 [CCNB1], cyclin G1 [CCNG1], cullin 1 [CUL1], general transcription factor IIH, polypeptide 1 [GTF2H1], and proliferating cell nuclear antigen [PCNA]), 3 cytokine genes (C3, chemokine (C-C motif) ligand 21 [CCL21], and chemokine (C-X-C motif) ligand 14 [CXCL14]) and 1 gene related to dendritic cell pathways (ICAM2), showing that differential gene expression is present in the endocervical epithelium of women with deep endometriosis.

Matrix metalloproteinases 2 and 9 and e-cadherin expression in the endometrium during the implantation window of infertile women before in vitro fertilization treatment.

OBJECTIVE: To evaluate the expression of endometrial matrix metalloproteinases (MMPs) 2 and 9 and E-cadherin in peri-implantation phase of infertile women who have undergone in vitro fertilization (IVF) cycles. METHODS: This prospective study included 51 patients who underwent endometrial biopsy during the receptive phase in a menstrual cycle prior to IVF treatment. The samples were evaluated by tissue microarray for immunohistochemical study. RESULTS: The expression of MMP-2, MMP-9, and E-cadherin in the endometrium prior to IVF treatment was not associated with pregnancy. There was a decrease in E-cadherin immunodetection, the higher the age of the patients, a negative relationship between E-cadherin and MMP-2, and a positive association between MMP-9 and E-cadherin. CONCLUSIONS: The MMP-2, MMP-9, and E-cadherin are expressed in the endometrium of infertile patients during the receptive phase of the natural menstrual cycle. However, there is no correlation between the expression of these molecules and the clinical IVF outcomes.

Letrozole Therapy for Obstructive Azoospermic Men before in vitro Fertilization (IVF) treatment with Percutaneous Epididymal Sperm Aspiration.

OBJECTIVE: The aim of this study was to report our preliminary experience regarding the use of letrozole in men with obstructive azoospermia (OA) undergoing percutaneous epididymal sperm aspiration (PESA) for in vitro fertilization treatment using intracytoplasmic sperm injection (ICSI), who had a very low sperm recovery upon PESA and unsuccessful ICSI. Our hypothesis was that letrozole therapy could improve testicular function by increasing serum gonadotropins and T levels, stimulate testicle germ cells and, most importantly, that it enhanced the motile sperm count at a second attempt. METHODS: We report on our preliminary experience with letrozole therapy in 11 men with OA, who failed to achieve pregnancy in the first PESA-ICSI and did not have spermatozoa cryopreserved for a second attempt. The patients received 3 months of letrozole at 2.5mg/day and underwent PESA-ICSI after 6.1±3.8 months. The patients were 48.6 ± 9.6 years old, and underwent at least two PESA procedures. We evaluated the total motile sperm count per PESA samples, as the increases in serum FSH, LH, and T levels after treatment. RESULTS: All parameters increased significantly at 3 months following letrozole therapy for most patients. The total motile sperm count increased from 100 to 500% compared to the first PESA. CONCLUSION: Letrozole can be considered a reliable treatment to improve sperm recovery for men with OA undergoing PESA-ICSI cycles by increasing serum gonadotropins and testosterone (T) levels, and-most importantly-the motile sperm count.

Intrafollicular soluble receptor for advanced glycation end products (sRAGE) and embryo quality in assisted reproduction.

The developmental potential of human embryos has important implications in assisted reproduction and depends, among other factors, on oocyte competency. The receptor for advanced glycation end products (RAGE) is a member of the superfamily of immunoglobulin cell-surface molecules that are constitutively expressed during embryonic development. RAGE is down-regulated in homeostasis in adult life. This study measured the concentration of soluble RAGE (sRAGE) in follicular fluid obtained from the leading follicle after ovarian stimulation of 54 women undergoing intracytoplasmic sperm injection. Corresponding embryos and sRAGE concentrations in follicular fluid were evaluated and correlations were investigated by multi-adjusted regression analysis. High intrafollicular sRAGE concentrations predicted poor-quality embryos (n=45, OR=0.986; P=0.026), adjusted for patient age, body mass index and oocyte quality, showing an inverse association between intrafollicular sRAGE concentrations and embryo development. The developmental potential of human embryos has important implications in assisted reproduction, and it depends, among other factors, on oocyte competency. The receptor for advanced glycation end products (RAGE) is a molecule constitutively expressed during embryonic development, but it is down-regulated in adult life. RAGE is frequently associated with pro-inflammatory responses, and it is implicated as an underlying condition in immune disorders, cardiovascular diseases, diabetes, Alzheimer's disease and cancer. In addition to activating the pro-inflammatory responses, RAGE down-regulates cellular defence mechanisms. The present study measured the concentrations of soluble RAGE (sRAGE) in follicular fluid samples obtained from leading follicles of women undergoing intracytoplasmic sperm injection (ICSI). This prospective cohort study included 54 patients undergoing ICSI, and follicular fluid samples were obtained from the leading follicle after ovarian stimulation. The corresponding embryos were evaluated and correlations with intrafollicular sRAGE concentrations were investigated using multi-adjusted regression analysis. We observed that high intrafollicular concentrations of sRAGE predicted poor embryo quality. Our findings suggest an association between high concentrations of intrafollicular sRAGE and poor embryo development following ovarian stimulation for ICSI.

Opinions concerning pre-implantation genetic diagnosis and sex selection among gynecologist-obstetricians in Brazil.

OBJECTIVE: The aim of this study was to assess a sample of obstetrician-gynecologists on their technical knowledge of PGD as well as their views of PGD and sex selection (SS). STUDY DESIGN: Cross-sectional observational study, carried out during a Congress on Gynecology and Obstetrics. A self-report questionnaire was offered to all professionals registered to attend the Congress and answers of 723 physicians were analyzed. RESULTS: Of the participants, 436 (60.3%) were female and 287 (39.7%) were male; 517 (71.5%) had children. Regarding knowledge of PGD techniques, 63.2% had heard something about PGD and/or preconception SS. Concerning SS, physicians believed that SS is always the couple's choice (36.4%), should be the couple's choice only in specific situations (42.6%), or did not agree that SS should be a couple's choice (17.4%). A majority of the respondents were in favor of PGD and believed that pre-embryos with 6-8 cells are human life in potency. CONCLUSIONS: In spite of a small percentage of gynecologists-obstetricians in Brazil had knowledge of PGD techniques, many of them recognized it to be a method to prevent genetic diseases and agreed with its use. The SS appears to be highly accepted for use in specific situations.

Sperm banking for male cancer patients: social and semen profiles.

PURPOSE: Report the characteristics of cryopreserved semen from a cohort of male cancer patients, attitudes towards cryopreservation and outcomes of semen samples based on a 12-year cryopreservation program. MATERIAL AND METHODS: Data from 98 male cancer patients whose sperm samples were banked were evaluated. Demographic parameters, semen characteristics, destination of sperm banked samples and questionnaires answered by the patients regarding cryopreservation time were evaluated. RESULTS: The cancer diagnoses were testicle (56.1%), prostate (15.3%), Hodgkin's lymphomas (9.2%), non-Hodgkin's lymphomas (7.1%), leukemia (3.1%) and other malignancies (9.2%). The patients with testicular cancer presented lower sperm concentration (p < 0.001); however, there were no differences with the percentage of normozoospermic patients among cancer type groups (p = 0.185). A shorter time between cancer diagnosis and sperm banking was observed for testicular and prostate cancer patients (p < 0.001). Most of the patients (89.5%) favored sperm banking as a fertility preservation method. CONCLUSIONS: Although less than 20% of banked sperm samples were disposed of, the majority of patients related sperm banking with safe for fertility preservation. Our results show that all male cancer patients of reproductive age facing cancer treatment could be offered sperm banking.

Sperm banking for male cancer patients: social and semen profiles

PURPOSE: Report the characteristics of cryopreserved semen from a cohort of male cancer patients, attitudes towards cryopreservation and outcomes of semen samples based on a 12-year cryopreservation program. MATERIAL AND METHODS: Data from 98 male cancer patients whose sperm samples were banked were evaluated. Demographic parameters, semen characteristics, destination of sperm banked samples and questionnaires answered by the patients regarding cryopreservation time were evaluated. RESULTS: The cancer diagnoses were testicle (56.1 percent), prostate (15.3 percent), Hodgkin’s lymphomas (9.2 percent), non-Hodgkin’s lymphomas (7.1 percent), leukemia (3.1 percent) and other malignancies (9.2 percent). The patients with testicular cancer presented lower sperm concentration (p < 0.001); however, there were no differences with the percentage of normozoospermic patients among cancer type groups (p = 0.185). A shorter time between cancer diagnosis and sperm banking was observed for testicular and prostate cancer patients (p < 0.001). Most of the patients (89.5 percent) favored sperm banking as a fertility preservation method. CONCLUSIONS: Although less than 20 percent of banked sperm samples were disposed of, the majority of patients related sperm banking with safe for fertility preservation. Our results show that all male cancer patients of reproductive age facing cancer treatment could be offered sperm banking.

Tetanus and diphtheria antibodies and response to a booster dose in Brazilian HIV-1-infected women.

Tetanus and diphtheria (Td) antibodies were studied in HIV-1-infected women during puerperium. HIV group (n=61) was compared with Control group (n=101). Twenty-one women from HIV and 13 from Control group who had antibody levels lower than 0.1 IU/mL received a booster with Td vaccine. Antibodies were assessed by double antigen ELISA. Mean tetanus and diphtheria antibody levels from HIV group were lower than those from Control group. Multiple linear regression analysis showed that tetanus and diphtheria antibody levels were decreased by HIV-1-infection, and that was independent of the reduction due to the time interval between last booster and antibody assessment. After a booster dose, both groups had an increase in mean tetanus and diphtheria antibody levels, but in Control group the levels were higher than in HIV group.