Improving Implantation Rate in 2nd ICSI Cycle through Ovarian Stimulation with FSH and LH in GNRH Antagonist Regimen / Melhorando a taxa de implantação no 2° ciclo de ICSI através do estímulo ovariano com FSH e LH no regime com antagonista do GnRH

Abstract Objective To investigate whether patients with a previous recombinant follicle stimulating hormone (rFSH)-stimulated cycle would have improved outcomes with rFSH + recombinant luteinizing hormone (rLH) stimulation in the following cycle. Methods For the present retrospective case-control study, 228 cycles performed in 114 patients undergoing intracytoplasmic sperm injection (ICSI) between 2015 and 2018 in an in vitro fertilization (IVF) center were evaluated. Controlled ovarian stimulation (COS) was achieved with rFSH (Gonal-f, Serono, Geneva, Switzerland) in the first ICSI cycle (rFSH group), and with rFSH and rLH (Pergoveris, Merck Serono S.p.A, Bari, Italy) in the second cycle (rFSH + rLH group). The ICSI outcomes were compared among the groups. Results Higher estradiol levels, oocyte yield, day-3 high-quality embryos rate and implantation rate, and a lower miscarriage rate were observed in the rFSH + rLH group compared with the rFSH group. In patients < 35 years old, the implantation rate was higher in the rFSH + rLH group compared with the rFSH group. In patients ≥ 35 years old, higher estradiol levels, oocyte yield, day-3 high-quality embryos rate, and implantation rate were observed in the rFSH + rLH group. In patients with ≤ 4 retrieved oocytes, oocyte yield, mature oocytes rate, normal cleavage speed, implantation rate, and miscarriage rate were improved in the rFSH + rLH group. In patients with ≥ 5 retrieved oocytes, higher estradiol levels, oocyte yield, and implantation rate were observed in the rFSH + rLH group. Conclusion Ovarian stimulation with luteinizing hormone (LH) supplementation results in higher implantation rates, independent of maternal age and response to COS when compared with previous cycles stimulated with rFSH only. Improvements were also observed for ICSI outcomes and miscarriage after stratification by age and retrieved oocytes.

Resumo Objetivo: Investigar se há algum efeito da suplementação com hormônio luteinizante (LH, na sigla em inglês) no regime com antagonista do hormônio liberador de gonadotropina (GnRH, na sigla em inglês) sobre os resultados dos ciclos consecutivos de injeção intracitoplasmática de espermatozoides (ICSI, na sigla em inglês). Métodos Para o presente estudo retrospectivo de caso-controle, foram avaliados 228 ciclos de microinjeção intracitoplasmática de espermatozoides (ICSI, na sigla em inglês) realizados em 114 pacientes entre 2015 e 2018 em um centro privado de fertilização in vitro (FIV) afiliado a uma universidade. O estímulo ovariano controlado (EOC) foi feito com hormônio folículo- estimulante recombinante (rFSH, na sigla em inglês) (Gonal-f, Serono, Genebra, Suíça) no primeiro ciclo de ICSI (grupo rFSH), e com rFSH e rLH (Pergoveris, Merck Serono S.p.A, Bari, Itália) no segundo ciclo (grupo rFSH + rLH). Os desfechos dos ciclos de ICSI foram comparados entre os grupos. Resultados Níveis mais elevados de estradiol, de recuperação oocitária, taxa de embriões de alta qualidade no 3° dia e taxa de implantação, e menor taxa de aborto foram observados no grupo rFSH + rLH. Em pacientes < 35 anos, a taxa de implantação foi maior no grupo rFSH + rLH em comparação com o grupo rFSH. Em pacientes com ≥ 35 anos, maiores níveis de estradiol, recuperação oocitária, a taxa de embriões de alta qualidade no 3° dia e a taxa de implantação foram observados no grupo rFSH + rLH. Em pacientes com baixa resposta ao EOC (≤ 4 oócitos recuperados), a recuperação oocitária, a taxa de oócitos maduros, a taxa de velocidade normal de clivagem, a taxa de implantação e a taxa de aborto foram melhoradas no grupo rFSH + rLH. Em pacientes com resposta normal ao EOC (≥ 5 oócitos recuperados), níveis mais elevados de estradiol, recuperação oocitária e taxa de implantação foram observados no grupo rFSH + rLH. Conclusão A estimulação ovariana com suplementação de LH resultou em taxas de implantação mais altas, independentemente da idade materna e da resposta ao EOC, em comparação com os ciclos anteriores estimulados apenas com rFSH. Melhorias também foram observadas nos resultados da ICSI e na taxa de aborto quando as pacientes foram estratificadas por idade e número de oócitos recuperados.

Increasing paternal age and ejaculatory abstinence length negatively influence the intracytoplasmic sperm injection outcomes from egg-sharing donation cycles.

BACKGROUND: The paternal role in embryogenesis is much more than providing a haploid genome. The spermatozoa have crucial roles related to embryogenesis' epigenetic regulation, fusion of gametes, and cleavage, which may influence intracytoplasmic sperm injection outcomes. OBJECTIVES: To study whether paternal age, length of ejaculatory abstinence and semen parameters influence the outcomes of intracytoplasmic sperm injection in the oocyte recipients from egg-sharing donation cycles. MATERIALS AND METHODS: This historical cohort study involved 427 oocyte recipient intracytoplasmic sperm injection cycles, from 321 patients undergoing egg-sharing donation cycles from January 2015 to May 2017, in a private in vitro fertilization center. Electronic medical records of donors and recipients intracytoplasmic sperm injection cycles were reviewed. The impact of male age, length of ejaculatory abstinence, and semen parameters on recipients' intracytoplasmic sperm injection outcomes were investigated using general mixed models. Implantation, pregnancy, and live birth rates were the primary outcome measures. RESULTS: Paternal age negatively affected fertilization, day 3 top-quality embryos, day 3 normal embryo cleavage speed, blastocyst formation, high-quality blastocysts, and implantation, and was correlated with diminished pregnancy and live birth rates. Length of ejaculatory abstinence negatively influenced the rates of day 3 top-quality embryos, normal embryo development on day 3, blastocyst development, and implantation. Sperm count positively affected the rates of fertilization, blastocyst development, and implantation. The percentage of progressive sperm motility positively affected day 3 normal embryo development rate and implantation rate. Total motile sperm count positively affected blastocyst development rate and implantation rate. DISCUSSION AND CONCLUSION: Additional tracking of the influence of male characteristics on intracytoplasmic sperm injection outcomes should be stimulated. Paternal age is not manageable, but reducing length of ejaculatory abstinence could be an alternative approach to improve the outcomes of intracytoplasmic sperm injection.

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.

Obtenção de gametas seguros por meio de associação de técnicas de processamento seminal para casais sorodiscordantes para HIV / Safe gametes acquisition through association of seminal processing techniques with HIV serodiscordant couples

OBJETIVO: o objetivo deste estudo foi avaliar os parâmetros laboratoriais e clínicos de pacientes submetidos à reprodução humana assistida, associando técnicas de processamento seminal para eliminação de partículas virais da amostra de sêmen em casais nos quais o homem é infectado pelo vírus da imunodeficiência humana adquirida (HIV). MÉTODOS: foram avaliados 11 ciclos de injeção intracitoplasmática de espermatozóide (ICSI) realizados em casais nos quais os homens eram infectados pelo HIV (Grupo HIV), e 35 ciclos de ICSI nos quais se utilizaram espermatozóides doados (Grupo Controle). As amostras de sêmen dos doadores foram submetidas à análise seminal completa, processamento seminal (lavagem) e criopreservação. Os homens do Grupo HIV receberam antibioticoterapia prévia e realizou-se análise seminal, lavagem e gradiente descontínuo de densidade antes da criopreservação. As amostras foram avaliadas para carga viral e a ICSI foi realizada quando não houve detecção do HIV. RESULTADOS: quanto aos resultados da análise seminal, os grupos se mostraram comparáveis em relação à concentração e motilidade progressiva dos espermatozóides. Entretanto, a porcentagem de espermatozóides morfologicamente normais foi maior no Grupo Controle (14,3 por cento) comparado ao HIV (5,8 por cento; p=0,002). Na avaliação dos parâmetros embrionários, as taxas de fertilização normal (Controle: 74,7 por cento e HIV: 71,7; p=0,838) e de bons embriões (Controle: 42,4 por cento e HIV: 65,1 por cento; p=0,312) foram semelhantes. Por outro lado, o Grupo Controle apresentou melhores resultados clínicos que o HIV (gestação continuada: 52,9 e 12,5 por cento; p=0,054; implantação: 42,6 e 10,4 por cento; p=0,059, respectivamente), apesar de as diferenças não serem estatisticamente significantes. Após o nascimento, não houve soroconversão das mães e das crianças nascidas. CONCLUSÕES: a associação de técnicas de processamento seminal para eliminação do HIV de amostras...

PURPOSE: the propose of this study was to analyze the clinical and laboratorial parameters of patients submitted to human assisted reproduction techniques with association of sperm processing techniques, in order to remove virus particles from semen samples of couples in which men was infected by human immunodeficiency virus (HIV). METHODS: it was assessed 11 intracytoplasmatic sperm injection (ICSI) cycles from couples whose men were HIV seropositive (HIV Group), and 35 cycles in which semen donors' samples were used in ICSI procedures (Control Group). Semen samples from Control Group were submitted to routine semen analysis, sperm wash and cryopreservation. The man from HIV Group received previous antibiotic therapy; the semen samples were analyzed routinely and prepared by sperm wash and density gradient method before cryopreservation. Those samples were evaluated to viral load and ICSI was performed when no HIV was detected. RESULTS: regards to semen analysis the groups were similar to sperm concentration and progressive motility. Nevertheless, the percentage of sperm with normal morphology were higher on Control Group (14.3 percent) than HIV (5.8 percent; p=0.002). On embryo parameters assessment, the normal fertilization (CT: 74.7 percent and HIV: 71.7; p=0.838, respectively) and good embryos rate (CT: 42.4 percent and HIV: 65.1 percent; p=0.312, respectively) were comparable. On the other hand, the Control Group presented better clinic results than HIV Group (ongoing pregnancy rate: 52.9 percent versus 12.5 percent; p=0.054, and implantation rate: 42.6 versus 10.4 percent; p=0.059, respectively), however the differences were not statistically significant. After delivery, no seroconversion was observed on mother and child. CONCLUSIONS: the association of sperm processing techniques in order to remove HIV from semen samples does not influence in laboratorial parameters of assisted reproduction techniques cycles. On the other hand, ...

[Safe gametes acquisition through association of seminal processing techniques with HIV serodiscordant couples]. / Obtenção de gametas seguros por meio de associação de técnicas de processamento seminal para casais sorodiscordantes para HIV.

PURPOSE: the propose of this study was to analyze the clinical and laboratorial parameters of patients submitted to human assisted reproduction techniques with association of sperm processing techniques, in order to remove virus particles from semen samples of couples in which men was infected by human immunodeficiency virus (HIV). METHODS: assessed were 11 intracytoplasmatic sperm injection (ICSI) cycles from couples whose men were HIV seropositive (HIV Group), and 35 cycles in which semen donors' samples were used in ICSI procedures (Control Group). Semen samples from Control Group were submitted to routine semen analysis, sperm wash and cryopreservation. The man from HIV Group received previous antibiotic therapy; the semen samples were analyzed routinely and prepared by sperm wash and density gradient method before cryopreservation. Those samples were evaluated to viral load and ICSI was performed when no HIV was detected. RESULTS: with regards to semen analysis, the groups were similar to sperm concentration and progressive motility. Nevertheless, the percentage of sperm with normal morphology were higher on Control Group (14.3%) than HIV (5.8%; p=0.002). On embryo parameters assessment, the normal fertilization (CT: 74.7% and HIV: 71.7; p=0.838, respectively) and good embryos rate (CT: 42.4% and HIV: 65.1%; p=0.312, respectively) were comparable. On the other hand, the Control Group presented better clinic results than HIV Group (ongoing pregnancy rate: 52.9% versus 12.5%; p=0.054, and implantation rate: 42.6 versus 10.4%; p=0.059, respectively), however the differences were not statistically significant. After delivery, no seroconversion was observed on mother and child. CONCLUSIONS: the association of sperm processing techniques in order to remove HIV from semen samples does not influence in laboratorial parameters of assisted reproduction techniques cycles. On the other hand, it has been demonstrated excellent results getting safety gametes to serodiscordant couples.

Preimplantation genetic diagnosis of Von Hippel-Lindau disease cancer syndrome by combined mutation and segregation analysis

Von Hippel-Lindau (VHL) disease is an autosomal dominant cancer syndrome, associated with the development of tumors and cysts in multiple organ systems, whose expression and age of onset are highly variable. The VHL disease tumor suppressor gene (VHL) maps to 3p25-p26 and mutations ranging from a single base change to large deletions have been detected in patients with VHL disease. We developed a single cell PCR protocol for preimplantation genetic diagnosis (PGD) of VHL disease to select unaffected embryos on the basis of the detection of the specific mutation and segregation analysis of polymorphic linked markers. Multiplex-nested PCR using single buccal cells of an affected individual were performed in order to test the accuracy and reliability of this single-cell protocol. For each locus tested, amplification efficiency was 83 percent to 87 percent and allelic drop-out rates ranged from 12 percent to 8 percent. Three VHL disease PGD cycles were performed on cells from a couple with paternal transmission of a 436delC mutation in exon 2 of the VHL gene, leading to the identification of three unaffected embryos. Independent of the mutation present, this general PGD protocol for the diagnosis of VHL disease can be used in families informative for either the D3S1038 or D3S1317 microsatellite markers.

Processamento seminal e utilização da técnica de ICSI em casais sorodiscordantes para o HIV-1 / Semen processing and ICSI technique in HIV-1 serodiscordant couples

Com os avanços na terapia antiretroviral para indivíduos infectados pelo HIV-1, aumentou a procura das técnicas de reprodução assistida por casais sorodiscordantes que almejam ter um filho biológico saudável.A associação de técnicas para processamento seminal em homens infectados pelo HIV-1 vêm sendo utilizadas com sucesso.Esse trabalho tem como objetivo relatar três ciclos de tratamento de reprodução assistida em dois casais sorodiscordantes, nos quais o homem é infectado pelo HIV-1.Em nosso estudo sugerimos uma modificação na técnica habitualmente utilizada para casais sorodiscordantes, que consiste na utilização de antiboticoterapia prévia com intuito de reduzir o número de linfócitos e macrófagos no ejaculado, seguida da associação de técnicas para processamento seminal, sperm wash, gradiente descontínuo de densidade e swim-up.A criopreservação e validação da amostra por PCR foram realizadas possibilitando maior confiabilidade para os procedimentos de reprodução assistida, e técnica de ICSI, que não depende do número final de espermatozóides móveis obtidos, além de contribuir para a redução no risco de transmissão do virus, parece ser o método de escolha nestes casos

Importância dos parâmetros seminais nos resultados de inseminaçäo intra-uterina / Importance of seminal parameters for intrauterine insemination outcomes

OBJETIVOS: analisar a relevância dos parâmetros seminais nos resultados dos ciclos de inseminaçäo intra-uterina (IIU) em pacientes com causa masculina de infertilidade e salientar o caráter prognóstico de cada parâmetro para o sucesso da técnica. MÉTODOS: duzentos e trinta e nove ciclos de IIU (155 casais) foram analisados durante período de 15 meses. Todos as pacientes foram submetidas à induçäo da ovulaçäo de acordo com protocolos do I Consenso Brasileiro de Induçäo de Ovulaçäo. A análise seminal foi realizada de acordo com os critérios da Organizaçäo Mundial da Saúde (OMS) para parâmetros como concentraçäo e motilidade e critério estrito de Kruger para avaliaçäo da morfologia. O preparo da amostra para IIU foi realizado por meio de gradientes descontínuos de densidade (ISolate®). Foram formados dois grupos de pacientes de acordo com o resultado de gestaçäo: grupo G - gestaçäo positiva e grupo NG - gestaçäo negativa. RESULTADOS: näo houve diferença significativa nos valores obtidos relacionados à concentraçäo total de espermatozóides/mL de sêmen, motilidade total, motilidade de formas progressivas antes e depois do processamento seminal. Quando a morfologia estrita de Kruger foi comparada entre os dois grupos, diferenças significativas foram encontradas (grupo G=10,6 por cento de formas normais; grupo NG=6,4 por cento de formas normais; p<0.05) quando o número de espermatozóides/mL inseminados foi superior a 15 milhöes/mL, a taxa de gravidez foi significantemente maior. CONCLUSÖES: tanto a morfologia estrita de Kruger como o número de espermatozóides inseminados parecem ser fatores prognósticos positivos para o estabelecimento da gestaçäo, devendo ser salientados na investigaçäo do homem infértil

Different intervals between vasectomy and sperm retrieval interfere in the reproductive capacity from vasectomized men.

PURPOSE: To evaluate the relationship between the postvasectomy period and sperm reproductive capacity after ICSI. METHODS: Seventy-seven ICSI cycles with percutaneous epididymal sperm aspiration (PESA) were reviewed. Patients were divided into 4 groups according to the interval after vasectomy: 0 - 5 years (G1); 6 - 8 years (G2); 9 - 14 years (G3), and > 15 years (G4). RESULTS: Clinical and ongoing pregnancy rates did not correlate significantly with the time period of vasectomy until 14 years. Although the higher implantation rate observed in G1, no significant differences were noted among Groups 1-3. The miscarriage rates increased from G1 to G4, reaching a statistical significance among G1, G2, and G3 compared with G4. When groups were also divided according to the maternal age, the same results were obtained. CONCLUSIONS: The interval between the vasectomy and the sperm retrieval procedure has no effect on the outcome until the interval of 14 years.

Testicular sperm results in elevated miscarriage rates compared to epididymal sperm in azoospermic patients

CONTEXT: Several sperm retrievel techniques are available for use on azoospeermic men. Comparisons between spermatozoa retrieved from the testicles and epidymis in relation to pregnancy and miscarriage rates are not well established. OBJECTIVE: To compare pregnancy and miscarriage rates using sperm retrieved from the testes and epididymis using intracytoplasmic sperm injection. Furthermore, we evaluated the fertilization and pregnancy rates according to the status of the spermatozoa retrieved (motile or immotile). DESIGN: Retrospective study. SETTING: A private center for assisted fertilization. PARTICIPANTS: One hundred and eight consecutive patients who presented with azoospermia were included in our study, on whom a total of 144 retrieval procedures were performed. PROCEDURES: Of the 144 retrieval procedures, 104 were testicular sperm aspirations (TESA) and 40 were epididymal sperm aspirations (PESA). PESA was the first approach in obstructive patients (n = 68), whereas TESA was used when the former failed. For non-obstructive patients (n = 40), TESA was the method of retrieval. MAIN MEASUREMENTS: Pregnancy and miscarriage rates according to spermatozoa characteristics (motile or immotile). RESULTS: The number of cycles performed using spermatozoa retrieved from the testicles and epididymis was 81 and 30, respectively. Motile spermatozoa had higher fertilization (2PN) and pregnancy rates compared to immotile spermatozoa (p < 0.05). Also, motile spermatozoa had lower miscarriage rates compared to immotile spermatozoa (p < 0.0001). No differences were seen in pregnancy rates with testicular spermatozoa (n = 28) compared to epididymal spermatozoa (n = 13) (p = 0.1). However, the miscarriages rates were higher in spermatozoa retrieved from the testicles (n = 12) compared to epididymis retrievals (n = 1) (p = 0.01). CONCLUSIONS: Although pregnancy rates were similar when the intracytoplasmic sperm injection was performed with spermatozoa retrieved from the testicles and epididymis, the use of testicular spermatozoa yields a significantly higher miscarriage rate. It is possible that the higher miscarriage rate seen in patients using spermatozoa retrieved from the testicles is linked to high genetic sperm abnormalities