From in vivo to in vitro: exploring the key molecular and cellular aspects of human female gametogenesis.

Human oogenesis is a highly complex and not yet fully understood process due to ethical and technological barriers that limit studies in the field. In this context, replicating female gametogenesis in vitro would not only provide a solution for some infertility problems, but also be an excellent study model to better understand the biological mechanisms that determine the formation of the female germline. In this review, we explore the main cellular and molecular aspects involved in human oogenesis and folliculogenesis in vivo, from the specification of primordial germ cells (PGCs) to the formation of the mature oocyte. We also sought to describe the important bidirectional relationship between the germ cell and the follicular somatic cells. Finally, we address the main advances and different methodologies used in the search for obtaining cells of the female germline in vitro.

Cervical cancer patients that respond to chemoradiation therapy display an intense tumor infiltrating immune profile before treatment.

Cervical cancer (CC) is a major cause of death and suffering to women globally with 570,000 new cases in 2017. It disproportionately affects those living in resource-constrained countries such as Brazil, with 90% of the deaths from CC happening in low and middle-income countries. Early detection is still the best strategy for improving response to therapy and survival and cases detected in advanced stages show variable response rates to the standard chemoradiation therapy protocols. Both the genetic landscape and the immune status of patients can dramatically affect cancer progression and response to therapy, as well as disease recurrence. Here we performed a comprehensive sequencing analysis using the cancer gene panel - Ion AmpliSeq™ Cancer hotspot Panel V2 CHPv2, as well as determined the immune infiltrate composition of a group of locally advanced CC patients with the goal of identifying genetic and immune characteristics associated with a clinical response to therapy. The expression levels of CD68+ tumor-associated macrophages (TAMs) and CD8+ tumor-infiltrating lymphocytes (TILs), as well as the immune checkpoint molecules PD-1, PD-L1 and PD-L2 in stroma and in tumor regions were analyzed by immunohistochemistry (IHC). The HPV infection status with high-risk strains was also determined. Twenty-one samples from patients with squamous cell carcinoma segregated into responder (11) and non-responder (10) groups according to standard chemoradiation therapy response were studied. Our findings indicate that responder patients showed an increase of an inflammatory tumor microenvironment as indicated by higher numbers of CD8+ and PD-L2+ TILs, as well as higher expression of PD-L1 immunoreactive area, as compared to the non-responder group. Additionally, our results demonstrate a correlation between the number of gene mutations and PD-L2+ TILs in the responder group. The genes PIK3CA and KDR/VEGFR were the most mutated genes, corroborating past findings. Together, these findings indicate an inflammatory tumor microenvironment present in patients that will respond to future chemoradiation treatment as compared to those that will not. This points to possible future predictors of response to therapy in CC patients.

Frequency of Chromosomal Abnormalities in Products of Conception.

Purpose To describe the frequencies of chromosomal abnormalities found in abortion material, and to observe its correlation to maternal age. Methods A retrospective study was conducted based on data obtained from the databank of a medical genetics laboratory in Belo Horizonte, MG, Brazil. A total of 884 results from products of conception analysis were included, 204 of which were analyzed by cytogenetics, and 680 by molecular biology based on quantitative fluorescence polymerase chain reaction (QF-PCR). The frequency of individual chromosomal aberrations and the relationship between the presence of anomalies and maternal age were also evaluated. Results The conventional cytogenetics technique was able to detect 52% of normal and 48% of abnormal results in the analyzed material. Quantitative fluorescence polymerase chain reaction revealed 60% of normal and 40% of abnormal results from the samples evaluated by this method. The presence of trisomy 15 was detected only by cytogenetics, as it was not included in the QF-PCR routine investigation in the laboratory. A significant increase in abnormal results was observed among women aged 35 years or older compared with younger women (p = 0.02). Conclusion Chromosomal aberrations are still a major cause of spontaneous abortion, and the conventional cytogenetics technique is efficient for miscarriage material analysis, but molecular methods such as QF-PCR are adequate complementary strategies to detect the major chromosomal anomalies, leading to technical reports with reliable results.

Frequency of Chromosomal Abnormalities in Products of Conception / Frequência de anomalias cromossômicas em material de aborto

Abstract Purpose To describe the frequencies of chromosomal abnormalities found in abortion material, and to observe its correlation to maternal age. Methods A retrospective study was conducted based on data obtained from the databank of a medical genetics laboratory in Belo Horizonte, MG, Brazil. A total of 884 results from products of conception analysis were included, 204 of which were analyzed by cytogenetics, and 680bymolecular biology basedon quantitative fluorescence polymerase chain reaction (QF-PCR). The frequency of individual chromosomal aberrations and the relationship between the presence of anomalies and maternal age were also evaluated. Results The conventional cytogenetics technique was able to detect 52% of normal and 48% of abnormal results in the analyzed material. Quantitative fluorescence polymerase chain reaction revealed 60% of normal and 40% of abnormal results from the samples evaluated by this method. The presence of trisomy 15 was detected only by cytogenetics, as it was not included in the QF-PCR routine investigation in the laboratory. A significant increase in abnormal results was observed among women aged 35 years or older compared with younger women (p = 0.02). Conclusion Chromosomal aberrations are still a major cause of spontaneous abortion, and the conventional cytogenetics technique is efficient for miscarriage material analysis, but molecular methods such as QF-PCR are adequate complementary strategies to detect the major chromosomal anomalies, leading to technical reports with reliable results.

Resumo Objetivos Descrever a frequência de anomalias cromossômicas encontradas em material de aborto, e observar se estas estão relacionadas com a idade materna. Métodos Foi realizado um estudo retrospectivo no banco de dados de um laboratório de genética médica em Belo Horizonte, MG. O estudo incluiu 204 resultados avaliados por citogenética, e 680 resultados por biologia molecular baseada em reação em ensaio fluorescente da reação em cadeia da polimerase (QF-PCR), totalizando um número de 884 análises. A frequência de diferentes anomalias cromossômicas e a relação entre a presença de anomalias e a idade materna também foi avaliada. Resultados A citogenética convencional foi capaz de detectar 52% de resultados normais e 48% de resultados anormais no material analisado. A QF-PCR revelou 60% de resultados normais e 40% de anormais nas amostras avaliadas por esta técnica. A presença da trissomia 15 foi detectada por citogenética,mas até então não era incluída na investigação por QF-PCR no laboratório. Umaumento significativo na quantidade de resultados anormais foi observado em mulheres comidade de 35 anos ou mais, quando comparado a mulheres mais jovens (p = 0,02). Conclusão As aberrações cromossômicas são causas importantes de abortos espontâneos, e o estudo citogenético é eficaz para a análise das amostras de material de aborto, mas as técnicas moleculares, como a QF-PCR, representam métodos complementares adequados para detectar as principais anomalias cromossômicas, possibilitando a liberação de laudos com resultados confiáveis.

Estudo das regulamentações de reprodução humana assistida no Brasil, Chile, Uruguai e na Argentina / Study of the regulations of assisted human reproduction in Brazil, Argentina, Chile and Uruguay

Objetivo: Estudar as regulamentac¸ões referentes à reproduc¸ão humana assistida no Brasil,Chile, Uruguai e na Argentina.Método: Estudo qualitativo transversal das regulamentac¸ões referentes à reproduc¸ãohumana assistida no Brasil, Chile, Uruguai e na Argentina entre novembro de 2013 e abrilde 2014.Resultados: O Brasil é regido pela Resoluc¸ão do Conselho Federal de Medicina n◦2.013/2013;a Argentina pela Lei n◦26.862/2013; no Chile, até o momento, não existe uma lei queregulamente o uso das técnicas; e no Uruguai o projeto de lei n◦19.167/2013 aguardaregulamentac¸ão.Conclusão: Brasil, Argentina, Chile e Uruguai são países sul-americanos que se encon-tram em distintas situac¸ões legais no que diz respeito à regulamentac¸ão das práticas dereproduc¸ão humana assistida.

tObjective: This research aimed to study the regulations of Assisted Human Reproduction inBrazil, Argentina, Chile and Uruguay.Method: A cross-sectional qualitative study of the regulations relating to Assisted HumanReproduction in Brazil, Argentina, Chile and Uruguay was conducted between November of2013 and April of 2014. Results: Regarding the Assisted Human Reproduction techniques, currently Brazil is regulatedby Resolution of the Federal Council of Medicine No. 2013/2013; Argentina by LawNo. 26,862/2013; in Chile, so far, there is no law regulating the use of these techniques; inUruguay the draft bill No. 19,167/2013 awaits regulation.Conclusion: Brazil, Argentina, Chile and Uruguay are South American countries that arein different legal situations regarding to the regulation of the practice of Assisted HumanReproduction.