Prenatal Exposure to Potentially Toxic Metals and Their Effects on Genetic Material in Offspring: a Systematic Review.

In recent years, the background level of environmental pollutants, including metals, has increased. Pollutant exposure during the earliest stages of life may determine chronic disease susceptibility in adulthood because of genetic or epigenetic changes. The objective of this review was to identify the association between prenatal and early postnatal exposure to potentially toxic metals (PTMs) and their adverse effects on the genetic material of offspring. A systematic review was carried out following the Cochrane methodology in four databases: PubMed, Scopus, Web of Science, and the Cochrane Library. Eligible papers were those conducted in humans and published in English between 2010/01/01 and 2021/04/30. A total of 57 articles were included, most of which evaluated prenatal exposure. Most commonly evaluated PTMs were As, Cd, and Pb. Main adverse effects on the genetic material of newborns associated with PTM prenatal exposure were alterations in telomere length, gene or protein expression, mitochondrial DNA content, metabolomics, DNA damage, and epigenetic modifications. Many of these effects were sex-specific, being predominant in boys. One article reported a synergistic interaction between As and Hg, and two articles observed antagonistic interactions between PTMs and essential metals, such as Cu, Se, and Zn. The findings in this review highlight that the problem of PTM exposure persists, affecting the most susceptible populations, such as newborns. Some of these associations were observed at low concentrations of PTMs. Most of the studies have focused on single exposures; however, three interactions between essential and nonessential metals were observed, highlighting that metal mixtures need more attention.

Advances and Challenges of Using Ovarian Preantral Follicles to Develop Biobanks of Wild Mammals.

The extinction rate of mammalian species has been accelerated in recent decades. It therefore is important to preserve and store genetic materials in cryobanks for research purposes and subsequent production of offspring through assisted reproductive techniques. Along with the systematic collection and storage of germplasm, research efforts focusing on in vitro culture to produce mature gametes are critical. Specifically, obtaining mature oocytes from in vitro culture of preantral follicles is a great option to enhance female fertility preservation since these early follicles represent 90%-95% of the follicular population on ovarian cortex biopsy. This review presents current advances and discusses limitations and prospects about isolation, cryopreservation/banking, and in vitro culture of preantral follicles from wild species.

Investigando a fragmentação não induzida e a susceptibilidade à fragmentação do DNA espermático: refinamento da avaliação espermática. Parte 2 / Investigating not induced fragmentation and susceptibility of sperm DNA: refinement of sperm evaluation. Second part

A integridade do DNA espermático está intimamente relacionada com a fertilidade. Técnicas queavaliam o DNA espermático possuem grande importância e potencial de serem aplicadas nas rotinas dasavaliações andrológicas. No entanto, são necessários maiores conhecimentos sobre as técnicas e seus princípiospara permitir que sejam empregadas apropriadamente de acordo com os objetivos e resultados esperados. Dessaforma, assim como abordado na Parte 1, nesta Parte 2 serão abordadas técnicas de avaliação do DNAespermático: TUNEL, laranja de acridina e Sperm Chromatin Structure Assay (SCSA). A técnica de TUNELfundamenta-se na adição de nucleotídeos modificados marcados com fluorescência às fitas fragmentadas. Atécnica avalia, portanto, diretamente a fragmentação de DNA da amostra. A sonda laranja de acridina é capaz dediferenciar, por meio de diferentes colorações, o DNA fragmentado do não fragmentado. Já a técnica de SCSAbaseia-se na imposição de um desafio ao espermatozoide; esta técnica avalia a susceptibilidade dosespermatozoides à fragmentação de DNA, sendo, portanto, uma avaliação indireta. A presente revisão permiteexpor a opinião dos autores sobre as diferentes técnicas e suas aplicações, mostrando o potencial emprego delasnos exames de rotina e nas pesquisas.(AU)

Sperm DNA integrity is directly related to male fertility. Techniques to assess sperm DNA are importantand have potential to be applied in routine evaluation of semen. However, the techniques and principles need tobe further studied to apply they correctly and to get relevant results. Thus, as discussed in Part 1, Part 2 willaddress evaluation techniques of sperm DNA: TUNEL, acridine orange and Sperm Chromatin Structure Assay(SCSA). TUNEL is based on addition of modified nucleotides labeled with fluorescence to fragmented tapesevaluating, therefore, directly, the DNA fragmentation of the sample. Acridine orange differentiates fragmentedDNA by different colors. On the other hand, SCSA are based on the imposition of a challenge to sperm. Thus,this technique assesses the susceptibility of sperm DNA fragmentation constituting indirect assessment. Thereview allows showing opinion of the authors about different techniques and their applications. Therefore,potential use of these techniques in routine exams and researches are addressed.(AU)

Investigando a fragmentação não induzida e a susceptibilidade à fragmentação do DNA espermático: refinamento da avaliação espermática. Parte 2 / Investigating not induced fragmentation and susceptibility of sperm DNA: refinement of sperm evaluation. Second part

A integridade do DNA espermático está intimamente relacionada com a fertilidade. Técnicas queavaliam o DNA espermático possuem grande importância e potencial de serem aplicadas nas rotinas dasavaliações andrológicas. No entanto, são necessários maiores conhecimentos sobre as técnicas e seus princípiospara permitir que sejam empregadas apropriadamente de acordo com os objetivos e resultados esperados. Dessaforma, assim como abordado na Parte 1, nesta Parte 2 serão abordadas técnicas de avaliação do DNAespermático: TUNEL, laranja de acridina e Sperm Chromatin Structure Assay (SCSA). A técnica de TUNELfundamenta-se na adição de nucleotídeos modificados marcados com fluorescência às fitas fragmentadas. Atécnica avalia, portanto, diretamente a fragmentação de DNA da amostra. A sonda laranja de acridina é capaz dediferenciar, por meio de diferentes colorações, o DNA fragmentado do não fragmentado. Já a técnica de SCSAbaseia-se na imposição de um desafio ao espermatozoide; esta técnica avalia a susceptibilidade dosespermatozoides à fragmentação de DNA, sendo, portanto, uma avaliação indireta. A presente revisão permiteexpor a opinião dos autores sobre as diferentes técnicas e suas aplicações, mostrando o potencial emprego delasnos exames de rotina e nas pesquisas.

Sperm DNA integrity is directly related to male fertility. Techniques to assess sperm DNA are importantand have potential to be applied in routine evaluation of semen. However, the techniques and principles need tobe further studied to apply they correctly and to get relevant results. Thus, as discussed in Part 1, Part 2 willaddress evaluation techniques of sperm DNA: TUNEL, acridine orange and Sperm Chromatin Structure Assay(SCSA). TUNEL is based on addition of modified nucleotides labeled with fluorescence to fragmented tapesevaluating, therefore, directly, the DNA fragmentation of the sample. Acridine orange differentiates fragmentedDNA by different colors. On the other hand, SCSA are based on the imposition of a challenge to sperm. Thus,this technique assesses the susceptibility of sperm DNA fragmentation constituting indirect assessment. Thereview allows showing opinion of the authors about different techniques and their applications. Therefore,potential use of these techniques in routine exams and researches are addressed.

Criopreservação de tecido ovariano: limitações e perspectivas para a preservação da fertilidade de fêmeas / Ovarian tissue cryopreservation: limitations and perspectives for female pertility preservation

Background: The ovarian tissue cryopreservation has been achieved a great notoriety in the Reproductive Biology area, due to its potential in preserving female fertility through the protection of exocrine and endocrine functions of the ovary. The association of this technique with in vitro culture and/or transplant in adult or young individuals who has not initiated its reproductive activities represents not only the conservation and perpetuation of the genetic material of economic valuable animals, but also the preservation of female gametes from endangered species, or even from young women who may have ovarian dysfunctions caused by gonadotoxic treatments. Studies with some species (human, mice and ovine) have demonstrated the recovery of the ovarian function and the birth of healthy offspring after transplant of ovarian tissue which has been previously cryopreserved. However, most studies have shown that ovarian cryopreservation process offer risks to different structures (follicle and stroma ) as well as to the different cell types (oocyte, granulosa, thecal and stromal cells), which constitute this tissue. Review: Extreme cold, intracellular ice crystallization, osmotic shock and the toxicity of the cryoprotectant agents are factors that are usually associated with the injuries caused by the cryopreservation process. As a direct or indirect consequence, those factors limit the success of the cryopreservation of ovarian tissue, since they affect the survival or alter the tissue functionality or cellular structure, like the ovarian follicles, for example, after the thawing/warming procedure. Among the injuries that may take place as a consequence of those factors, we can mention: cell death by the necrotic or apoptotic pathways; alterations in normal levels of genic expression; ischemia and changes of communication and interaction between the oocyte and follicular cells. As a result, many authors have studied and developed protocols of cryopreservation that may prevent or minimize the cryoinjuries, since the cryopreservation per si or combined to other techniques (in vitro culture and/or transplant) can compromise the ovarian integrity, leading consequently to a significant loss of follicles. In this regard, the present review seeks to app roach the advantages of the cryopreservation of ovarian tissue; indicating the difficulties and challenges that encompass this procedure, with purpose of pointing out solutions to overcome the damages of ovarian tissue cryopreservation, through of convenient cryopreservation protocols that avoid those follicular losses. For this purpose, it is necessary the preservation of the follicular viability as well as the preservation of the tissue integrity and the contact between reproductive (oocytes) or somatic cells, which are essentials to the follicle development, and, consequently, to the embryo production. Conclusion: The use of cryopreserved ovarian tissue is an important strategy to the preservation of female fertility. This tool has being pointed as an alternative way to the cryopreservation of mature oocytes and embryos. However, additional studies are necessary to diminishing the cellular damages inherent to this procedure, especially those related to the comprehension of the obstacles and mechanisms associated to the exposition to extreme cold.(AU)

Criopreservação de tecido ovariano: limitações e perspectivas para a preservação da fertilidade de fêmeas / Ovarian tissue cryopreservation: limitations and perspectives for female pertility preservation

Background: The ovarian tissue cryopreservation has been achieved a great notoriety in the Reproductive Biology area, due to its potential in preserving female fertility through the protection of exocrine and endocrine functions of the ovary. The association of this technique with in vitro culture and/or transplant in adult or young individuals who has not initiated its reproductive activities represents not only the conservation and perpetuation of the genetic material of economic valuable animals, but also the preservation of female gametes from endangered species, or even from young women who may have ovarian dysfunctions caused by gonadotoxic treatments. Studies with some species (human, mice and ovine) have demonstrated the recovery of the ovarian function and the birth of healthy offspring after transplant of ovarian tissue which has been previously cryopreserved. However, most studies have shown that ovarian cryopreservation process offer risks to different structures (follicle and stroma ) as well as to the different cell types (oocyte, granulosa, thecal and stromal cells), which constitute this tissue. Review: Extreme cold, intracellular ice crystallization, osmotic shock and the toxicity of the cryoprotectant agents are factors that are usually associated with the injuries caused by the cryopreservation process. As a direct or indirect consequence, those factors limit the success of the cryopreservation of ovarian tissue, since they affect the survival or alter the tissue functionality or cellular structure, like the ovarian follicles, for example, after the thawing/warming procedure. Among the injuries that may take place as a consequence of those factors, we can mention: cell death by the necrotic or apoptotic pathways; alterations in normal levels of genic expression; ischemia and changes of communication and interaction between the oocyte and follicular cells. As a result, many authors have studied and developed protocols of cryopreservation that may prevent or minimize the cryoinjuries, since the cryopreservation per si or combined to other techniques (in vitro culture and/or transplant) can compromise the ovarian integrity, leading consequently to a significant loss of follicles. In this regard, the present review seeks to app roach the advantages of the cryopreservation of ovarian tissue; indicating the difficulties and challenges that encompass this procedure, with purpose of pointing out solutions to overcome the damages of ovarian tissue cryopreservation, through of convenient cryopreservation protocols that avoid those follicular losses. For this purpose, it is necessary the preservation of the follicular viability as well as the preservation of the tissue integrity and the contact between reproductive (oocytes) or somatic cells, which are essentials to the follicle development, and, consequently, to the embryo production. Conclusion: The use of cryopreserved ovarian tissue is an important strategy to the preservation of female fertility. This tool has being pointed as an alternative way to the cryopreservation of mature oocytes and embryos. However, additional studies are necessary to diminishing the cellular damages inherent to this procedure, especially those related to the comprehension of the obstacles and mechanisms associated to the exposition to extreme cold.