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Infertility is a growing worldwide public health problem, and stress is a main factor exerting detrimental effects on female reproduction. However, knowledge regarding the neuroendocrine changes caused by chronic stress in females is limited. Therefore, this study assessed the effects of stress on hormones that control female reproduction during the proestrus and diestrus stages of the estrous cycle, as well as its effects on fertility. Adult females were assigned to either a control or a stress group. Stress consisted of exposure, for 15 min, to cold-water immersion daily for 30 days. Estrous cyclicity, female sexual behavior, as well as hypothalamic kisspeptin, gonadotropin releasing hormone (GnRH) content, serum luteinizing hormone (LH), estradiol (E2), progesterone (P4), corticosterone (CORT) and fertility were assessed after chronic stress. The results show that chronically stressed females exhibited disrupted estrous cyclicity, decreased receptivity, low pregnancy rates and lower numbers of fetuses. The content of Kisspeptin and GnRH in the Anteroventral Periventricular/medial Preoptic Area decreased during proestrus, while Kisspeptin increased in the Arcuate nucleus in proestrus and diestrus. Serum LH decreased only during proestrus, whereas E2 and P4 concentrations decreased during proestrus and diestrus, with a concomitant increase in CORT levels in both stages. As a whole, these results indicate that chronic stress decreases Kisspeptin content in AVPV nucleus and GnRH in POA in females, and might induce disruption of the LH surge, consequently disrupting estrous cyclicity and fertility, leading to lower rates of pregnancy and number of fetuses.

The reactive oxygen species (ROS) play an important role in various aspects of male reproductive function, for spermatozoa to acquire the ability to fertilize. However, the increase in ROS generation, both due to internal and external factors, can induce oxidative stress, causing alterations in the structure and function of phospholipids and proteins. In the nucleus, ROS attack DNA, causing its fragmentation and activation of apoptosis, thus altering gene and protein expression. Accumulating evidence also reveals that endogenously produced ROS can act as second messengers in regulating cell signalling pathways and in the transduction of signals that are responsible for regulating spermatogonia self-renewal and proliferation. In the epididymis, they actively participate in the formation of disulphide bridges required for the final condensation of chromatin, as well as in the phosphorylation and dephosphorylation of proteins contained in the fibrous sheath of the flagellum, stimulating the activation of progressive motility in epididymal spermatozoa. In this review, the role of small amounts of ROS during spermatogenesis and epididymal sperm maturation was discussed.

Background The emergence of assisted reproductive technology (ART) in humans has been an important tool for the treatment of infertility. The number of treatments performed in Latin America has been increasing, and Mexico is the third country with the most assisted reproduction cycles performed in the region. However, Mexico lacks a national regulation for assisted reproduction. Therefore, it is necessary to implement regulations that allow for a safe clinical practice based on ethics which can be available to any social group. Main body The aim of this review was to examine the existing legislation that regulates human assisted reproduction practices in Mexico, but also to examine the legal analysis of the policies, laws, and regulations in effect in some countries in Latin America, North America, and Europe. For this, seven databases were consulted, and 34 articles from 2004 to 2021 referring to the practice of ART within the legal framework and the anthropological analysis that this entails were analyzed. Eight documents were also consulted such as the Mexican General Health Law of the Official Journal of the Federation (February 7, 1984) with its last published reform (DOF 01-06-2021). And three official agency websites were also consulted. No specific legislation was found for human assisted reproduction practices in Mexico; however, assisted reproduction clinics are ruled under some agreements implemented by national organizations such as the Mexican Association of Reproductive Medicine and, at the Latin America level, the Latin America Network of Assisted Reproduction (abbreviated REDLARA in Spanish); in addition, the practice of ART is considered, although not explicitly, in the General Health Law. Conclusion In Mexico, there is no legal regulation in charge of assisted reproduction practices, which is why there is an urgent need to establish human assisted reproduction laws without incurring discriminatory and unconstitutional acts, and at the same time, be in accordance with scientific advances. This will allow a considerable reduction in the violation of human rights., Plain language summary The emergence of ART in humans has been an important tool for the treatment of infertility. It is reported that one in four couples in developing countries has fertility problems. In 2009, the International Committee for Monitoring Assisted Reproductive Technology (ICMART) established ART as "all treatments or procedures involving in vitro manipulation of oocytes, sperm or embryos for the purpose of establishing a pregnancy". The number of treatments performed in Latin America has been increasing, and Mexico is the third country with the most assisted reproduction cycles performed in the region. However, Mexico lacks a national regulation for human assisted reproduction. This has caused Mexico to become a medical tourism paradise, which increases the possibility of abuses, fraud, and clinical risks. In addition, it allows each institution offering assisted reproduction services, whether public or private, to establish arbitrary requirements for inclusion. Thus, the emergence of a regulation that allows a safe clinical practice based on ethics, which will also make this reproductive tool available to any social group, is a social need. Therefore, the aim of this review was to examine the existing legislation that regulates human assisted reproduction practices in Mexico, but also to examine the legal analysis of the policies, laws, and regulations in use in some countries in Latin America, North America, and Europe, as well as highlighting the importance of working on the establishment of regulations that allow for safe and ethically based clinical practices.

Background The evaluation of the DNA damage generated in cumulus cells after mature cumulus-oocyte complexes vitrification can be considered as an indicator of oocyte quality since these cells play important roles in oocyte developmental competence. Therefore, the aim of this study was to determine if matured cumulus-oocyte complexes exposure to cryoprotectants (CPAs) or vitrification affects oocytes and cumulus cells viability, but also if DNA damage is generated in cumulus cells, affecting fertilization and embryo development. Results The DNA damage in cumulus cells was measured using the alkaline comet assay and expressed as Comet Tail Length (CTL) and Olive Tail Moment (OTM). Results demonstrate that oocyte exposure to CPAs or vitrification reduced oocyte (75.5 ± 3.69%, Toxicity; 66.7 ± 4.57%, Vitrification) and cumulus cells viability (32.7 ± 5.85%, Toxicity; 7.7 ± 2.21%, Vitrification) compared to control (95.5 ± 4.04%, oocytes; 89 ± 4.24%, cumulus cells). Also, significantly higher DNA damage expressed as OTM was generated in the cumulus cells after exposure to CPAs and vitrification (39 ± 17.41, 33.6 ± 16.69, respectively) compared to control (7.4 ± 4.22). In addition, fertilization and embryo development rates also decreased after exposure to CPAs (35.3 ± 16.65%, 22.6 ± 3.05%, respectively) and vitrification (32.3 ± 9.29%, 20 ± 1%, respectively). It was also found that fertilization and embryo development rates in granulose-intact oocytes were significantly higher compared to denuded oocytes in the control groups. However, a decline in embryo development to the blastocyst stage was observed after CPAs exposure (1.66 ± 0.57%) or vitrification (2 ± 1%) compared to control (22.3 ± 2.51%). This could be attributed to the reduction in both cell types viability, and the generation of DNA damage in the cumulus cells. Conclusion This study demonstrates that oocyte exposure to CPAs or vitrification reduced viability in oocytes and cumulus cells, and generated DNA damage in the cumulus cells, affecting fertilization and embryo development rates. These findings will allow to understand some of the mechanisms of oocyte damage after vitrification that compromise their developmental capacity, as well as the search for new vitrification strategies to increase fertilization and embryo development rates by preserving the integrity of the cumulus cells.

Phytoestrogens are considered to be endocrine disruptors, since they can alter the endocrine system, thus disturbing many reproductive events. The intake of diets containing a high content of phytoestrogens has increased worldwide in human populations and in domestic animals. Phytoestrogens in maternal blood can pass through the placenta to the fetus in high amounts and can have long-term organizational effects. Mesquite (Prosopis sp) is a leguminous plant widely used to feed several livestock species, and is also used in the human diet. In this study we assessed the effects of exposure to mesquite pod extract during the periconception and pregnancy periods on the reproduction of male and female descendants. The females of three experimental groups received one of the following treatments: 1) vehicle injection; 2) mesquite pod extract or 3) the isoflavone daidzein during the periconception and pregnancy periods. Estrous cyclicity, sexual behavior and hormones, as well as uterine and vaginal epithelia were evaluated in the female descendants. In the males, sexual behavior and hormones, apoptosis in testicular cells and sperm quality were evaluated. In females the following was observed: alterations in estrous cycles, decreased sexual behavior, estradiol and progesterone levels, increased uterine and vaginal epithelia. In males, we observed a decrease in sexual behavior, testosterone and sperm quality, and apoptosis increased in testicular cells. All these effects were similar to those caused by daidzein. These results indicate that prenatal exposure to mesquite pod extract or daidzein, administered to females before and during pregnancy, can disrupt normal organizational-activational programming of reproductive physiology in female and male descendants.

Simple Summary Recently, a significant relationship between stress and reproductive failure in women was reported; being one of the possible causes of infertility. The World Health Organization recognizes infertility as a global public health issue; therefore, the interest in understanding the main causes of this issue has increased over the last few decades. Thus, many studies have reported that stress can adversely alter the functionality of the hypothalamic-pituitary-gonadal axis; as well as being one of the reasons of subfertility in patients undergoing in vitro fertilization. Therefore, it can be assumed that stress is closely related to poor in vitro fertilization outcomes. In chronically stressed female rats, irregular estrous cyclicity, increased corticosterone levels, decreased oocyte viability, and increased percentage of abnormal oocytes were obtained in all estrous cycle phases, resulting in reduced oocyte maturation during proestrus. Oocyte maturation disturbed by chronic stress is a crucial factor by which chronic stress disrupts female reproduction. Abstract Background: Stress has been considered as one of the causes of decreased reproductive function in women. However, direct evidence of the effect of chronic stress on oocytes depending on estrous cycle phases is limited. Objective: The present study aimed to evaluate the impact of chronic stress on the viability, integrity, and maturation of rat oocytes depending on estrous cycle phases, specifically proestrus, estrus, and diestrus. Methods: For this purpose, adult female rats were stressed daily by cold water immersion (15 °C) for 30 consecutive days. Results: In chronically stressed female rats, irregular estrous cyclicity, increased corticosterone levels, decreased oocyte viability, and an increased percentage of abnormal oocytes were obtained in all the estrous cycle phases, resulting in reduced oocyte maturation during proestrus. Conclusion: Oocyte maturation disturbed by chronic stress is a crucial factor by which chronic stress disrupts female reproduction

Vitrification may severely affect cumulus cells and oocyte morphology and viability, limiting their maturation and developmental potential. The aim of this study was to evaluate the gap junction intercellular communication (GJIC) integrity after the vitrification of porcine immature cumulus-oocyte complexes (COCs). Fresh COCs were randomly distributed in three groups: untreated (control), toxicity (cryoprotectants exposure), and vitrification, then subjected to in vitro maturation (IVM). Oocyte viability and IVM were measured in all groups. The evaluation of GJIC was expressed as relative fluorescence intensity (RFI). Vitrification significantly decreased oocyte viability and maturation after 44 h of culture compared to control. Also, significantly reduced RFI was observed in vitrified COCs during the first hours of culture (4–8 h) compared to control. This study demonstrates that porcine oocyte viability and maturation after 44 h of culture decreased after vitrification. GJIC was also affected during the first hours of culture after the vitrification of immature oocytes, being one of the possible mechanisms by which oocyte maturation decreased.

Introducción: La mayoría de los estudios cuyo objetivo es evaluar las tasas de recuperación y desarrollo post-vitrificación de ovocitos porcinos han reportado mejores tasas cuando estos son criopreservados en estados embrionarios, pero no así en ovocitos inmaduros. Por esta razón, para mejorar los protocolos de la vitrificación de ovocitos inmaduros, se ha evaluado el uso de diferentes tipos y concentraciones de crioprotectores, recipientes, y tiempos de incubación, pero muy pocos han evaluado cual técnica de fertilización permite incrementar las tasas de producción de blastocistos en ovocitos vitrificados. El objetivo de esta tesis fue evaluar el desarrollo embrionario hasta el estado de blastocisto a partir de ovocitos inmaduros vitrificados madurados con un sistema de co-cultivo con células de la granulosa y fertilizados por fertilización in vitro (FIV), inyección intracitoplasmática de espermatozoides (ICSI) e inyección fisiológica intracitoplasmática de espermatozoides (PICSI). Así como, evaluar si la selección espermática con ácido hialurónico (AH) o con polivinilpirrolidona (PVP) antes de la inyección intracitoplasmática participan en forma importante en el incremento de las tasas de fertilización y producción de blastocistos. Resultados: Los espermatozoides seleccionados con AH-PICSI mostraron un mayor porcentaje de espermatozoides vivos/reaccionados en comparación con el control y los expuestos a la PVP. Altas tasas de espermatozoides muertos/reaccionados se obtuvieron después de la exposición a la PVP en comparación con el control y el AH. En ovocitos, la viabilidad disminuyó significativamente en el control de ovocitos denudados (DO) y en ovocitos vitrificados. Además, la maduración disminuyó significativamente en el control DO. En cuanto a los parámetros de fertilización, la FIV mostró mayores porcentajes de monospermia en comparación con la ICSI y la PICSI. Sin embargo, los resultados demostraron que la fertilización con PICSI incrementó la formación de blastocistos en los grupos control complejo ovocito-células del cúmulo (COCs), control ovocitos denudados co-cultivados con células de la granulosa (DO-GC) y ovocitos vitrificados en comparación con la FIV e ICSI. Conclusión: Para lograr altas tasas de formación de blastocitos a partir de ovocitos vitrificados en etapa de vesícula germinal (VG), se recomienda que el espermatozoide se seleccione antes de ser inyectado al ovocito con AH en lugar de la PVP, debido a que se obtienen mayores tasas de viabilidad y reacción acrosomal. Además, la fertilización por PICSI es el mejor método para producir mayores tasas de blastocistos en comparación con los procedimientos de FIV e ICSI.

Parabens (PBs) are compounds widely used in industry for food and personal care products as antimicrobials and preservatives. Their indiscriminate use has resulted in their detection in different ecosystems so that humans and other organisms are highly exposed. Methylparaben (MePB), compared with other PBs, is mostly detected in food, personal care and baby care products. PBs could be linked to the generation of hormonal disorders and fertility impairment since their recent classification as endocrine disruptors. The knowledge of the effects that MePB can exert is of great importance as, in terms of reproduction, information is limited. Therefore, the objective of the present study was to evaluate the effect of MePB on porcine oocyte viability and in vitro maturation (IVM), as well as to determine the lethal concentration at 50% (LC50 ) and the maturation inhibition concentration at 50% (MIC50 ). Oocytes were exposed to different MePB concentrations 0 (control), 50, 100, 500, 750 and 1000 μm during 44 h of IVM. Cytoplasmic alterations and reduced cumulus cell expansion were observed in oocytes exposed to MePB; however, viability was not affected. In addition, oocyte maturation decreased in a concentration-dependent manner after exposure to MePB. The estimated LC50 was 2028.38 μm, whereas MIC50 was 780.31 μm. To our knowledge, this is the first study that demonstrates that MePB altered porcine oocyte morphology, and caused a reduction in cumulus cell expansion, both of which resulted in decreased oocyte maturation. Therefore, MePB exposure may be one of the factors involved in fertility impairment in mammals, including that of humans.

Chronic stress induces decreased sperm motility, viability and concentration in stressed males. Also, stress modifies oxidative status and causes apoptosis in testes, as well as a decrease in the epithelial area of seminiferous tubules. However, there are no studies that analyze the alterations caused by stress in testicular cells. Thus, in this study, alterations in the morphology of testicular germ cells caused by different days of chronic stress were assessed. Adult male rats were exposed to stress by immersion in cold water (ICW) daily for 3, 8, 20 or 50 consecutive days. Plasma testosterone and corticosterone were also assessed. Results showed that chronic stress causes loss of germ cells, and alteration of spermatogenesis. Seminiferous tubules from stressed males showed several degenerative signs, such as vacuoles in the basal epithelium, with picnotic indicia; moderate to severe exfoliation of degenerative germinal cells in the tubule lumen was also observed. These alterations were observed in all days of stress in a gradual way, from day 3-50. Testosterone levels were decreased at all those times, and corticosterone concentrations were increased on the same days. These results show that chronic stress causes severe damage to germ cells, which can account for infertility problems in males. These alterations are related to a decrease in testosterone as well as an increase in corticosterone caused by stress.

The administration of Mesquite pod extract (containing mesquitol, daidzein and genistein) to female and male rats disrupts reproductive variables. However, its effect on fertility is not known. This study evaluated fertility in male and female rats treated with mesquite pod extract, comparing its effects with those of daidzein and estradiol. The following treatments were given for 30 days to groups of female and male rats: vehicle, mesquite pod extract, DAI and E2. Treatments were administered subcutaneously for 30 days. These extract disrupted both the female and male sexual behavior in a similar way to DAI, but less than E2. Mesquite pod extract increased the number of days in estrus and decreased lordosis intensity during proestrus. Mesquite pod extract-treated males showed lower testicular and glandular weights, as well as decreased sperm motility, viability and count. In females treated with mesquite pod extract, the number of pups was lower than in control females, and 10 to 20% of pups were dead. These effects were similar to those with DAI-treatment. Despite the lower sperm quality, the fertility of mesquite pod extract- and DAI-treated males seem not to be disrupted, as they could impregnate control females. These results show that mesquite pod extract can disrupt female but not male fertility. Key words: Mesquite pod extract, daidzein, fertility, offspring, sexual behavior, phytoestrogens.

Perfluorohexane sulfonate (PFHxS) is one of the most abundant perfluorinated compounds in the environment. Exposure to this compound has been correlated to a decrease in human fertility, although the molecular and cellular mechanisms underlying this correlation have not been described. The adverse reproductive effects of PFHxS could be based on alterations in oocyte maturation, the process rendering oocytes competent for fertilization. The aim of this study was to evaluate the effect of PFHxS on porcine oocyte viability and maturation in vitro, as well as on gap-junctional intercellular communication (GJIC) in cumulus–oocyte complexes (COCs), oocyte mitochondrial membrane potential (mΔΨ) and DNA damage in cumulus cells, as possible mechanisms of action. PFHxS caused cytotoxicity (medium lethal concentration, LC50 = 329.1 μM) and inhibition of oocyte maturation (medium inhibitory concentration, MIC50 = 91.68 μM). GJIC was not affected in exposed COCs. However, the mitochondrial membrane potential was significantly decreased in PFHxS-exposed oocytes at the germinal vesicle breakdown (GVBD) stage. In addition, exposure to PFHxS induced DNA damage in cumulus cells. Thus, inhibition of oocyte maturation by PFHxS could be attributed to a decreased oocyte mΔΨ at the GVBD and to DNA damage of the cumulus cells that support the oocyte.

Background In order to improve ICSI, appropiate sperm selection and oocyte activation is necessary. The objective of the present study was to determine the efficiency of fertilization using ICSI with chemically activated ovine oocytes and sperm selected by swim up (SU) or swim up + zona pellucida (SU + ZP) binding. Results Experiment 1, 4–20 replicates with total 821 in vitro matured oocytes were chemically activated with ethanol, calcium ionophore or ionomycin, to determine oocyte activation (precense of one PN). Treatments showed similar results (54, 47, 42 %, respectively) but statistically differents (P 0.05). Conclusions Chemical activation induces higher ovine oocyte activation than mechanical activation. Ethanol slightly displays higher oocyte activation than calcium ionophore and ionomicine. Sperm selection with SU + ZP increased AR/A and AR/D rates in comparison with SU in fresh and frozen-thawed sperm. According to this, in terms of fertilization rates, chemical activation after ICSI increased oocyte PN formation compared to mechanical activation. Also, fresh sperm treated with SU and SU + ZP were significantly different than frozen-thawed sperm, but between sperm treatments no significant differences were obtained.

This study was designed to evaluate the capacity of vitrified-warmed porcine immature oocytes to mature and to be fertilized using in vitro fertilization or intracytoplasmic sperm injection, and to determine the subsequent embryo development. Immature oocytes were vitrified using ethylene glycol and dimethylsulphoxide as cryoprotectants and the Cryolock method. After warming oocytes were cultured 44 h for maturation. Oocytes were randomly distributed in three treatment groups and subjected to in vitro fertilization (Experiment 1) or intracytoplasmic sperm injection (Experiment 2) procedures. The results indicate that the embryo development was higher in denuded oocytes co-cultured with granulosa cells (NkO-CC group) fertilized by in vitro fertilization or intracytoplasmic sperm injection compared to cumulus-cell oocyte complexes (COCs group), showing no significant differences with control. Vitrified denuded oocytes matured with a co-culture system NkO-CC group, displayed higher cleavage rate and blastocyst production than vitrified COCs group. Blastocysts were successfully obtained after IVF and ICSI procedures; however, the development to the blastocyst stage was better after IVF. These results show that the vitrification-warming media, the employment of a granulosa cell co-culture system and the Cryolock method during vitrification, increased the nuclear and cytoplasmic maturation of vitrified porcine immature oocytes. Further experiments are required to enhance porcine embryo production after vitrification.

In 1985, the first study achieving ice-free cryopreservation by vitrification of mammalian cells was performed. Up to now, many studies have been carrying out to increase the oocytes and embryos survival rate in order to obtain live offspring. Immature oocytes vitrification continues to be a wide area of research because one of the greatest challenges to reproductive cryobiologists is to establish a vitrification-warming procedure that allows the banking of oocytes and the improvement of assisted reproductive technologies. Vitrification is used to cryopreserve cells, tissues, organs and even organisms. This technique requires the use of cryoprotectant agents (CPAs) added at a high concentration, and devices with high cooling–warming rates to minimize ice crystal formation and subsequently cell damage; but its use should be the most suitable because they are highly toxic. There are few repots testing immature oocyte viability, maturation and fertilization rates after warming. Most studies are conducted to matured oocytes and embryos obtained in vivo rather than in vitro. So far, there are no reports testing the cryolock device, and only one study has reported the birth of live offspring from vitrified porcine immature oocytes with the solid surface vitrification (SSV) method. Therefore, this study was designed to evaluate the efficiency of a vitrification–warming procedure using the cryolock device as well as the effect of the co-culture of vitrified immature oocytes with fresh granulosa cells to improve in vitro maturation (IVM) and embryo development (ED) by in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) techniques. In conclusion, maintaining the viability of granulosa cells is essential to improve maturation and fertilization rates in vitrified immature oocytes. En 1985 se logró el primer estudio de criopreservación sin la formación de cristales de hielo a través de la vitrificación de células de mamíferos. Hasta ahora, muchos estudios se han realizado para aumentar la tasa de supervivencia de los ovocitos y embriones con la finalidad de obtener crías vivas. La vitrificación de ovocitos inmaduros sigue siendo una gran área de investigación debido a que uno de los mayores retos para los criobiólogos de la reproducción es establecer un procedimiento de vitrificacióncalentamiento, que permita la creación de bancos de ovocitos y mejoras en las técnicas de reproducción asistida. La vitrificación se utiliza para criopreservar células, tejidos, órganos e incluso organismos. Esta técnica requiere del uso de agentes crioprotectores (CPAs) añadidos a una alta concentración y recipientes con altas velocidades de enfriamiento-calentamiento para minimizar la formación de cristales de hielo y daño celular. Sin embargo, su uso debe ser el más adecuado debido a su alta toxicidad. Existen pocos trabajos que reportan las tasas de viabilidad, maduración, fertilización de los ovocitos inmaduros después del calentamiento. La mayoría de los trabajos se han realizado en ovocitos maduros y embriones obtenidos in vivo en lugar de in vitro. Hasta el momento, no hay reportes donde utilicen el recipiente cryolock durante el proceso de vitrificación, y sólo un estudio ha reportado el nacimiento de crías vivas a partir de ovocitos inmaduros porcinos vitrificados con el método de vitrificación en superficie sólida (SSV). Por lo que, este estudio fue diseñado para evaluar la eficiencia de un procedimiento de vitrificacióncalentamiento utilizando el recipiente cryolock así como el efecto del co-cultivo de ovocitos inmaduros vitrificados con células de la granulosa frescas para incrementar las tasas de maduración in vitro (MIV) y desarrollo embrionario (DE) mediante las técnicas de fertilización in vitro (FIV) e inyección intracitoplasmática de espermatozoides (ICSI). En conclusión, mantener la viabilidad de las células de la granulosa es esencial para mejorar las tasas de maduración y fertilización de los ovocitos inmaduros vitrificados.

This study was designed to evaluate the efficiency of two oocyte vitrification-warming procedures using two different devices: Superfine Open Pulled Straws (SOPS) and Cryolock, as well as the effect of the co-culture of vitrified immature oocytes with fresh granulosa cells to improve in vitro maturation (IVM). Immature oocytes were vitrified with two procedures: A) Oocytes were exposed to an increasing concentration of ethylene glycol (EG) from 4% to 35% with 0.5 M trehalose. They then, were loaded in SOPS or Cryolock. For warming, oocytes were exposed to decreasing concentrations of trehalose 0.3, 0.2 and 0.1 M for IVM. B) Oocytes were exposed to two mixtures of EG and dimethylsulfoxide (Me2SO), at 7.5% and 16%, both with 0.4 M of sucrose and then loaded in SOPS or Cryolock and stored in liquid nitrogen. For warming, oocytes were exposed to a single concentration of sucrose 0.5M. After warming, viability was determined; and after 44 h of IVM both viability and meiotic stages were evaluated. The results indicate no significant differences between procedures A and B with SOPS in all maturation stages, reaching a maximum maturation rate of 21%. As to Cryolock, significant differences were observed between both procedures, being procedure B, more efficient with a yield of 38% in MII stage and increased to 49% due to the co-culture with fresh granulosa cells. In conclusion, viability and maturation rates were improved with Cryolock and procedure B with the co-culture system in vitrified immature oocytes.

En los últimos años se ha registrado una alta incidencia de problemas de fertilidad y defectos en el desarrollo sexual masculino,que podría estar relacionado a la contaminación ambiental. El desarrollo embrionario y fetal comprende una serie de eventos importantes para la formación de un nuevo individuo, siendo un período susceptible a agresiones químicas por contaminantes. El sulfonato de perfluorooctano (PFOS), es un contaminante ambiental que en el humano ha sido detectado en suero sanguíneo, líquido amniótico, sangre del cordón umbilical y diferentes órganos como hígado, riñón y en pequeñas concentraciones en testículo y cerebro. Tanto el humano, como muchas otras especies animales están expuestas a PFOS desde etapas tempranas del desarrollo embrionario. En el varón se ha descrito que PFOS altera la proporción de espermatozoides normales,disminuye la concentración espermática y altera la concentración de hormonas sexuales. Sin embargo,actualmente no existen estudios que describen las alteraciones en el individuo macho causadas por la exposición a PFOS durante la gestación, por lo que en,este trabajo se utilizó un modelo murino,para evaluar las alteraciones en el crecimiento, desarrollo sexual y calidad espermática de ratones macho expuestos a PFOS durante el desarrollo fetal. Se obtuvieron ratones hembra gestantes de la cepa CD-1, a las cuales se les administró una dosis de PFOS (5 mg/kg/día) por vía intraperitoneal en las etapas 10.5, 12.5 y 16.5 dpc, que abarcan el periodo de gametogénesis gestacional en las crías.Demostrando que la exposición gestacional a PFOS puede alterar patrones morfométricos en la descendencia masculina como la talla y la distancia anogenital, además, en ratones de 90 días se encontraron alteraciones en la viabilidad, movilidad y concentración espermática.y aunque no afecta la capacidad reproductiva, se encontró una disminución en el número de crías, que puede ser explicada por la afectación en la calidad espermática.

La producción de embriones in vitro (PEIV) es una tecnología que permite obtener una gran cantidad de embriones en condiciones de laboratorio. La primera etapa que es la maduración in vitro (MIV) es una de las más imprescindibles para el éxito del potencial embrionario, sin embargo, el estrés oxidativo es considerado como un interruptor de la competencia del ovocito y para disminuir sus efectos sobre la viabilidad celular se han probado compuestos conocidos como antioxidantes, los cuales procuran disminuir los niveles suprafisiológicos de moléculas oxidantes conocidas como especies reactivas de oxígeno (ERO). La mayoría de los trabajos en donde suplementan medios de cultivo con antioxidantes son realizados en especies animales de interés económico. Hasta el momento la mayoría de los suplementos son agregados al medio de MIV esperando que los resultados positivos se vean reflejados en la calidad del desarrollo embrionario. Por lo tanto, los programas para la PEIV buscan estrategias que permitan optimizar los medios de cultivo y disminuir los niveles de ERO para proteger al ovocitoy/o embrión del estrés oxidantivo. Teniendo en cuenta la importancia de este último y su impacto en la competencia del ovocito, es trascendente el estudio a profundidad de compuestos que puedan ayudar a establecer mejores protocolos para la PEIV y la transferencia embrionaria. In vitro embryo production (IVP) is a technology that makes it possible to obtain manyembryos under laboratory conditions.The first stage, which is in vitro maturation (IVM), is one of the most essential for the success of the embryonic potential, however, oxidative stress is considered as a switch of the oocyte's competition and to reduce its effects on cell viability. A myriad of compounds known as antioxidants have been tested, which seek to lower the supra-physiological levels of oxidant molecules known as reactive oxygen species (ROS). Most of the works in which the culture media are supplemented with antioxidants are carried out in species of economic interest. Until now, most of the additives are added to the IVM medium, hoping that the positive results will be reflected in the quality of embryonic development. Therefore, IVP programs seek strategies that optimize culture media and decrease ROS levels to protect the oocyte from oxidative stress. Consideringthe importance of the latter and its impact on the competence of the oocyte. The in-depth study of compounds that can help establish better protocols for IVP and embryo transfer is conclusive.

Introducción:Los parabenos (PBs) son compuestos ampliamente utilizadoscomo antimicrobianos y fungicidasen alimentos y productos del cuidado personal. Su uso indiscrimidado ha permitido que sean detectados en diferentes ecosistemas, por lo que tanto humanos como otros organismos se encuentran altamente expuestos. El metil-parabeno (MePB), comparado con otros PBs, es el más abundante en alimentos, productos de cuidado personal y productos para el cuidado de bebés. El uso de los PBs podría estar ligado a desórdenes hormonales y provocarproblemas de fertilidad, ya que recientemente han sido catalogados como disruptores endocrinos (DEs). El conocimento de los efectos que el MePB puede ocasionar es de gran importancia, ya que se conoce pocoen cuanto a sus efectos reproductivos. El objetivo de este trabajo fue evaluar el efecto del MePB en concentraciones de 0, 50, 100, 500, 750 y 1000 μM de la MIV enla viabilidad y la maduración in vitro(MIV) de ovocitos porcinos, así como determinar la concentración letal 50 (CL50) y la concentración de la inhibición de la maduración 50 (CIM50). Resultados:Después de laMIV, se observó una dispersión de las células del cúmulo así como alteraciones citoplasmáticas en los ovocitos expuestos a las diferentes concentraciones de MePB; sin embargo, la viabilidad no se vió afectada en lasconcentraciones evaluadas. Además, la maduración de los ovocitos disminuyóde forma concentración-dependiente despuésde suexposición al MePB. La CL50estimada fue de 2028.38 μM, mientras que la CIM50fue 780.31 μM. Conclusión: Hasta nuestro conocimiento, este es el primer trabajo que demuestra el efecto del MePB sobre la maduración de los ovocitos porcinos in vitro.Los resultados demuestran que el MePB alteróla morfología de los ovocitos, afectando la expansión de las células del cúmulo y disminuyendo su maduración,por lo quela exposición al MePB podría ser una de las causas de infertilidad en los mamíferos, incluido el humano. Background: Parabens (PBs) are compounds widely used in food and personal care products industry as antimicrobials and preservatives. Their indiscriminate use has allowed them to be detected in different ecosystems so that humans and other organisms are highly exposed. Methylparaben (MePB), compared with other PBs, is mostly detected in food, personal, and baby care products. PBs could be linked to hormonal disorders generation and fertility impairment since they have been recently classified as endocrine disruptors. The knowledge of the effects that MePB can exert is of great importance since, in terms of reproduction, information is limited. Therefore, the objective of this study was to evaluate the effect of MePB on concentrations of 0, 50, 100, 500, 750 y 1000 μM porcine oocyte viability and in vitromaturation(IVM), as well as to determine the lethal concentration 50 (LC50) and maturation inhibition concentration 50 (MIC50). Results:AfterIVM, dispersed cumulus cells and cytoplasmic alterations were observed in oocytes exposed to MePB; however, viability was not affected in the concentration range assayed. In addition, oocyte maturation decreased in a concentration-dependent manner after exposure to MePB. The estimated LC50was 2028.38 μM, whereas MIC50was 780.31 μM. Conclusions: As far as we know, this is the first study demonstrating that MePB can cause deleterious effects to the in vitro maturation of porcine oocytes. This study demonstratesthat MePB altered porcine oocytes morphology and caused cumulus cells dispersion, which resulted in decreased oocyte maturation. Therefore, MePB exposure may be one of the factors involved in fertility impairment in mammals, including the human.

La vitrificación es una técnica de criopreservación empleada principalmente en gametos. Esta técnica tiene como finalidad mantener la viabilidad celular, su funcionalidad y su potencial de desarrollo a -196°C. Durante este proceso se requiere de la adición de agentes crioprotectores (CPAs), que son sustancias que confieren protección a las células durante el proceso. La selección de la técnica de vitrificación adecuada; depende de la especie animal, el tipo celular y la naturaleza de los CPAs que se utilizan. La toxicidad producida por el uso de los CPAs durante la vitrificación puede causar alteraciones en los microfilamentos (MF) y en la cromatina (CR), esto puede repercutir en la viabilidad, la maduración, la fertilización y el desarrollo embrionario (DE). En estudios previos, se ha evaluado el efecto de la vitrificación en ovocitos en estados de VG y MII, cigotos y blastocistos en la misma etapa de desarrollo en la que fueron vitrificados. Además, estos estudios atribuyeron el papel de los MF y la CR a la disminución de la fertilización y la producción embrionaria, pero no se evaluó la distribución de estas estructuras. Por lo que en este estudio se examinó de qué manera la vitrificación de ovocitos porcinos inmaduros afecta a la distribución de los MF y la CR, así como su repercusión en el DE temprano. La vitrificación, produjo alteraciones en la distribución de la actina cortical (AC) y desarreglo en la compactación de la CR en los embriones, por lo que la fertilización y el DE temprano disminuyeron The vitrification technique is a type of cryopreservation used mainly in gametes. This technique alows to maintain cell viability, functionality and development potential at -196 ° C. During this technique, the addition of cryoprotective agents (CPAs) is required, which are substances that confer protection to the cells during cooling and warming. Also, the animal species, the cell type and the nature of the CPAs that are used to select the appropriate vitrification technique depends on its success. The toxicity produced by the CPAs during vitrification can cause alterations in microfilaments (MF) and chromatin (CR), also affecting cell viability, maturation, fertilization and embryo development (ED). Previous studies, have evaluated the vitrification effects on oocytes in VG and MII stages, zygotes and blastocysts in the same stage of development in which they were vitrified. In addition, these studies evaluated the role of MF and CR based on the rate of fertilization and embryo production, but not on the distribution of these structures. Therefore, this study was designed to evaluate how the vitrification of immature porcine oocytes affects the distribution of MF and CR, as well as its repercussion in the early ED. In the present study, alterations in the distribution of cortical actin (CA) and disarrangement in the compaction of CR in embryos produced from vitrified oocytes were obtained, reducing the fertilization and early ED rates