Impacto de la fragmentación del ADN espermático y la tecnología de microfluidos en los resultados de fecundación in vitro / Impact of sperm DNA fragmentation and microfluidic technology on IVF outcomes

Un diagnóstico acertado en los pacientes infértiles es clave para determinar el tratamiento de elección en un programa de reproducción asistida. En el caso del varón, el diagnóstico inicial se basa en el resultado del seminograma, el cual permite hallar problemas relacionados con la esterilidad de la pareja, pero es insuficiente para la correcta detección de la infertilidad masculina, puesto que no predice la capacidad funcional de los espermatozoides. En los últimos años, han aparecido múltiples estudios que relacionan la integridad del ADN espermático con la fertilidad. Al mismo tiempo, los laboratorios de fecundación in vitro (FIV) tienen a su alcance nuevos métodos de selección del esperma, como los microfluidos, que ayudarían a disminuir el grado de fragmentación del ADN espermático (SDF) en la muestra. En este trabajo revisamos el impacto que tienen la SDF y el uso de los dispositivos de microfluidos en los resultados de FIV con base en una selección de estudios relevantes publicados hasta febrero de 2023.(AU)

An accurate diagnosis in infertile patients is key to determine the treatment of choice in an assisted reproduction program. In the case of the male, the initial diagnosis is based on the result of the semen analysis. The semen analysis can detect problems related to the couple's infertility, but it is insufficient for the correct diagnosis of male infertility, since it does not predict the functional capacity of the spermatozoa. In recent years, multiple studies have appeared that relate sperm ADN integrity to fertility. At the same time, IVF laboratories have within their reach new methods of sperm selection, such as microfluidics, which would make it possible to reduce the degree of ADN fragmentation in the sample. In this paper we review the impact of sperm ADN fragmentation and the use of microfluidic devices on IVF outcomes based on a selection of relevant studies published up to February 2023.(AU)

Correlation of DNA fragments with routine semen parameters and lifestyle and their impact on assisted reproductive outcomes

Objective: This study investigated the correlation between sperm DNA integrity and routine semen evaluation parameters in male infertile patients, the influencing factors, and the impact of the DNA fragmentation index (DFI) on embryo quality and clinical outcomes in in vitro fertilization and embryo transfer (IVF-ET). Methods: Sperm DFI and semen routine parameters of 6160 infertile men admitted between June 2017 and June 2018 were analyzed. Patients were divided into three groups according to their DFI: low-DFI (DFI<15%), medium-DFI (15%30%). The correlations of DFI with patients’ age, sperm concentration, sperm percentage of forward movement and sperm percentage of normal shape were analyzed. The clinical data of 5040 infertile couples who received IVF treatment between June 2016 and 2021 and had embryos transferred in a fresh cycle were reviewed. The fertilization rate, cleavage rate, blastocyst rate, and pregnancy rate in different DFI groups were compared. Results: Semen evaluation parameters (concentration, spermatozoa with progressive motility, and the normal morphology rate), the high-quality embryo rate, blastocyst development rate, and pregnancy rate in the high-DFI group were significantly lower than those in the other two groups. The correlation analysis revealed that sperm DFI was negatively correlated with semen concentration, sperm motility, and normal sperm morphology and positively correlated with the man's age, BMI, and unhealthy habits (smoking and drinking). There was no significant difference in the number of mature eggs and normal fertilization rate among groups. Conclusion: A strong correlation exists between sperm DFI and semen evaluation parameters. Smoking, drinking, and other unhealthy habits lead to an increase in DFI, reducing the high-quality embryo rate and blastocyst development rate and affecting pregnancy outcomes. (AU)

Objetivo: Avaliar a correlação entre a integridade do DNA espermático de pacientes com infertilidade masculina e os parâmetros de avaliação seminal convencional, os fatores que a impulsionam, a influência do índice de fragmentação de DNA (DFI) na qualidade embrionária, o desfecho clínico na fertilização in vitro e a transferência de embriões (fiv-te). Métodos: Foram analisados o DFI espermático e os parâmetros de rotina seminal de 6.160 homens com infertilidade admitidos entre junho de 2017 e junho de 2018. Os pacientes foram divididos de acordo com o DFI em baixo (DFI <15%), médio (15% 30%). Foi analisada a correlação do DFI com a idade dos pacientes, concentração espermática, porcentagem de espermatozoides motilizados para frente e porcentagem de espermatozoides com morfologia normal. Foram revisados dados clínicos de 5.040 casais inférteis que receberam tratamento de fertilização in vitro entre junho de 2016 e 2021 e transferiram embriões no novo ciclo. As taxas de fertilização, clivagem, blastocisto e concepção foram comparadas entre os diferentes grupos de DFI. Resultados: Os índices de avaliação seminal (concentração, espermatozoides com movimento progressivo e morfologia normal), embriões de boa qualidade, desenvolvimento de blastocistos e prenhez foram significativamente menores no grupo com alto DFI. As análises de correlação mostraram que o DFI espermático está negativamente relacionado com a concentração seminal, motilidade espermática, morfologia espermática normal e positivamente relacionado com a idade dos homens, IMC e hábitos adversos (tabagismo e consumo de álcool). Conclusão: Houve forte correlação entre o DFI espermático e os parâmetros de avaliação seminal. Hábitos indesejáveis, como tabagismo e consumo de álcool, levam ao aumento do DFI, diminuição da taxa de embriões de boa qualidade e desenvolvimento de blastocistos e comprometem o desfecho gestacional. (AU)

Microfluidic sperm sorting improves ICSI outcomes in patients with increased values of Double-Strand Breaks in sperm DNA / La selección espermática mediante un dispositivo microfluídico mejora los resultados de ICSI en pacientes con valores elevados de fragmentación de cadena doble en el ADN espermático

Background: Delays in embryo kinetics, implantation failures in ICSI treatments and recurrent miscarriages have been associated with high values of Double-Strand Breaks (DSB) in sperm DNA. While conventional methods for semen preparation have been shown to be inefficient reducing DSB values, Microfluidic Sperm Sorting (MSS) devices are promising tools to reduce this damage. Objective: To study the clinical utility of an MSS device in ICSI treatments when the male partner presents increased DSB values, as compared to the use of conventional methods based on sperm motility. Methods: This retrospective cohort study included 28 infertile couples undergoing ICSI treatments. Only couples where the male partner presented increased values of DSB were included. DSB values were evaluated in semen samples by the Neutral Comet assay. Couples performed a first ICSI cycle using conventional methods for semen preparation (Density Gradients and Swim-up) and a second ICSI cycle using the ZyMōt™ICSI (formerly named FertileChip®) microfluidic device. Embryology and clinical outcomes were compared between ICSI cycles. Results: Semen parameters and the number of obtained and fertilized oocytes did not show differences between ICSI rounds. Clinical outcomes were statistically better when MSS was used: the biochemical pregnancy rate increased 28.31%; the clinical pregnancy rate increased 35.56% and the number of live births increased 35.29%, as compared to the first ICSI cycle in this group of patients. (AU)

Antecedentes: Valores elevados de fragmentación de cadena doble (DSB) en el ADN de los espermatozoides se han asociado con retrasos en la cinética embrionaria, fallos de implantación en ciclos de ICSI y con abortos de repetición. Actualmente no hay evidencias de que los métodos convencionales para la preparación del semen puedan reducir los niveles de DSB. Por el contrario, los nuevos dispositivos microfluídicos de selección espermática (MSS) han mostrado resultados prometedores en cuanto a la reducción de la fragmentación. Objetivo: Evaluar el uso de un dispositivo MSS en ciclos de ICSI donde el varón presenta niveles elevados de DSB, en comparación con el uso de métodos convencionales basados en la selección por motilidad. Métodos: Este estudio retrospectivo ha incluido a 28 parejas infértiles que han realizado ciclos de ICSI y donde se han detectado valores elevados de DSB en la muestra seminal del varón. Los niveles de DSB se han analizado mediante el test Cometa Neutro. Las parejas realizaron un primer ciclo de ICSI utilizando métodos convencionales para la preparación del semen (gradientes de densidad y Swim-up). Posteriormente, las parejas realizaron un segundo ciclo de ICSI utilizando el dispositivo microfluídico ZyMōt™ICSI (antes FertileChip®). Se han comparado los resultados de embriología y los resultados clínicos entre ambos tratamientos. Resultados: No se han encontrado diferencias entre ambos ciclos de ICSI en cuanto a parámetros seminales y el número de ovocitos obtenidos y fecundados. Los resultados clínicos fueron mejores cuando se usó el dispositivo MSS: se observó un incremento del 28,31% en la tasa de embarazo bioquímico, del 35,56% en la tasa de embarazo clínico y del 35,29% en la tasa de nacidos vivos, en comparación con el uso de métodos convencionales. (AU)

Correlation of DNA fragments with routine semen parameters and lifestyle and their impact on assisted reproductive outcomes.

OBJECTIVE: This study investigated the correlation between sperm DNA integrity and routine semen evaluation parameters in male infertile patients, the influencing factors, and the impact of the DNA fragmentation index (DFI) on embryo quality and clinical outcomes in in vitro fertilization and embryo transfer (IVF-ET). METHODS: Sperm DFI and semen routine parameters of 6160 infertile men admitted between June 2017 and June 2018 were analyzed. Patients were divided into three groups according to their DFI: low-DFI (DFI<15%), medium-DFI (15%30%). The correlations of DFI with patients' age, sperm concentration, sperm percentage of forward movement and sperm percentage of normal shape were analyzed. The clinical data of 5040 infertile couples who received IVF treatment between June 2016 and 2021 and had embryos transferred in a fresh cycle were reviewed. The fertilization rate, cleavage rate, blastocyst rate, and pregnancy rate in different DFI groups were compared. RESULTS: Semen evaluation parameters (concentration, spermatozoa with progressive motility, and the normal morphology rate), the high-quality embryo rate, blastocyst development rate, and pregnancy rate in the high-DFI group were significantly lower than those in the other two groups. The correlation analysis revealed that sperm DFI was negatively correlated with semen concentration, sperm motility, and normal sperm morphology and positively correlated with the man's age, BMI, and unhealthy habits (smoking and drinking). There was no significant difference in the number of mature eggs and normal fertilization rate among groups. CONCLUSION: A strong correlation exists between sperm DFI and semen evaluation parameters. Smoking, drinking, and other unhealthy habits lead to an increase in DFI, reducing the high-quality embryo rate and blastocyst development rate and affecting pregnancy outcomes.

Microfluidic sperm sorting improves ICSI outcomes in patients with increased values of Double-Strand Breaks in sperm DNA.

BACKGROUND: Delays in embryo kinetics, implantation failures in ICSI treatments and recurrent miscarriages have been associated with high values of Double-Strand Breaks (DSB) in sperm DNA. While conventional methods for semen preparation have been shown to be inefficient reducing DSB values, Microfluidic Sperm Sorting (MSS) devices are promising tools to reduce this damage. OBJECTIVE: To study the clinical utility of an MSS device in ICSI treatments when the male partner presents increased DSB values, as compared to the use of conventional methods based on sperm motility. METHODS: This retrospective cohort study included 28 infertile couples undergoing ICSI treatments. Only couples where the male partner presented increased values of DSB were included. DSB values were evaluated in semen samples by the Neutral Comet assay. Couples performed a first ICSI cycle using conventional methods for semen preparation (Density Gradients and Swim-up) and a second ICSI cycle using the ZyMot™ICSI (formerly named FertileChip®) microfluidic device. Embryology and clinical outcomes were compared between ICSI cycles. RESULTS: Semen parameters and the number of obtained and fertilized oocytes did not show differences between ICSI rounds. Clinical outcomes were statistically better when MSS was used: the biochemical pregnancy rate increased 28.31%; the clinical pregnancy rate increased 35.56% and the number of live births increased 35.29%, as compared to the first ICSI cycle in this group of patients. CONCLUSIONS: The ZyMot™ICSI microfluidic device improved the reproductive outcomes in couples where the male partner presented increased DSB values, when compared to the use of conventional semen preparation techniques.

Evaluación de la integridad del ADN espermático y su repercusión en los parámetros seminales de varones infértiles / Evaluation of the integrity of sperm DNA and its impact on the seminal parameters of infertile males

Resumen OBJETIVO: Determinar las repercusiones del daño al ADN espermático en los parámetros seminales más estudiados en diagnóstico clínico de varones infértiles. MATERIALES Y MÉTODOS: Estudio de casos y controles, prospectivo y comparativo efectuado en pacientes masculinos atendidos en el Centro Integral de la Mujer y Reproducción Asistida de Puebla, México. Parámetros de estudio: edad, movilidad, morfología, diagnóstico seminal, leucocitos y factor de infertilidad. Los resultados se analizaron con Graphpad Prisma 5.0 y se consideraron estadísticamente significativos con p < 0.005. RESULTADOS: Se estudiaron 110 pacientes: 33 con mala integridad del ADN espermático (grupo 1) y 77 con buena integridad (grupo 2). La concentración espermática y la movilidad tipo A+B en el grupo 2 fue significativamente más alta que en el grupo 1 (p < 0.0001) en donde se registró mayor número de móviles no progresivos e inmóviles. La morfología normal fue más alta en el grupo 2 (p = 0.0063). En los varones menores de 40 años se observó un número significativamente mayor de casos de buena integridad espermática (p = 0.013). El diagnóstico seminal demostró que los varones con mala integridad tuvieron alteraciones espermáticas más severas. Los factores de infertilidad más frecuentes implicados en ambos grupos fueron: aborto de repetición, edad de la pareja, falla previa en la técnica de reproducción asistida, factor masculino severo y factor tubárico. CONCLUSIONES: La mala integridad del ADN espermático tiene repercusiones en la concentración espermática, movilidad y morfología, además de alterar el diagnóstico seminal, pues los varones tuvieron trastornos más severos cuando no hubo algún factor de infertilidad que describiera un comportamiento específico relacionado con la mala integridad espermática.

Abstract OBJECTIVE: To determine the impact of sperm DNA damage on the most studied seminal parameters in clinical diagnosis of infertile males. MATERIALS AND METHODS: Prospective and comparative case-control study that included male patients seen at Centro Integral de la Mujer y Reproducción Asistida de Puebla, Mexico. Study parameters: age, mobility, morphology, seminal diagnosis, leukocytes and infertility factor. The results were analyzed with Graphpad Prism 5.0 and were considered statistically significant with p < 0.005. RESULTS: 110 male patients were studied: 33 patients with poor sperm DNA integrity (group 1) and 77 patients with good integrity (group 2). Sperm concentration and type A + B mobility in group 2 was significantly higher than in group 1 (p <0.0001), where a greater number of non-progressive and immobile mobiles was recorded. The normal morphology was higher in group 2 (p = 0.0063). In men under 40 years of age, a significantly higher number of cases of good sperm integrity was observed (p = 0.013). The seminal diagnosis showed that males with poor integrity had more severe sperm alterations. The most frequent infertility factors involved in both groups were: repeat abortion, age of the couple, previous failure in the technique of assisted reproduction, severe male factor and tubal factor. CONCLUSIONS: The poor integrity of the sperm DNA has repercussions on sperm concentration, mobility and morphology, alters the seminal diagnosis, since males had more severe alterations when there was no infertility factor that described a specific behavior related to poor sperm integrity.

Fragmentación del ADN espermático e infertilidad masculina / Fragmentation of sperm DNA and male infertility

La integridad del ADN de los espermatozoides, es un indicador importante de la fertilidad, se utiliza como una variable adicional para evaluar, junto al espermograma, la calidad de una muestra seminal. En el presente trabajo se describen algunos aspectos de interés relacionados con la fragmentación del ADN espermático, cuya etiología es multifactorial y está relacionada con factores intrínsicos, como el empaquetamiento anormal de la cromatina durante la espermiogénesis, la apoptosis defectuosa antes de la eyaculación y la producción excesiva de especies reactivas del oxígeno; y con factores extrínsecos, como son los cambios en los hábitos de vida, la exposición a agentes tóxicos, así como la edad avanzada. La fragmentación está asociada al deterioro de las variables seminales, además afecta la fertilidad natural y la realizada por tratamientos de reproducción asistida, por lo que la implementación en los laboratorios de Andrología, de la tecnología para detectar si el ADN de los espermatozoides está íntegro o fragmentado, incorpora un nuevo conocimiento en el estudio de los hombres con trastornos de fertilidad, lo que contribuye a mejorar el diagnóstico y pronóstico de la infertilidad masculina. Para realizar este trabajo se revisaron 122 artículos, de los cuales 84 cumplieron con los criterios de calidad esperados. La búsqueda se realizó a través de los buscadores habituales(AU)

Sperm DNA´s integrity is an important indicator of fertility and it is used as an additional variable to evaluate the quality of a seminal sample, together with the spermogram. This paper describes some interesting aspects related to the fragmentation of sperm DNA, whose etiology is multifactorial and is related to intrinsic factors, such as the abnormal packing of chromatin during spermiogenesis, defective apoptosis before ejaculation, and the excessive production of oxygen´s reactive species; and with extrinsic factors, such as changes in life habits, exposure to toxic agents, as well as aging. Fragmentation is associated with the deterioration of seminal variables, it also affects natural fertility and that produced by assisted reproduction treatments, so the implementation in the andrology laboratories of the technology to detect if the DNA of the sperm is intact or fragmented incorporates a new knowledge in the study of men with fertility disorders, which contributes to improve the diagnosis and prognosis of male infertility. To carry out this work, 122 articles were reviewed, of which 84 met the expected quality criteria. The search was made through the usual search engines(AU)

Fragmentación del ADN espermático e infertilidad masculina / Fragmentation of sperm DNA and male infertility

La integridad del ADN de los espermatozoides, es un indicador importante de la fertilidad, se utiliza como una variable adicional para evaluar, junto al espermograma, la calidad de una muestra seminal. En el presente trabajo se describen algunos aspectos de interés relacionados con la fragmentación del ADN espermático, cuya etiología es multifactorial y está relacionada con factores intrínsicos, como el empaquetamiento anormal de la cromatina durante la espermiogénesis, la apoptosis defectuosa antes de la eyaculación y la producción excesiva de especies reactivas del oxígeno; y con factores extrínsecos, como son los cambios en los hábitos de vida, la exposición a agentes tóxicos, así como la edad avanzada. La fragmentación está asociada al deterioro de las variables seminales, además afecta la fertilidad natural y la realizada por tratamientos de reproducción asistida, por lo que la implementación en los laboratorios de Andrología, de la tecnología para detectar si el ADN de los espermatozoides está íntegro o fragmentado, incorpora un nuevo conocimiento en el estudio de los hombres con trastornos de fertilidad, lo que contribuye a mejorar el diagnóstico y pronóstico de la infertilidad masculina. Para realizar este trabajo se revisaron 122 artículos, de los cuales 84 cumplieron con los criterios de calidad esperados. La búsqueda se realizó a través de los buscadores habituales(AU)

Sperm DNA´s integrity is an important indicator of fertility and it is used as an additional variable to evaluate the quality of a seminal sample, together with the spermogram. This paper describes some interesting aspects related to the fragmentation of sperm DNA, whose etiology is multifactorial and is related to intrinsic factors, such as the abnormal packing of chromatin during spermiogenesis, defective apoptosis before ejaculation, and the excessive production of oxygen´s reactive species; and with extrinsic factors, such as changes in life habits, exposure to toxic agents, as well as aging. Fragmentation is associated with the deterioration of seminal variables, it also affects natural fertility and that produced by assisted reproduction treatments, so the implementation in the andrology laboratories of the technology to detect if the DNA of the sperm is intact or fragmented incorporates a new knowledge in the study of men with fertility disorders, which contributes to improve the diagnosis and prognosis of male infertility. To carry out this work, 122 articles were reviewed, of which 84 met the expected quality criteria. The search was made through the usual search engines(AU)

Modelo predictivo de fragmentación de ADN espermático usando parámetros evaluados en un espermatograma

Introducción: El análisis de espermatograma es utilizado como una prueba diagnóstica de la calidad seminal. Recientemente, el análisis de fragmentación espermática ha tomado importancia dado los diversos estudios que han demostrado que la integridad del ADN en el espermatozoide afectaría los resultados clínicos en los tratamientos de reproducción asistida. Objetivos: Identificar las variables analizadas en un espermatograma que predecirían independientemente el índice de fragmentación de ADN espermático (IFE). Diseño: Estudio retrospectivo, comparativo. Instituciones: Grupo PRANOR, Reprogenetics Latinoamerica, Clínica Concebir, Lima, Perú. Material biológico: Espermatozoides. Métodos: Se comparó variables individuales y dos modelos: el primero consideró el porcentaje de vialidad espermática y la edad del paciente; el segundo modelo incluyó el porcentaje de espermatozoides motiles y la edad. Se hizo análisis de regresión logística. Principales medidas de resultados: Viabilidad espermática, edad. Resultados: El análisis multivariado demostró que los dos modelos fueron significantemente superiores a las variables individuales (p<0,01). El primer modelo tuvo valores de coeficiente no estandarizados (IC95%) de 0,200 (0,082 a 0,318) y -0,146 (-0,206 a -0,086), respectivamente. El segundo modelo tuvo valores de coeficiente no estandarizados (IC95%) de -0,099 (-0,157 a -0,042) y 0,219 (0,99 a 0,339), respectivamente. El análisis de regresión logística demostró que el porcentaje de viabilidad espermática y la edad del paciente predijeron la probabilidad de tener un IFE superior al 30% con valores de coeficiente no estandarizados de edad de IC95% 0,034 (0,015 a 0,053) y porcentaje de viabilidad de -0,043 (0,034 a 0,052). Adicionalmente, el segundo modelo tuvo IC95% de -0,04 (-0,031 a -0,049) y 0,035 (0,017 a 0,053), respectivamente. Finalmente, una curva ROC construida para determinar la superioridad de algún modelo sobre las variables individuales demostró que las áreas bajo la curva (ABC) del modelo 1 (edad y viabilidad espermática) fue 0,727 (IC95% = 0,665 a 0,790) y el modelo 2 (edad y motilidad total de espermatozoides) 0,675 (IC95% = 0,606 a 0,744), comparadas con las ABC del porcentaje de viabilidad espermática = 0,295 (IC95% = 0,229 a 0,362), motilidad total de espermatozoides ABC = 0,333 (IC95% = 0,264 a 0,403) y edad de paciente con ABC 0,584 (IC95% = 0,510 a 0,658). Conclusiones: La edad, motilidad y viabilidad espermática correlacionaron de manera independiente con el IFE y, por tanto, estas variables podrían ser usadas como predictores del porcentaje de fragmentación de ADN.

Introduction: The spermatogram is used as a test of seminal quality. Recently, the sperm fragmentation test has demonstrated importance as sperm DNA integrity would affect clinical results in assisted reproduction treatments. Objectives: To determine spermatogram variables that would independently predict sperm DNA fragmentation index (SFI). Design: Retrospective, comparative study. Settings: Grupo PRANOR, Reprogenetics Latinoamerica, Clinica Concebir, Lima, Peru. Biologic material: Sperm. Methods: Individual variables and two models were compared: the first model considered percentage of sperm viability and patients age; the second model included percentage of motile sperms and age. Logistic regression analysis was done. Main outcome measures: Sperm viability, age. Results: Multivariate analysis showed that both models were significantly superior to individuals variables (p<0.01). The first model had non standardized coefficient values (95%CI) respectively of 0.200 (0.082 to 0.318) and -0.146 (-0.206 to -0.086). The second model had non standardized coefficient values (95%CI) respectively of -0.099 (-0.157 to -0.042) and 0.219 (0.99 to 0.339). Logistic regression analysis showed that the percentage of sperm viability and patients age predicted the probability of having an SFI over 30% with age non standardized coefficient values of 95%IC 0.034 (0.015 to 0.053) and viability percentage of -0.043 (0.034 to 0.052). Additionally the second model had 95%CI respectively of -0.04 (-0.031 to -0.049) and 0.035 (0.017 to 0.053). Finally, a ROC curve to determine the superiority of some model over individual variables showed that areas under the curve (ABC) of model 1 (age and sperm viability) was 0.727 (95%CI = 0.665 to 0.790) and model 2 (age and sperm total motility) 0.675 (95%CI = 0.606 to 0.744), compared with ABC of percentage of sperm viability = 0.295 (95%CI = 0.229 to 0.362), sperm total motility ABC = 0.333 (95%CI = 0.264 to 0.403) and patients age with ABC 0.584 (95%CI = 0.510 to 0.658). Conclusions: Age, and sperm motility and viability independently correlated with SFI and consequently these variables could be used as predictors of DNA fragmentation percentage.