False positive rate of an arrayCGH platform for single-cell preimplantation genetic screening and subsequent clinical application on day-3

In this work, false positive rate of an arrayCGH platform for its use in day-3 single-blastomere analysis was calculated. For this purpose, 38 embryos diagnosed as abnormal on day-3 by FISH were re-biopsied on day-4. Single-cell day-4 arrayCGH diagnosis was then performed. A successful amplification was obtained in 97.4 % (37/38) of the day-4 cells analysed by arrayCGH. Day-3 FISH and day-4 arrayCGH diagnosis were concordant in 35/37 cases. The two discordant embryos were spread and all the cells from each embryo were re-analysed by FISH on day 5. The same error rate (2.7 %) for day-3 FISH and day-4 arrayCGH was obtained when comparing day-5 FISH re-analysis. After this pre-clinical phase, the platform was used for day-3 arrayCGH clinical application in 320 patients (1,760 embryos). Day-3 amplification rate was 98.6 %. An optimal reproductive outcome was obtained when applying arrayCGH to a clinical program: clinical pregnancy rate per cycle of 38.4 % and 60.3 % per transference were obtained, with an implantation rate of 53.5 %. Overall miscarriage rate was 10.6 %. Additionally, day-5 FISH re-analysis was performed in 42 of the embryos from the clinical phase, obtaining a concordance rate of 97.6 % with day-3 arrayCGH.Electronic supplementary materialThe online version of this article (doi:10.1007/s10815-012-9918-4) contains supplementary material, which is available to authorized users

New Tools for Embryo Selection: Comprehensive Chromosome Screening by Array Comparative Genomic Hybridization

The objective of this study was to evaluate the usefulness of comprehensive chromosome screening (CCS) using array comparative genomic hybridization (aCGH). The study included 1420 CCS cycles for recurrent miscarriage (n = 203); repetitive implantation failure (n = 188); severe male factor (n = 116); previous trisomic pregnancy (n = 33); and advanced maternal age (n = 880). CCS was performed in cycles with fresh oocytes and embryos (n = 774); mixed cycles with fresh and vitrified oocytes (n = 320); mixed cycles with fresh and vitrified day-2 embryos (n = 235); and mixed cycles with fresh and vitrified day-3 embryos (n = 91). Day-3 embryo biopsy was performed and analyzed by aCGH followed by day-5 embryo transfer. Consistent implantation (range: 40.5–54.2%) and pregnancy rates per transfer (range: 46.0–62.9%) were obtained for all the indications and independently of the origin of the oocytes or embryos. However, a lower delivery rate per cycle was achieved in women aged over 40 years (18.1%) due to the higher percentage of aneuploid embryos (85.3%) and lower number of cycles with at least one euploid embryo available per transfer (40.3%). We concluded that aneuploidy is one of the major factors which affect embryo implantation

Estudio de anomalías meióticas y aneuploidías en pacientes con azoospermia secretora

La contribución de anomalías meióticas a la infertilidad masculina humana ha sido reconocida desde hace tiempo. Los errores meióticos ligados a los procesos de sinapsis, recombinación y reparación del ADN, interfieren en el proceso normal de la meiosis pudiendo dar lugar a una segregación anómala de los cromosomas homólogos (meiosis I) ó las cromátidas hermanas (meiosis II) y generar espermatozoides portadores de alteraciones cromosómicas numéricas (aneuploides o diploides). El objetivo principal de la presente tesis doctoral fue estudiar las diferencias en el proceso de la meiosis entre varones de fertilidad probada y varones infértiles con azoospermia secretora de origen idiopático, diferenciando dentro de este grupo aquellos que inicialmente fueron criptozoospermicos y evolucionaron a azoospérmicos. Mediante técnicas de inmunocitogenética e hibridación in situ fluorescente se evaluó la progresión meiótica, la frecuencia de recombinación meiótica y la longitud de los complejos sinaptonémicos, en espermatocitos primarios en estadío de paquitene, así como la incidencia de disomías y diploidías en espermatozoides testiculares en ambos grupos. Los varones con azoospermia secretora presentaron una acumulación de espermatocitos primarios en los estadios iniciales de la profase I, con menor porcentaje de meiocitos que superaron el punto de control de paquitene. La frecuencia de recombinación meiótica en los pacientes con azoospermia secretora de origen idiopático mostró una disminución significativa al compararla con el grupo control de pacientes post-vasectomizados. Sin embargo, en el grupo de pacientes criptozoospérmicos esta disminución no mostró diferencias significativas con el grupo control. Se observó una variación de la frecuencia de recombinación entre individuos, tanto en los grupos de estudio, como en el grupo control que no se correlacionó ni con la edad de los pacientes, ni con el tiempo transcurrido desde la vasectomía en el caso de los individuos pertenecientes al grupo control. El estudio de la frecuencia específica de recombinación meiótica por cromosoma mostró en los pacientes con azoospermia secretora una disminución en los niveles de recombinación para la mayoría de cromosomas analizados, independientemente de su tamaño y estructura. La medición de la longitud total de los complejos sinaptonémicos en los espermatocitos primarios en estadio de paquitene resultó similar en el grupo control y en los grupos de estudio, por lo que no se observó una correlación entre longitud de complejos sinaptonémicos con los niveles de recombinación. En pacientes con azoospermia secretora se observó un incremento significativo en la incidencia de disomías y diploidías para todos los cromosomas analizados al compararlo con el grupo control. Los pacientes criptozoospérmicos sólo mostraron este incremento significativo en las disomías para el cromosoma 18, los cromosomas sexuales y para la incidencia de diploidías. El 81,2% de los pacientes del grupo de azoospermia secretora que presentaron unos niveles de recombinación meiótica significativamente inferiores a los del grupo control mostraron un incremento de anomalías cromosómicas en sus espermatozoides.The contribution of meiotic abnormalities in human male infertility has been recognized. Meiotic errors linked to synaptic processes, recombination and repair of DNA interfere with the normal process of meiosis and can produce an abnormal segregation of homologous chromosomes (meiosis I) or sister chromatids (meiosis II),resulting in the generation of sperm with numerical chromosome abnormalities (aneuploid or diploid). The aim of this PhD project was to assess the differences in the process of meiosis between fertile males and idiopathic non-obstructive azoospermic patients.Our control group was formed of 10 obstructive azoospermic vasectomized patients and our study group was formedof 23 non-obstructive azoospermic patients and 5 criptozoospermic patients. We performed inmunocitogenetic assay and fluorescence in situ hybridization (FISH) technique to analyse meiotic progression, sinaptonemal complex total length, recombination frequency in primary spermatocytes at pachytene stage and the incidence of aneuploidy and diploidy in testicular sperm. Non-obstructive azoospermic patients and criptozoospermic patients showed a significantly higher percentage of cells at the earlier stages of meiotic progression compared to the control group, before to complete the synapsis of homologues chromosomes. Suggesting partial meiotic arrest before reaching pachytene stage. Non-obstructive azoospermic patients group showed a significant decrease of recombination levels compared to the control group. However,the criptozoospermic group showed a decrease in recombination levels that was not significant compared to the control group. We observed a variation in the recombination frequency among individuals that was not correlated with the age of the patients and in the case of the control group was not related to the time from vasectomy. The decrease in recombination levels was observedfor the majority of the chromosomes analyzed. Independently of the chromosome size or structure. Regarding the synaptomal complex lenght, no differences were observed among groups, suggesting that there is no correlation between recombination frequency and synaptonemal complex length. FISH analysis in testicular spermatozoa in the non-obstructive azoospermic group showed an increase of aneuploidy and diploidy rate for the chromosomes analyzed compared to the control group. The criptozoospermic group only showed significant increase in the incidence of disomies for chromosome 18, sex chromosomes and diploidy. A significant increase in the incidence of sperm chromosomal abnormalities was detected in the 81.2% of non-obstructive azoospermic patients with significantly decrease of recombination levels compared to the control group

Relación entre el apego y la sintomatología depresiva en personas con Trastorno mental Grave y Duradero: El papel de la evitación experiencial

Severe and Chronic Mental Disorder (SMD) presents a high rate of comorbidity with depressive symptoms, causing a great personal and economic impact and reducingthe quality of life of people who suffer it. In recent years, insecure attachment styles and the avoidance of internal negative experiences seem to be variables related, not only, to the presence of severe psychopathology but it could, also, be involved in the development and maintenance of these comorbidity rates. The aim of this study was to explore the relationships between insecure attachment styles, experiential avoidance and depressive symptomatology in 41 individuals diagnosed with SMD. The results showed that insecure attachment styles were associated with an increase in depression. This relationship was intermediate by experiential avoidance. In sum, the widespread presence of depression in SMD highlights the urgent need to develop new treatments focused on working with attachment styles and the development of strategies based on acceptanceEl Trastorno Mental Grave y Duradero (TMGD) presenta una alta tasa de comorbilidad con síntomas depresivos, causando un gran impacto personal, económico y reduciendo la calidad de vida de las personas que lo sufren. En los últimos años, los estilos de apego inseguro y la evitación de experiencias negativas internas parecen variables relacionadas no sólo con la presencia de psicopatología grave, sino que podrían estar implicados en el desarrollo y mantenimiento de estas altas tasas de comorbilidad. El objetivo deeste estudio fue explorar las relaciones entre los estilos de apego inseguros, la evitación experiencial y la sintomatología depresiva en 41 individuos con diagnóstico de TMGD. Los resultados mostraron que los estilos de apego inseguros se asociaron con mayor depresión. Esta relación fue mediada por la evitación experiencial. En definitiva, la presencia generalizada de depresión en los TMGD subraya la necesidad urgente de desarrollar nuevos tratamientos que incluyan el trabajo con los estilos de apego y el desarrollo de estrategias basadas en la aceptació