Interchromosomal effect in carriers of translocations and inversions assessed by preimplantation genetic testing for structural rearrangements (PGT-SR).

PURPOSE: Balanced carriers of structural rearrangements have an increased risk of unbalanced embryos mainly due to the production of unbalanced gametes during meiosis. Aneuploidy for other chromosomes not involved in the rearrangements has also been described. The purpose of this work is to know if the incidence of unbalanced embryos, interchromosomal effect (ICE) and clinical outcomes differ in carriers of different structural rearrangements. METHODS: Cohort retrospective study including 359 preimplantation genetic testing cycles for structural rearrangements from 304 couples was performed. Comparative genomic hybridisation arrays were used for chromosomal analysis. The results were stratified and compared according to female age and carrier sex. The impact of different cytogenetic features of chromosomal rearrangements was evaluated. RESULTS: In carriers of translocations, we observed a higher percentage of abnormal embryos from day 3 biopsies compared with day 5/6 biopsies and for reciprocal translocations compared with other rearrangements. We observed a high percentage of embryos with aneuploidies for chromosomes not involved in the rearrangement that could be attributed to total ICE (aneuploid balanced and unbalanced embryos). No significant differences were observed in these percentages between types of rearrangements. Pure ICE (aneuploid balanced embyos) was independent of female age only for Robertsonian translocations, and significantly increased in day 3 biopsies for all types of abnormalities. Furthermore, total ICE for carriers of Robertsonian translocations and biopsy on day 3 was independent of female age too. High ongoing pregnancy rates were observed for all studied groups, with higher pregnancy rate for male carriers. CONCLUSION: We observed a higher percentage of abnormal embryos for reciprocal translocations. No significant differences for total ICE was found among the different types of rearrangements, with higher pure ICE only for Robertsonian translocations. There was a sex effect for clinical outcome for carriers of translocations, with higher pregnancy rate for male carriers. The higher incidence of unbalanced and aneuploid embryos should be considered for reproductive counselling in carriers of structural rearrangements.

High sperm DNA fragmentation delays human embryo kinetics when oocytes from young and healthy donors are microinjected.

BACKGROUND: Time-lapse monitoring (TLM) technology has been implemented in the clinical setting for the culture and selection of human embryos. Many studies have assessed the association between sperm DNA fragmentation (sDNAf) and clinical outcomes after ART, but little is known about the influence of sDNA on embryo morphokinetics. OBJECTIVES: The objective of this retrospective study, which includes 971 embryos from 135 consecutive ICSI cycles (56 cases with own oocytes, 79 with oocytes from young and healthy donors), was to assess if sDNAf has an impact on embryo morphokinetics. MATERIALS AND METHODS: Samples used to perform ICSI were analyzed by the flow cytometry TUNEL assay, and embryo development was assessed through an EmbyoScope® system. The association between sDNAf and the timings of cell cleavage was analyzed by categorizing the first variable into quartiles: ≤6.50%; 6.51-10.70%; 10.71-20.15%; >20.15%. RESULTS: In cases where sDNAf was above 20.15% (the upper quartile), embryos derived from donated oocytes (n = 644) showed significantly slower divisions. Such association was not observed in embryos obtained from the patients' own oocytes (n = 327). The embryo cleavage pattern (either normal, direct from 1 to 3 blastomeres, direct from 1 to 4 blastomeres, incomplete, reversed or asynchronous) was independent of the sDNAf level. Blastocyst arrival rate was 63.0% and the rate of good quality embryos (transferred and frozen embryos divided by the number of zygotes) was 45.49%. Neither parameter was related to the levels of sDNAf. DISCUSSION: According to our results, the association between high sDNAf and donated oocytes led to delayed cell division. To our knowledge, this is the first study suggesting that sDNAf can delay human embryo cleavage timings when oocytes from donors are inseminated. CONCLUSIONS: This finding may indicate that, in the presence of increased DNA damage, time is needed before the first embryonic cell division for the activation of the optimal DNA repairing machinery in higher quality oocytes.

Spermatozoa with numerical chromosomal abnormalities are more prone to be retained by Annexin V-MACS columns.

Colloidal super-paramagnetic microbeads conjugated with annexin V are effective for separating apoptotic spermatozoa by MACS as a result of the high affinity of annexin V for externalized PS molecules. The effectiveness of the procedure in reducing the percentage of sperm with fragmented DNA and abnormal morphology has also been reported. However, it is still unknown if it could decrease the percentage of aneuploid spermatozoa. The objective of our prospective study, performed on 16 males with abnormal FISH on spermatozoa, was to assess if MACS columns were useful tools to retain spermatozoa carrying chromosomal abnormalities in semen samples processed after density gradient centrifugation (DGC). The pellet obtained after DGC was subjected to MACS, and sperm FISH analyses were performed both in the eluded fraction and in the fraction retained in the column. The observed frequencies of disomy and nullisomy 13, 18, and 21, X and Y, as well as the diploidy rates in the MACS eluded fraction and the fraction retained in the MACS column were recorded. We observed that the frequencies of aneuploidies in the eluded fraction were lower than in the fraction retained in the MACS column (0.59% vs. 0.75%; p = 0.010). DGC determined a significant reduction in sperm concentration (z-ratio = 2.83; p = 0.005) and a significant increase in sperm progressive motility (z-ratio = -3.5; p < 0.001). MACS also led to a significant reduction in sperm concentration (z-ratio = 3.14; p = 0.002) and a significant increase in progressive motility (z-ratio = -2.59; p = 0.01) when compared with the post-DGC sample. Sperm concentration was similar in the two fractions generated by MACS (z-ratio = 0.63; p = 0.52), while progressive motility was significantly higher in the MACS eluded fraction (z-ratio = 2.42; p = 0.02). According to our results, MACS columns are able to selectively retain spermatozoa carrying chromosomal abnormalities. Furthermore, the performance of DGC and MACS on semen samples leads to an enrichment of progressive motility.

The use of morphokinetics as a predictor of  implantation: a multicentric study to define and validate an algorithm for embryo selection.

STUDY QUESTION: Can we use morphokinetic markers to select the embryos most likely to implant and are the results likely to be consistent across different clinics? SUMMARY ANSWER: Yes, morphokinetic markers can be used to select the embryos most likely to implant and the results were similar in different IVF clinics that share methods and organization to some extent. WHAT IS KNOWN ALREADY: With the introduction of time-lapse technology several authors have proposed the use of kinetic markers to improve embryo selection. The majority of these markers can be detected as early as Day 2 of development. Morphology remains the gold standard but kinetic markers have been proven as excellent tools to complement our decisions. Nevertheless, the majority of time-lapse studies are based on small data sets deriving from one single clinic. STUDY DESIGN, SIZE, DURATION: Retrospective multicentric study of 1664 cycles of which 799 were used to develop an algorithm (Phase 1 of the study) and 865 to test its predictive power (Phase 2 of the study). PARTICIPANTS/MATERIALS, SETTING, METHODS: University-affiliated infertility centres patients undergoing first or second ICSI cycle using their own or donated oocytes. Embryo development was analysed with a time-lapse imaging system. Variables studied included the timing to two cells (t2), three cells (t3), four cells (t4) and five cells (t5) as well as the length of the second cell cycle (cc2 = t3 - t2) and the synchrony in the division from two to four cells (s2 = t4 - t3). Implantation (IR) and clinical pregnancy (CPR) rates were also analysed. MAIN RESULTS AND THE ROLE OF CHANCE: During Phase 1 of the study we identified three variables most closely related to implantation: t3 (34-40 h), followed by cc2 (9-12 h) and t5 (45-55 h). Based on these results we elaborated an algorithm that classified embryos from A to D according to implantation potential. During Phase 2 of the study the algorithm was validated in a different group of patients that included 865 cycles and 1620 embryos transferred. In this phase of the study, embryos were categorized based on the algorithm and significant differences in IR were observed between the different categories ('A' 32%, 'B' 28%, 'C' 26%, 'D' 20% and 'E' 17%, P < 0.001). In addition we identified three quality criteria: direct cleavage from one to three cells, uneven blastomere size in second cell cycle and multinucleation in third cell cycle. LIMITATIONS, REASONS FOR CAUTION: The retrospective nature of the study limits its potential value, although the use of one database to generate the algorithm (embryos from this database were not selected by any morphokinetic criteria) and one database to validate it reinforces our conclusions. WIDER IMPLICATIONS OF THE FINDINGS: The elaboration of an algorithm based on a larger database derived from different (albeit related) clinics raises the possibility that such algorithms could be applied in different clinical settings.

The role of the adapter LAT in T cell activation and maturation

El receptor de células T (TCR) reconoce péptidos unidos alcomplejo mayor de histocompatibilidad (MHC) y transmite estainformación a la célula T a través de proteínas adaptadoras. Eladaptador LAT (de «Linker for Activation of T cells») es una proteínatransmembrana que, una vez fosforilada en sus residuos detirosina, coordina la unión de muchas proteínas implicadas enseñalización intracelular, de modo que promueve la formaciónde complejos multi-moleculares que regulan la activación y maduraciónde las células T. Estudios funcionales y estructurales, tantoin vitro como in vivo, han revelado un papel central de LAT comoplataforma de distribución de señales procedentes del TCR y pre-TCR, así como una inesperada función en la regulación del desarrolloy homeostasis de las células T. En esta revisión se discutenalgunos de los más recientes avances acerca de las funciones deeste adaptador en la maduración y activación de los linfocitos T

The T Cell Receptor (TCR) recognizes peptides bound to majorhistocompatibility complex (MHC) molecules and relays this informationto the T cell through adapter proteins. The adapter LAT(Linker for Activation of T cells) is a transmembrane protein that,once phosphorylated in its tyrosine residues, coordinates the bindingof many signaling proteins in order to assemble multi-molecularcomplexes that regulate T cell activation and maturation.Structure/function studies, both in vitro and in vivo, have revealeda central role of LAT as a platform for the distribution of signalscoming from the TCR and the pre-TCR, and also an unexpectedfunction in the regulation of T cell development and homeostasis.Thus, in the present review we discuss some of the recentadvances on the role of this adaptor in T lymphocyte developmentand activation

The role of the adapter LAT in T cell activation and maturation

El receptor de células T (TCR) reconoce péptidos unidos al complejo mayor de histocompatibilidad (MHC) y transmite esta información a la célula T a través de proteínas adaptadoras. Eladaptador LAT (de «Linker for Activation of T cells») es una proteína transmembrana que, una vez fosforilada en sus residuos detirosina, coordina la unión de muchas proteínas implicadas en señalización intracelular, de modo que promueve la formación de complejos multi-moleculares que regulan la activación y maduraciónde las células T. Estudios funcionales y estructurales, tanto in vitro como in vivo, han revelado un papel central de LAT como plataforma de distribución de señales procedentes del TCR y pre-TCR, así como una inesperada función en la regulación del desarrollo y homeostasis de las células T. En esta revisión se discuten algunos de los más recientes avances acerca de las funciones de este adaptador en la maduración y activación de los linfocitos T (AU)

The T Cell Receptor (TCR) recognizes peptides bound to majorhistocompatibility complex (MHC) molecules and relays this information to the T cell through adapter proteins. The adapter LAT(Linker for Activation of T cells) is a transmembrane protein that,once phosphorylated in its tyrosine residues, coordinates the binding of many signaling proteins in order to assemble multi-molecular complexes that regulate T cell activation and maturation.Structure/function studies, both in vitro and in vivo, have revealeda central role of LAT as a platform for the distribution of signalscoming from the TCR and the pre-TCR, and also an unexpectedfunction in the regulation of T cell development and homeostasis.Thus, in the present review we discuss some of the recent advances on the role of this adaptor in T lymphocyte developmentand activation (AU)