Euploid blastocysts implant irrespective of their morphology after NGS-(PGT-A) testing in advanced maternal age patients.

PURPOSE: Does blastocyst morphology following euploid elective single embryo transfer (eSET) after preimplantation genetic testing for aneuploidies (PGT-A) via next generation sequencing impact clinical outcome? METHODS: Two hundred ninety-six patients underwent PGT-A. Of 1549 blastocysts, 1410 blastocysts had a conclusive result after PGT-A and were included for analysis. An eSET policy was followed in a frozen embryo replacement cycle. A total of 179 euploid blastocysts were thawed and transferred. Clinical outcomes were categorized in four different embryo quality groups: excellent, good, average and poor. RESULTS: Euploidy rate was 19/36 (52.7%, 95% CI 37-68), 199/470 (42.3%, 95% CI 38-47), 156/676 (23.0%, 95% CI 20-26) and 39/228 (17.1%, 95% CI 13-23) in the excellent, good, average and poor quality blastocyst groups, respectively. Fitted logistic regression analysis taking into account the following covariables: female, age, embryo chromosomal status and day of blastocyst development/biopsy showed that morphology was predictive of the comprehensive chromosome screening result (p < 0.05). A logistic regression analysis was also performed on clinical outcomes taking into account the effect of blastocyst morphology and day of blastocyst development/biopsy. None of the parameters were shown to be significant, suggesting morphology and day of blastocyst development/biopsy do not reduce the competence of euploid embryos (p > 0.05). CONCLUSIONS: After eSET, implantation rate was 80-86%; live birth rate per embryo transfer was 60-73% and clinical miscarriage rate was found to be < 10% and were not significantly affected by the embryo morphology. Results are concordant with those reported when using aCGH and highlights the competence of poor-quality euploid embryos.

Investigation of microRNA expression and DNA repair gene transcripts in human oocytes and blastocysts.

PURPOSE: The aim of the study is to investigate the regulation of DNA repair genes by microRNAs (miRNAs). miRNAs are short non-coding RNAs that regulate transcriptional and post-transcriptional gene silencing. Several miRNAs that are expressed during preimplantation embryo development have been shown or are predicted to target genes that regulate cell cycle checkpoints and DNA repair in response to DNA damage. METHODS: This study compares the expression level of 20 miRNAs and 9 target transcripts involved in DNA repair. The statistical significance of differential miRNA expression between oocytes and blastocysts was determined by t test analysis using the GraphPad Prism v6 software. The possible regulatory roles of miRNAs on their target messenger RNAs (mRNAs) were analysed using a Pearson correlation test. RESULTS: This study shows for the first time that several miRNAs are expressed in human oocytes and blastocysts that target key genes involved in DNA repair and cell cycle checkpoints. Blastocysts exhibited statistically significant lower expression levels for the majority of miRNAs compared to oocytes (p < 0.05). Correlation analyses showed that there was both inverse and direct association between miRNAs and their target mRNAs. CONCLUSIONS: miRNAs target many mRNAs including ones involved in DNA repair mechanisms. This study suggests that miRNAs and their target mRNAs involved in DNA repair are expressed in preimplantation embryos. Similar to the miRNAs expressed in adult tissues, these miRNAs seem to have regulatory roles on their target DNA repair mRNAs during preimplantation embryo development.

Accurate and rapid prenatal diagnosis of the most frequent East Mediterranean beta-thalassemia mutations.

Beta-thalassemia is the most common genetic disorder in the Lebanese population. Of the 200 different mutations in the beta-globin gene that leads to thalassemia, the IVSI-110 (29.87%), IVSI-6 (20.74%), IVSI-1 (14.07%), IVSII-1 (9.13%), Cd29 (9.13%), and Cd30 (3.95%) mutations are the most frequent among Lebanese thalassemic patients. These mutations are also present at high frequencies in the East Mediterranean region. Due to this high prevalence of certain beta-thalassemia mutations, a rapid technique for the prenatal diagnosis of these mutations was implemented. The technique used is based on Real-Time PCR quantification and melting curve analysis of the amplified fragment using the LightCycler. The DNA samples used for amplification were obtained from CVS or amniotic fluid. Six mutations were easily and efficiently detected using only 3 sets of probes. With this method, mutant genotypes can be easily distinguished from normal alleles. In prenatal diagnosis, the accuracy and the speed of testing are paramount. The method of prenatal beta-thalassemia mutations detection described here is efficient and fast, with the entire procedure including DNA preparation taking less than half a workday. It is safe, does not involve radioactivity, and is accurate showing 100% concordance with conventional DNA sequencing methods.