9 research outputs found
Identificazioni di nuove variabili predittive di qualità ovocitaria ed embrionaria nei programmi di procreazione medicalmente assistita
Attualmente, nel protocollo standard di fecondazione in vitro, gli embrioni più idonei al trasferimento in utero vengono scelti in base ad una valutazione morfologica, due o tre giorni dopo la fecondazione. D’altro canto alcuni lavori suggeriscono che anche l’analisi morfologica degli zigoti può essere indicativa di successo nella fertilizzazione in vitro. Su tali evidenze, note da diversi anni in letteratura, si basano i primi passi del lavoro di ricerca svolto nel corso di dottorato, avendo come obiettivo quello di elaborare un sistema di scoring combinato (weighted score) che tenesse conto dello score degli embrioni pre-impianto, dello score degli zigoti e del numero di blastomeri degli embrioni pre-transfer. Il sistema, una volta elaborato, è stato testato mediante un analisi retrospettiva, al fine di valutare il suo valore predittivo in termini di percentuale di gravidanza e di tasso d’impianto. I dati ottenuti suggeriscono che il presente sistema di score combinato può essere utilizzato per stabilire una strategia nei cicli di fecondazione assistita per ridurre il numero di embrioni selezionati per il transfer, tutto ciò al fine di mantenere alte le percentuali di gravidanza riducendo l’incidenza di gravidanze multiple. In una seconda fase del dottorato di ricerca si è cercato di andare a determinare l’esistenza di una eventuale correlazione tra livelli intra-follicolari di leptina, i tassi di fertilizzazione ovocitaria ed il precedentemente testato sistema scoring embrionario. I risultati ottenuti evidenziano una correlazione positiva tra i livelli intrafollicolari di leptina e qualità ovocitaria intesa come “attitudine” alla fertilizzazione. Risulta inoltre evidente come la concentrazione intrafollicolare di leptina non sia correlata in maniera significativa con lo sviluppo e la qualità degli embrioni nelle tecniche di fecondazione assistita. Se i risultati riportati dovessero trovare conferma, la valutazione della concentrazione intrafollicolare di leptina potrebbe rappresentare un utile strategia nella selezione degli ovociti da destinare alle tecniche di fecondazione assistita
Effect of the male factor on the clinical outcome of intracytoplasmic sperm injection combined with preimplantation aneuploidy testing: Observational longitudinal cohort study of 1,219 consecutive cycles
The effect of ease of transfer and type of catheter used on pregnancy and implantation rates in an IVF program
Purpose: To test the effects of type of embryo transfer catheter, transfer difficulty, and observations after the transfer procedure on pregnancy and implantation rates in an IVF programme. Methods: Patients were prepared for IVF using standard protocols. Embryo transfer was performed using either Edwards-Wallace or TDT catheter. The difficulty of transfer was graded by a clinician and biologist. Blood observed inside the catheter after the transfer procedure was scored as endometrial damage. Pregnancy and implantation rates were scored. Results: Type of embryo transfer catheter and the observation of blood did not significantly affect pregnancy and implantation rates when transfer was performed by a single operator. Conclusions: In the hands of experienced, skilled operators, neither choice of transfer catheter and difficulty of transfer nor observations of blood on the transfer catheter caused any significant reduction in outcome to the patient
Oocyte competence is comparable between progestin primed ovarian stimulation with Norethisterone acetate (NETA-PPOS) and GnRH-antagonist protocols: A matched case-control study in PGT-A cycles
Objective: To outline oocyte competence after progestin primed ovarian stimulation with Norethisterone acetate (NETA-PPOS) compared to conventional GnRH-antagonist protocol. Study design: Retrospective matched case-control study involving advanced-maternal-age women undergoing ICSI with PGT-A. 89 NETA-PPOS were matched with 178 control patients based on maternal age and ovarian reserve biomarkers. Both groups underwent recombinant-FSH OS with GnRH-agonist ovulation trigger and collected ≥1 MII. In the study group, NETA (10 mg/day) was administered orally starting from day2 of the menstrual cycle. Euploid blastocyst rate per cohort of metaphase-II oocytes (EBR per MII) was the primary outcome. All other embryological and clinical outcomes were reported. Gestational age, birthweight and length were also assessed. Results: The EBR per MII was comparable among PPOS and control (13.9 % ± 19.3 % versus 13.3 % ± 17.9 %; the sample size allowed to exclude up to a 10 % difference). Blastocysts morphology and developmental rate were similar. No difference was reported for all clinical outcomes among the 61 and 107 vitrified-warmed euploid single blastocyst transfers respectively conducted. The cumulative live birth delivery rate per concluded cycles was also comparable (24.7 % versus 21.9 %). Neonatal outcomes were analogous. Conclusions: Oocyte competence after NETA-PPOS and standard OS is comparable. This evidence is reassuring and, because of its lower cost and possibly higher patients' compliance, supports PPOS administration whenever the patients are indicated to freeze-all (e.g., fertility preservation, PGT-A, oocyte donation). More data are required about follicle recruitment, oocyte yield, gestational and perinatal outcomes. Randomized-controlled-trials are advisable to confirm our evidence
Definition and validation of a custom protocol to detect miRNAs in the spent media after blastocyst culture: searching for biomarkers of implantation
STUDY QUESTION: Can miRNAs be reliably detected in the spent blastocyst media (SBM) after IVF as putative biomarkers of the
implantation potential of euploid embryos?
SUMMARY ANSWER: Adjustment of the data for blastocyst quality and the day of full-expansion hinders the predictive power of a fast,
inexpensive, reproducible and user-friendly protocol based on the detection of 10 selected miRNAs from SBM.
WHAT IS KNOWN ALREADY: Euploidy represents so far the strongest predictor of blastocyst competence. Nevertheless, ∼50% of
the euploid blastocysts fail to implant. Several studies across the years have suggested that a dialogue exists between the embryo and the
endometrium aimed at the establishment of a pregnancy. MicroRNAs have been proposed as mediators of such a dialogue and investigated
in this respect. Several expensive, time-consuming and complex protocols have been adopted and promising results have been produced, but
conclusive evidence from large clinical studies is missing.
STUDY DESIGN, SIZE, DURATION: This study was conducted in two phases from September 2015 to December 2017. In Phase 1,
the human blastocyst miRNome profile was defined from the inner cell mass (ICM) and the corresponding whole-trophectoderm (TE) of six
donated blastocysts. Two different protocols were adopted to this end. In parallel, 6 pools of 10 SBM each were run (3 from only implanted
euploid blastocysts, IEBs; and 3 from only not-implanted euploid blastocysts, not-IEBs). A fast, inexpensive and user-friendly custom protocol
for miRNA SBM profiling was designed. In Phase 2, 239 SBM from IEB and not-IEB were collected at three IVF centres. After 18 SBM from
poor-quality blastocysts were excluded from the analysis, data from 107 SBM from not-IEB and 114 from IEB were produced through the
previously developed custom protocol and compared. The data were corrected through logistic regressions.
PARTICIPANT/MATERIALS, SETTINGS, METHODS: Donated blastocysts underwent ICM and whole-TE isolation. SBM were
collected during IVF cycles characterized by ICSI, blastocyst culture in a continuous media, TE biopsy without zona pellucida opening in
Day 3, quantitative PCR (qPCR)-based aneuploidy testing and vitrified-warmed single euploid embryo transfer. Not-IEB and IEB were clustered
following a negative pregnancy test and a live birth, respectively. The Taqman Low Density Array (TLDA) cards and the Exiqon microRNA
human panel I+II qPCR analysis protocols were adopted to analyse the ICM and whole-TE. The latter was used also for SBM pools. A custom
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Customized miRNA analysis in spent IVF media 1747
protocol and plate was then designed based on the Exiqon workflow, validated and finally adopted for SBM analysis in study Phase 2. This
custom protocol allows the analysis of 10 miRNAs from 10 SBM in 3 hours from sample collection to data inspection.
MAIN RESULTS AND ROLE OF THE CHANCE: The TLDA cards protocol involved a higher rate of false positive results (5.6% versus
2.8% with Exiqon). There were 44 miRNAs detected in the ICM and TE from both the protocols. One and 24 miRNAs were instead detected
solely in the ICM and the TE, respectively. Overall, 29 miRNAs were detected in the pooled SBM: 8 only from not-IEB, 8 only from IEB and 13
from both. Most of them (N = 24/29, 82.7%) were also detected previously in both the ICM and TE with the Exiqon protocol; two miRNAs
(N = 2/29, 6.9%) were previously detected only in the TE, and three (N = 3/29, 10.3%) were never detected previously. In study Phase 2,
significant differences were shown between not-IEB and IEB in terms of both miRNA detection and relative quantitation. However, when the
data were corrected for embryo morphology and day of full development (i.e. SBM collection), no significant association was confirmed.
LIMITATIONS, REASONS FOR CAUTION: This study did not evaluate specifically exosomal miRNAs, thereby reducing the chance of
identifying the functional miRNAs. Ex-vivo experiments are required to confirm the role of miRNAs in mediating the dialogue with endometrial
cells, and higher throughput technologies need to be further evaluated for miRNA profiling from clinical SBM samples.
WIDER IMPLICATIONS OF THE FINDINGS: Although no clinical predictive power was reported in this study, the absence of invasiveness
related with SBM analysis and the evidence that embryonic genetic material can be reliably detected and analysed from SBM make this waste
product of IVF an important source for further investigations aimed at improving embryo selection.
STUDY FUNDING/COMPETING INTEREST(S): This project has been financially supported by Merck KgaA (Darmstadt, Germany)
with a Grant for Fertility Innovation (GFI) 2015. The authors have no conflict of interest to declare related with this stud
Effetto della somministrazione di aspirina e prednisone a basse dosi dopo il prelievo ovocitario sui tassi di impianto e di aborto in donne non affette da patologie autoimmunitarie sottoposte a ICSI: Studio prospettico randomizzato
Endometriosis shows no impact on the euploid blastocyst rate per cohort of inseminated metaphase-II oocytes: A case-control study
Objective: To evaluate the true impact of endometriosis on oocytes' competence defined as blastulation, euploidy and implantation rates. Design: Retrospective multicenter case-control study involving infertile couples undergoing ICSI with qPCR and trophectoderm biopsy-based PGT-A. Patients affected from endometriosis (n = 210) were diagnosed through transvaginal sonography or surgical history with histological confirmation. Each case was matched to two controls (n = 420) according to IVF clinic, maternal age at retrieval (38.6 ± 2.7 yr), number of previous failed IVF treatments (0.5 ± 0.8) and number of metaphase-II oocytes retrieved (6.1 ± 3.7 per patient). The primary outcome was the mean euploid blastocyst rate per cohort of inseminated metaphase-II oocytes. Other embryological, clinical, obstetric and neonatal outcomes were also evaluated. Results: The mean euploid blastocyst rate per cohort of inseminated metaphase-II oocytes was identical in the two groups (18 %±22 %) independently of maternal age. No difference was shown for all embryological outcomes investigated. The live birth rates per vitrified-warmed single euploid blastocyst transfer were also similar (67/158, 42 % in patients affected from endometriosis versus 132/327, 40 % in matched-controls). No difference was reported in the gestational and neonatal outcomes. The cumulative live birth delivery rates among completed cycles were also identical (61/201, 30 % versus 117/391, 30 % in endometriosis and matched-control groups, respectively) independently of maternal age. Conclusions: Endometriosis might not impair oocyte developmental and reproductive competence, although its potential impact on the number of metaphase-II oocytes retrieved cannot be ignored. This information is critical for clinicians during counseling to outline an effective strategy to treat infertile patients affected from this condition. Future prospective studies are needed to evaluate the impact of endometriosis stage on euploidy rates
Progesterone for preparation of the endometrium for frozen–thawed blastocyst transfer in vitro fertilization cycles: a prospective study on patients’ opinions on a new subcutaneous formulation
Inconclusive chromosomal assessment after blastocyst biopsy: prevalence, causative factors and outcomes after re-biopsy and re-vitrification. A multicenter experience
Study question: Can a second round of biopsy, vitrification and chromosomal testing provide a valid diagnosis where the first attempt fails? Summary answer: The risk of inconclusive chromosomal-assessment after trophectoderm biopsy was 2.5% but a further biopsy and vitrification-warming appeared not to impair the competence of euploid blastocysts. What is known already: The increasing implementation of multicell trophectoderm biopsy has significantly reduced the risk of inconclusive diagnosis after preimplantation-genetic-testing (PGT). Yet, few reports have defined the variables that influence the risk of failure or described the technical and clinical outcomes after re-biopsy. Study design, size, duration: Retrospective multicenter study involving 8990 blastocyst biopsies conducted between April 2013 and September 2017 at six IVF centers but analyzed at a single genetic laboratory. A total of 206 blastocysts were successfully re-biopsied after warming and re-expansion, then re-vitrified. And 49 of these blastocysts were diagnosed euploid and used in single-embryo-transfers (SETs). Logistic regression analyses were conducted. Participants/materials, setting, methods: A total of 3244 PGT-for-aneuploidies (PGT-A) cycles with a freeze-all approach, vitrification and qPCR-based analysis were performed by 2687 consenting couples. DNA amplification failure (AF) or non-concurrent data resulted in inconclusive diagnoses. In case of DNA amplification, the cellularity of the biopsy was estimated according to a previously validated method. Euploid SETs were performed. Clinical pregnancy, miscarriage, live birth rates (LBR) and perinatal outcomes were monitored. Main results and the role of chance: Overall, 2.5% of trophectoderm biopsies resulted in an inconclusive diagnosis (N = 228/8990). Specifically, 2% (N = 176/8990) resulted in AF and 0.5% (N = 52/8990) in non-concurrent results. The only parameters significantly associated with inconclusive diagnoses were the IVF center and the embryo age (days) at biopsy. Among samples with successful amplification, the number of cells in the biopsy and the day of biopsy were critical to limit non-concurrent results. In total, 213 blastocysts with an inconclusive diagnosis were warmed for re-analysis and the survival rate was 96.7% (N = 206/213). The euploidy rate in blastocysts biopsied twice was 51.9% (N = 107/206) and the euploid embryos were re-vitrified. Overall, 49 euploid embryos were warmed for replacement and all survived. The LBR after SET was 38.8% (N = 19/49). No minor/major obstetrical/perinatal complication was reported. Limitations, reasons for caution: A single aneuploidy-testing method was adopted in this retrospective analysis. A more powered report of the clinical and obstetrical/perinatal outcomes after re-biopsied and re-vitrified blastocysts euploid SET requires a larger sample size. Wider implications of the findings: It is important to re-biopsy and re-vitrify undiagnosed blastocysts since healthy live births can result from them. Study funding/competing interest(s): None. Trial registration number: None