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1.
Fertil Steril ; 2024 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-39069216

RESUMO

OBJECTIVE: To evaluate the positive predictive value (PPV) of prenatal cell-free DNA (cfDNA) screening for chromosomal aneuploidies in pregnancies achieved either after single euploid transfer in in vitro fertilization or Preimplantation Genetic Testing for Aneuploidy (PGT-A) cycles or transfer of single untested embryo, and to assess the concordance of prenatal-cfDNA-screening and PGT-A results. DESIGN: Single center retrospective cohort study. SETTING: Fertility clinic. PATIENT(S): A total of 2,973 prenatal-cfDNA-screening results for the most common trisomies (T13, T18, T21, X, and Y) and microdeletions (1p36, 4p16.3, 5p15.2, 15q11.2, and 22q11.2) from singleton pregnancies allocated into two groups: PGT-A group (n = 1,204) pregnant after single euploid transfer and non-PGT-A group (n = 1769) pregnant after transfer of single untested embryo, between 2016 and 2023. INTERVENTION(S): Not applicable. MAIN OUTCOME MEASURE(S): The primary outcome measure was the accuracy of prenatal-cfDNA screening. Positive and negative prenatal-cfDNA-screening results and subsequent prenatal or postnatal diagnostic testing were used to classify each positive prenatal-cfDNA-screening result as a true or a false positive. Secondary endpoints were to evaluate the concordance of PGT-A and prenatal-cfDNA-screening results and to assess the differences in the fetal fraction of cfDNA used for prenatal-cfDNA-screening reports between the study groups. RESULT(S): Prenatal-cfDNA screening was performed at a mean 11.3 ± 1.8 weeks gestational age, and yielded results in 99.9% of the patients (0.1% cancellation rate). There was no difference in the fetal fraction between PGT-A tested and not tested pregnancies (9.5% ± 4% vs. 10.3% ± 4%). 13 positive prenatal-cfDNA-screening results (two T21, two X0, four XXX, one XYY, one indeterminate sex, two 22q11 del/dup, and one 15q11.2del) were received for PGT-A group. Only one (22q11 dup) was confirmed with amniocentesis and fetal autopsy, giving a PPV for an abnormal prenatal-cfDNA screening of 7.7%, the rest had results concordant with PGT-A. Sex chromosomes were 100% concordant between prenatal-cfDNA-screening and PGT-A results, giving a 100% PPV for PGT-A for sex chromosomes and 100% negative predictive value for aneuploidies. Positive prenatal-cfDNA-screening results were received for 27 pregnancies from untested embryos (1.5%), follow-up testing was electively performed for 21, and 8 had confirmed the prenatal-cfDNA-screening result, giving a PPV for the non-PGT-A group of 38%. CONCLUSION(S): This study demonstrates that patients undergoing in vitro fertilization/PGT-A and single euploid embryo transfer can reliably do prenatal-cfDNA screening during their first trimester. Fetal fraction in singleton pregnancies after PGT-A tested embryos is not different from pregnancies with untested embryos. Positive predictive value for an abnormal prenatal-cfDNA-screening result after euploid embryo transfer was reassuringly low (7.7%). PGT-A reliably selects against aneuploidy with 100% concordance with fetal sex.

2.
Genome Res ; 34(1): 70-84, 2024 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-38071472

RESUMO

Meiotic recombination is crucial for human genetic diversity and chromosome segregation accuracy. Understanding its variation across individuals and the processes by which it goes awry are long-standing goals in human genetics. Current approaches for inferring recombination landscapes rely either on population genetic patterns of linkage disequilibrium (LD)-capturing a time-averaged view-or on direct detection of crossovers in gametes or multigeneration pedigrees, which limits data set scale and availability. Here, we introduce an approach for inferring sex-specific recombination landscapes using data from preimplantation genetic testing for aneuploidy (PGT-A). This method relies on low-coverage (<0.05×) whole-genome sequencing of in vitro fertilized (IVF) embryo biopsies. To overcome the data sparsity, our method exploits its inherent relatedness structure, knowledge of haplotypes from external population reference panels, and the frequent occurrence of monosomies in embryos, whereby the remaining chromosome is phased by default. Extensive simulations show our method's high accuracy, even at coverages as low as 0.02×. Applying this method to PGT-A data from 18,967 embryos, we mapped 70,660 recombination events with ∼150 kbp resolution, replicating established sex-specific recombination patterns. We observed a reduced total length of the female genetic map in trisomies compared with disomies, as well as chromosome-specific alterations in crossover distributions. Based on haplotype configurations in pericentromeric regions, our data indicate chromosome-specific propensities for different mechanisms of meiotic error. Our results provide a comprehensive view of the role of aberrant meiotic recombination in the origins of human aneuploidies and offer a versatile tool for mapping crossovers in low-coverage sequencing data from multiple siblings.


Assuntos
Aneuploidia , Testes Genéticos , Masculino , Humanos , Feminino , Testes Genéticos/métodos , Aberrações Cromossômicas , Desequilíbrio de Ligação , Linhagem
3.
Fertil Steril ; 120(5): 957-966, 2023 11.
Artigo em Inglês | MEDLINE | ID: mdl-37532168

RESUMO

OBJECTIVE: To understand the clinical risks associated with the transfer of embryos classified as a mosaic using preimplantation genetic testing for aneuploidy. DESIGN: Analysis of data collected between 2017 and 2023. SETTING: Multicenter. PATIENTS: Patients of infertility treatment. INTERVENTION: Comparison of pregnancies resulting from embryos classified as euploid or mosaic using the 20%-80% interval in chromosomal intermediate copy numbers to define a mosaic result. MAIN OUTCOME MEASURES: Rates of spontaneous abortion, birth weight, length of gestation, incidence of birth defects, and chromosomal status during gestation. RESULTS: Implanted euploid embryos had a significantly lower risk of spontaneous abortion compared with mosaic embryos (8.9% [n = 8,672; 95% confidence interval {CI95} 8.3, 9.5] vs. 22.2% [n = 914; CI95 19.6, 25.0]). Embryos with mosaicism affecting whole chromosomes (not segmental) had the highest risk of spontaneous abortion (27.6% [n = 395; CI95 23.2, 32.3]). Infants born from euploid, mosaic, and whole chromosome mosaic embryos had average birth weights and lengths of gestation that were not statistically different (3,118 g and 267 days [n = 488; CI95 3,067, 3,169, and 266, 268], 3052 g and 265 days [n = 488; CI95 2,993, 3,112, and 264,267], 3,159 g and 268 days [n = 194; CI95 3,070, 3,249, and 266,270], respectively). Out of 488 infants from mosaic embryo transfers (ETs), one had overt gross abnormalities as defined by the Centers for Disease Control and Prevention. Most prenatal tests performed on pregnancies from mosaic ETs had normal results, and only three pregnancies produced prenatal test results reflecting the mosaicism detected at the embryonic stage (3 out of 250, 1.2%; CI95 0.25, 3.5). CONCLUSION: Although embryos classified as mosaic experience higher rates of miscarriage than euploid embryos (with a particularly high frequency shortly after implantation), infants born of mosaic ETs are similar to infants of euploid ETs. Prenatal testing indicates that mosaicism resolves during most pregnancies, although this process is not perfectly efficient. In a small percentage of cases, the mosaicism persists through gestation. These findings can serve as risk-benefit considerations for mosaic ETs in the fertility clinic.


Assuntos
Aborto Espontâneo , Diagnóstico Pré-Implantação , Gravidez , Feminino , Recém-Nascido , Humanos , Aborto Espontâneo/etiologia , Aborto Espontâneo/genética , Diagnóstico Pré-Implantação/métodos , Fertilização in vitro/efeitos adversos , Fertilização in vitro/métodos , Blastocisto , Testes Genéticos/métodos , Aneuploidia , Mosaicismo , Cromossomos
4.
bioRxiv ; 2023 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-37333422

RESUMO

Meiotic recombination is crucial for human genetic diversity and chromosome segregation accuracy. Understanding its variation across individuals and the processes by which it goes awry are long-standing goals in human genetics. Current approaches for inferring recombination landscapes either rely on population genetic patterns of linkage disequilibrium (LD)-capturing a time-averaged view-or direct detection of crossovers in gametes or multi-generation pedigrees, which limits dataset scale and availability. Here, we introduce an approach for inferring sex-specific recombination landscapes using data from preimplantation genetic testing for aneuploidy (PGT-A). This method relies on low-coverage (<0.05×) whole-genome sequencing of in vitro fertilized (IVF) embryo biopsies. To overcome the data sparsity, our method exploits its inherent relatedness structure, knowledge of haplotypes from external population reference panels, as well as the frequent occurrence of monosomies in embryos, whereby the remaining chromosome is phased by default. Extensive simulations demonstrate our method's high accuracy, even at coverages as low as 0.02×. Applying this method to PGT-A data from 18,967 embryos, we mapped 70,660 recombination events with ~150 kbp resolution, replicating established sex-specific recombination patterns. We observed a reduced total length of the female genetic map in trisomies compared to disomies, as well as chromosome-specific alterations in crossover distributions. Based on haplotype configurations in pericentromeric regions, our data indicate chromosome-specific propensities for different mechanisms of meiotic error. Our results provide a comprehensive view of the role of aberrant meiotic recombination in the origins of human aneuploidies and offer a versatile tool for mapping crossovers in low-coverage sequencing data from multiple siblings.

5.
J Ovarian Res ; 15(1): 124, 2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36457002

RESUMO

BACKGROUND: Intracytoplasmic sperm injection (ICSI) has become a common method of fertilization in assisted reproduction worldwide. However, there are still gaps in knowledge of the ideal IVF-ICSI workflow including the optimal duration of time between induction of final oocyte maturation, oocyte denudation and ICSI. The aim of this study was to examine outcomes following different workflow protocols in IVF-ICSI procedures in blastocysts that have undergone undisturbed incubation and preimplantation genetic testing for aneuploidy (PGT-A) prior to transfer. METHODS: Retrospective secondary analysis of 113 patients (179 IVF cycles, 713 embryos), all of whom have gone through IVF-ICSI and PGT-A using undisturbed culture. Predictive test variables were the length of time from: trigger to OPU, OPU to denudation, and denudation to ICSI. Outcome metrics assessed were: maturation, fertilization, blastulation and euploid rates. Generalized Estimated Equations Linear Model was used to examine the relationship between key elements of a given cycle and continuous outcomes and LOESS curves were used to determine the effect over time. RESULTS: In a paired multi-regression analysis, where each patient served as its own control, delaying OPU in patients with unexplained infertility improved both maturation and blastulation rates (b = 29.7, p < 0.0001 and b = 9.1, p = 0.06, respectively). Longer incubation with cumulus cells (CCs) significantly correlated with improved ploidy rates among patients under 37, as well as among patients with unexplained infertility (r = 0.22 and 0.29, respectively), which was also evident in a multiple regression analysis (b = 6.73, p < 0.05), and in a paired analysis (b = 6.0, p < 0.05). Conversely, among patients with a leading infertility diagnosis of male factor, longer incubation of the denuded oocyte prior to ICSI resulted in a significantly higher euploid rate (b = 15.658, p < 0.0001). CONCLUSIONS: In this study we have demonstrated that different IVF-ICSI workflows affect patients differently, depending on their primary infertility diagnosis. Thus, ideally, the IVF-ICSI workflow should be tailored to the individual patient based on the primary infertility diagnosis. This study contributes to our understanding surrounding the impact of IVF laboratory procedures and highlights the importance of not only tracking "classic" IVF outcomes (maturation, fertilization, blastulation rates), but highlights the importance that these procedures have on the ploidy of the embryo.


Assuntos
Infertilidade , Injeções de Esperma Intracitoplásmicas , Masculino , Feminino , Humanos , Fluxo de Trabalho , Estudos Retrospectivos , Sêmen , Aneuploidia , Ploidias , Testes Genéticos
6.
Fertil Steril ; 116(5): 1212-1219, 2021 11.
Artigo em Inglês | MEDLINE | ID: mdl-34627598

RESUMO

Chromosomal mosaicism, the coexistence of cells with different chromosomal content, has been documented in human embryos for 3 decades. Early versions of preimplantation genetic testing for aneuploidy (PGT-A) did not measure mosaicism, either because typically only a single cell was assessed or because the technique could not accurately identify it. Although this led to a straightforward diagnosis (an embryo was considered either normal or abnormal), it simply avoided the issue and, in hindsight, may have led to numerous misdiagnoses with negative clinical consequences. Modern PGT-A evaluates a multicellular biopsy specimen with techniques capable of recognizing intermediate copy number signals for chromosomes or subchromosomal regions. We are, therefore, inevitably confronted with the issue of mosaicism and the challenge of managing embryos producing such results in the clinic. Here we discuss recent data showing that not only mosaicism in general, but specific features of mosaicism detected with PGT-A, are associated with variable clinical outcomes. The conclusion is evident: mosaicism should be considered for more informed and improved embryo selection in the clinic.


Assuntos
Blastocisto/patologia , Testes Genéticos , Infertilidade/terapia , Mosaicismo , Diagnóstico Pré-Natal , Técnicas de Reprodução Assistida/efeitos adversos , Aneuploidia , Transferência Embrionária , Feminino , Aconselhamento Genético , Humanos , Infertilidade/diagnóstico , Infertilidade/fisiopatologia , Masculino , Valor Preditivo dos Testes , Gravidez , Reprodutibilidade dos Testes , Resultado do Tratamento
7.
Hum Reprod Update ; 27(3): 486-500, 2021 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-33355342

RESUMO

BACKGROUND: Delayed parenthood, by both women and men, has become more common in developed countries. The adverse effect of advanced maternal age on embryo aneuploidy and reproductive outcomes is well known. However, whether there is an association between paternal age (PA) and embryonic chromosomal aberrations remains controversial. Oocyte donation (OD) is often utilized to minimize maternal age effects on oocyte and embryo aneuploidy, thus providing an optimal model to assess the effect of PA. Several studies have revealed a higher than expected rate of aneuploidy in embryos derived from young oocyte donors, which warrants examination as to whether this may be attributed to advanced PA (APA). OBJECTIVE AND RATIONALE: The objective of this systematic review and individual patient data (IPD) meta-analysis is to evaluate existing evidence regarding an association between PA and chromosomal aberrations in an OD model. SEARCH METHODS: This review was conducted according to PRISMA guidelines for systematic reviews and meta-analyses. Medline, Embase and Cochrane databases were searched from inception through March 2020 using the (MeSH) terms: chromosome aberrations, preimplantation genetic screening and IVF. Original research articles, reporting on the types and/or frequency of chromosomal aberrations in embryos derived from donor oocytes, including data regarding PA, were included. Studies reporting results of IVF cycles using only autologous oocytes were excluded. Quality appraisal of included studies was conducted independently by two reviewers using a modified Newcastle-Ottawa Assessment Scale. A one-stage IPD meta-analysis was performed to evaluate whether an association exists between PA and aneuploidy. Meta-analysis was performed using a generalized linear mixed model to account for clustering of embryos within patients and clustering of patients within studies. OUTCOMES: The search identified 13 032 references, independently screened by 2 reviewers, yielding 6 studies encompassing a total of 2637 IVF-OD cycles (n = 20 024 embryos). Two 'low' quality studies using FISH to screen 12 chromosomes on Day 3 embryos (n = 649) reported higher total aneuploidy rates and specifically higher rates of trisomy 21, 18 and 13 in men ≥50 years. One 'moderate' and three 'high' quality studies, which used 24-chromosome screening, found no association between PA and aneuploidy in Day 5/6 embryos (n = 12 559). The IPD meta-analysis, which included three 'high' quality studies (n = 10 830 Day 5/6 embryos), found no significant effect of PA on the rate of aneuploidy (odds ratio (OR) 0.97 per decade of age, 95% CI 0.91-1.03), which was robust to sensitivity analyses. There was no association between PA and individual chromosome aneuploidy or segmental aberrations, including for chromosomes X and Y (OR 1.06 per decade of age, 95% CI 0.92-1.21). Monosomy was most frequent for chromosome 16 (217/10802, 2.01%, 95% CI 1.76-2.29%) and trisomy was also most frequent for chromosome 16 (194/10802, 1.80%, 95% CI 1.56-2.06%). WIDER IMPLICATIONS: We conclude, based on the available evidence, that APA is not associated with higher rates of aneuploidy in embryos derived from OD. These results will help fertility practitioners when providing preconception counselling, particularly to older men who desire to have a child.


Assuntos
Idade Paterna , Diagnóstico Pré-Implantação , Idoso , Aneuploidia , Feminino , Fertilização in vitro , Humanos , Masculino , Doação de Oócitos , Oócitos , Gravidez , Diagnóstico Pré-Implantação/métodos
8.
Fertil Steril ; 114(2): 293-300, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32654815

RESUMO

OBJECTIVE: To investigate a possible correlation between chromosomal aberrations and paternal age, analyzing embryos derived from young oocyte donors, with available preimplantation genetic testing for aneuploidy results from day 5/6 trophectoderm biopsy obtained by next-generation sequencing for all 24 chromosomes. DESIGN: Retrospective cohort study. SETTING: Canadian fertility centre. PATIENT(S): A total of 3,118 embryos from 407 male patients, allocated into three paternal age groups: group A, ≤39 years (n = 203); group B, 40-49 years (n = 161); group C, ≥50 years (n = 43). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): The primary outcomes were aneuploidy, euploidy, mosaicism, and blastocyst formation rates. Secondary endpoints were comparison of specific chromosome aneuploidy, segmental and complex (involving two chromosomes + mosaicism >50%) aneuploidy, and analysis of overall percentage of chromosomal gains and losses within each group. RESULT(S): The study included 437 in vitro fertilization (IVF) antagonist cycles using 302 oocyte donors in which preimplantation genetic testing for aneuploidy was performed. Overall, 70.04% of embryos were euploid, 13.9% were aneuploid, and 16.06% were mosaic. No significant differences among paternal age groups A, B, and C were found in euploidy rates (69.2%, 70.6%, 71.4%, respectively), aneuploidy rates (14.7%, 12.8%, 13.9%, respectively) or mosaicism rates (16.1%, 16.6%, 13.6%; respectively). The fertilization rate was lower in group C compared with group B (76.35% vs. 80.09%). No difference was found in blastocyst formation rate between the study groups (median 52% [interquartile range, 41%, 67%] vs. 53% [42%, 65%] vs. 52% [42%, 64%], respectively). A generalized linear mixed model regression analysis for embryo ploidy rates found older oocyte donor age to be independently associated with embryo aneuploidy (odds ratio = 1.041; 95% CI, 1.009-1.074). The rate of segmental aneuploidies was significantly higher in the older versus younger paternal age group (36.6% vs. 19.4%). CONCLUSION(S): No association was found between paternal age and aneuploidy rates in embryos derived from IVF cycles using young oocyte donors, after adjusting for donor, sperm, and IVF cycle characteristics. Advanced paternal age ≥ 50, compared with younger paternal ages, was associated with a lower fertilization rate and increased rate of segmental aberrations.


Assuntos
Aneuploidia , Blastocisto/patologia , Fertilização in vitro , Infertilidade/terapia , Doação de Oócitos , Idade Paterna , Adulto , Biópsia , Feminino , Fertilidade , Fertilização in vitro/efeitos adversos , Testes Genéticos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Infertilidade/diagnóstico , Infertilidade/fisiopatologia , Masculino , Pessoa de Meia-Idade , Mosaicismo , Doação de Oócitos/efeitos adversos , Diagnóstico Pré-Implantação , Estudos Retrospectivos , Medição de Risco , Fatores de Risco , Adulto Jovem
10.
Sci Rep ; 10(1): 7244, 2020 04 29.
Artigo em Inglês | MEDLINE | ID: mdl-32350403

RESUMO

Preimplantation genetic testing for aneuploidies (PGT-A) using trophectoderm (TE) biopsy samples is labour intensive, invasive, and subject to sampling bias. In this study, we report on the efficacy and factors affecting accuracy of a technique we pioneered for minimally invasive preimplantation genetic testing for aneuploidy (miPGT-A). Our technique uses cell-free embryonic DNA (cfeDNA) in spent embryo culture medium (SEM) combined with blastocoel fluid (BF) to increase the amount of assayable cfeDNA. We compared miPGT-A results (n = 145 embryos) with standard PGT-A analysis of the corresponding trophectoderm biopsy. We found that accuracy of miPGT was not related to blastocyst morphological grade. The overall concordance rate per sample for euploidy/aneuploidy status between miPGT-A and TE biopsy samples was 88/90 (97.8%), and was not different between good 47/48 (97.9%) and moderate/low quality blastocysts 41/42 (97.9%) (p > 0.05). Importantly, we also discovered that for cfeDNA analysis, the SurePlex whole genome amplification (WGA) kit can be utilized without an additional cell lysis/extraction DNA step; this efficiency likely reduces the risk of maternal contamination. Regarding origin of embryonic cfeDNA, the average amount of miPGT-A WGA-DNA we obtained from blastocysts with different morphological grades, as well as the size miPGT-A WGA-DNA fragments, suggest that it is unlikely that apoptosis and necrosis are only mechanisms of DNA release from the inner cell mass (ICM) and TE into BF and SEM.


Assuntos
Aneuploidia , Blastocisto/citologia , Ácidos Nucleicos Livres/análise , Embrião de Mamíferos/citologia , Meios de Cultura , Humanos , Masculino , Diagnóstico Pré-Implantação
11.
Sci Rep ; 9(1): 2853, 2019 02 27.
Artigo em Inglês | MEDLINE | ID: mdl-30814554

RESUMO

Improved embryo prioritization is crucial in optimizing the results in assisted reproduction, especially in light of increasing utilization of elective single embryo transfers. Embryo prioritization is currently based on morphological criteria and in some cases incorporates preimplantation genetic testing for aneuploidy (PGT-A). Recent technological advances have enabled parallel genomic and transcriptomic assessment of a single cell. Adding transcriptomic analysis to PGT-A holds promise for better understanding early embryonic development and implantation, and for enhancing available embryo prioritization tools. Our aim was to develop a platform for parallel genomic and transcriptomic sequencing of a single trophectoderm (TE) biopsy, that could later be correlated with clinical outcomes. Twenty-five embryos donated for research were utilized; eight for initial development and optimization of our method, and seventeen to demonstrate clinical safety and reproducibility of this method. Our method achieved 100% concordance for ploidy status with that achieved by the classic PGT-A. All sequencing data exceeded quality control metrics. Transcriptomic sequencing data was sufficient for performing differential expression (DE) analysis. All biopsies expressed specific TE markers, further validating the accuracy of our method. Using PCA, samples clustered in euploid and aneuploid aggregates, highlighting the importance of controlling for ploidy in every transcriptomic assessment.


Assuntos
Blastocisto/metabolismo , Desenvolvimento Embrionário , Fertilização in vitro , Sequenciamento de Nucleotídeos em Larga Escala , Adulto , Biópsia , Blastocisto/patologia , Feminino , Humanos
12.
PLoS One ; 13(5): e0197262, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29746572

RESUMO

OBJECTIVE: To assess whether embryonic DNA isolated from blastocyst culture conditioned medium (BCCM) combined with blastocoel fluid (BF) could be used for blastocyst stage non-invasive preimplantation genetic testing for chromosomal aneuploidy (non-invasive preimplantation genetic screening, NIPGS). PATIENTS: 47 embryos from 35 patients undergoing IVF. INTERVENTIONS: DNA analysis of combined BCCM plus BF in comparison with trophectoderm (TE) biopsy and/or whole blastocyst (WB)using next generation sequencing (NGS). RESULTS: Embryonic DNA was successfully amplified in 47/47 NIPGS samples (28 frozen-thawed and 19 fresh culture samples) ranging from 6.3 to 44.0 ng/µl. For frozen-thawed embryos, the concordance rate for whole chromosome copy number per sample was equivalent between NIPGS vs. TE biopsy, NIPGS vs. WB and TE vs. WB samples taken from the same embryo was 87.5%; 96.4% and 91.7% respectively (P>0.05), and the rate of concordance per single chromosome was 99.3%, 99.7% and 99.7%, respectively (P>0.05). In fresh cases (Day 4 to Day 5/6 culture), the concordance rate for whole chromosome copy number per sample between NIPGS vs. TE samples taken from the same embryo was 100%, and the rate of concordance per single chromosome was 98.2% (P>0.05). CONCLUSIONS: A combination of BCCM and BF contains sufficient embryonic DNA for whole genome amplification and accurate aneuploidy screening. Our findings suggest that aneuploidy screening using BCCM combined with BF could potentially serve as a novel NIPGS approach for use in human IVF.


Assuntos
Blastocisto , Meios de Cultivo Condicionados , Testes Genéticos , Diagnóstico Pré-Implantação , Aneuploidia , Blastocisto/metabolismo , DNA/metabolismo , Técnicas de Cultura Embrionária , Fertilização in vitro , Humanos , Estudo de Prova de Conceito , Análise de Sequência de DNA
13.
PLoS One ; 11(1): e0146264, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26745889

RESUMO

BACKGROUND: Although age-related loss of chromosome Y (LOY) in normal hematopoietic cells is a well-known phenomenon, the phenotypic consequences of LOY have been elusive. However, LOY has been found in association with smoking, shorter survival and higher risk of cancer. It was suggested that LOY in blood cells could become a predictive biomarker of male carcinogenesis. AIMS, METHODS & FINDINGS: To investigate the association of LOY in blood cells with the risk for development of colorectal (CC) and prostate cancers (PC), we have analyzed DNA samples from peripheral blood of 101 CC male patients (mean age 60.5±11.9 yrs), 70 PC patients (mean age 68.8±8.0 yrs) and 93 healthy control males (mean age 65.8±16.6 yrs). The methodology included co-amplification of homologous sequences on chromosome Y and other chromosomes using multiplex quantitative fluorescent (QF) PCR followed by automatic detection and analysis on ABI 3500 Genetic Analyzer. The mean Y/X ratio was significantly lower in the whole group of cancer patients (0.907±0.12; p = 1.17x10-9) in comparison to the controls (1.015±0.15), as well as in CC (0.884±0.15; p = 3.76x10-9) and PC patients (0.941±0.06; p = 0.00012), when analyzed separately. Multivariate logistic regression analysis adjusting for LOY and age showed that LOY is a more significant predictor of cancer presence than age, and that age probably does not contribute to the increased number of subjects with detectable LOY in cancer patients cohort. CONCLUSION: In conclusion, our results support the recent findings of association of LOY in blood cells with carcinogenesis in males.


Assuntos
Biomarcadores Tumorais/genética , Carcinogênese/genética , Deleção Cromossômica , Cromossomos Humanos Y , Neoplasias Colorretais/diagnóstico , Neoplasias da Próstata/diagnóstico , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/sangue , Carcinogênese/metabolismo , Carcinogênese/patologia , Estudos de Casos e Controles , Neoplasias Colorretais/sangue , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Humanos , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/patologia , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase Multiplex , Estadiamento de Neoplasias , Prognóstico , Neoplasias da Próstata/sangue , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia
15.
PLoS One ; 9(11): e112498, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25386751

RESUMO

Congenital bilateral absence of vas deferens (CBAVD) is the most common CFTR-related disorder (CFTR-RD) that explains about 1-2% of the male infertility cases. Controversial data have been published regarding the involvement of CFTR mutations in infertile men with non-obstructive azoospermia and oligozoospermia. Here, we describe single base extension (SNaPshot) assay for detection of 11 common CFTR mutations: F508del, G542X, N1303K, 621+1G->T, G551D, R553X, R1162X, W1282X, R117H, 2184insA and 1717-1G->A and IVS8polyT variants. The assay was validated on 50 previously genotyped samples and was used to screen a total of 369 infertile men with different impairment of spermatogenesis and 136 fertile controls. Our results show that double heterozygosity of cystic fibrosis (CF) and CFTR-related disorder (CFTR-RD) mutations are found in a high percentage (22.7%) of infertile men with obstructive azoospermia, but not in other studied groups of infertile men. The SNaPshot assay described here is an inexpensive, fast and robust method for primary screening of the most common CFTR mutations both in patients with classical CF and CFTR-RD. It can contribute to better understanding of the role of CFTR mutations in impaired spermatogenesis, ultimately leading to improved management of infertile men.


Assuntos
Regulador de Condutância Transmembrana em Fibrose Cística/genética , Testes Genéticos/métodos , Infertilidade Masculina/genética , Análise Mutacional de DNA , Genótipo , Humanos , Masculino
16.
Reprod Toxicol ; 47: 77-80, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24893173

RESUMO

UNLABELLED: Studies have suggested that nausea and vomiting of pregnancy (NVP) may confer favorable pregnancy outcome, when compared to women not experiencing NVP. However, this was never examined systematically. METHODS: We systematically reviewed all human studies examining potential effects of NVP on rates of miscarriage, intrauterine growth restriction, congenital malformations, prematurity and developmental achievements. RESULTS: Our analysis reveals a consistent favorable effect of NVP on rates of miscarriages, congenital malformations, prematurity, and developmental achievements. The effect size was clinically important for miscarriage, malformations and prematurity. In a few studies the protective effects were more prominent in women with moderate-severe NVP than among those with mild or no NVP. CONCLUSIONS: NVP is associated with favorable fetal outcome, and therefore studies of drug exposure in pregnancy should either match their exposed and control cases for existence and severity of NVP, or adjust for these confounders in their multivariate analysis.


Assuntos
Náusea/epidemiologia , Complicações na Gravidez/epidemiologia , Resultado da Gravidez/epidemiologia , Gravidez , Vômito/epidemiologia , Feminino , Feto , Humanos
17.
Can Fam Physician ; 60(5): 441-2, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24829004

RESUMO

QUESTION: Since the thalidomide disaster, medicine is practised as if every drug is teratogenic, when in fact very few medications are. Pregnant women are often ready to refuse treatment even for life-threatening conditions owing to misinformation and misperceptions about fetal risks. How can I reassure my patients and prevent misinformation from affecting their treatment? ANSWER: Physicians must provide evidence-based counseling to their patients. For example, antihistamines for morning sickness have been proven safe in numerous studies, but are commonly the subject of media reports overstating the risks to the fetus. Family physicians and obstetricians must take an active role in preventing pregnant patients from being misinformed.


Assuntos
Anormalidades Induzidas por Medicamentos , Viés , Aconselhamento Diretivo , Conhecimentos, Atitudes e Prática em Saúde , Educação de Pacientes como Assunto , Cuidado Pré-Natal/métodos , Medicina Baseada em Evidências , Feminino , Humanos , Gravidez , Risco
18.
Prenat Diagn ; 34(10): 970-6, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-24801814

RESUMO

OBJECTIVE: The aim of this study was to determine whether cell free fetal (cff) DNA in residual amniotic fluid (AF) supernatant obtained from bloody, low-volume and late gestation samples can be used for prenatal diagnosis by quantitative fluorescence polymerase chain reaction (QF-PCR) and array comparative genomic hybridization (aCGH). METHOD: A total of 49 compromised AFs were analyzed in this case-control, double-blinded study. The samples were processed through: a conventional cytogenetic approach utilizing Fluorescence in situ Hybridization and/or karyotype (Approach I); QF-PCR analysis to establish the presence of maternal cell contamination (MCC) (Approach II) and a newly proposed approach using AF supernatant cff DNA (Approach III). Data on clinical impact and turn-around-time was collected. RESULTS: Evidence of MCC was not detected in any of the cff DNA samples, and informative results were provided for all cases, including nine aneuploidies. In contrast, the conventional approach (I) failed to provide results either due to MCC or culture failure in a significant proportion of cases. An adequate amount of quality cff DNA was obtained for successful aCGH testing. CONCLUSION: We have shown that it is feasible to isolate pure cff DNA from routinely discarded AF supernatant to perform QF-PCR and microarray analyses, providing timely and informative results even for problematic grossly bloody and otherwise compromised AF samples or culture failures.


Assuntos
Líquido Amniótico/química , Aneuploidia , DNA/química , Diagnóstico Pré-Natal , Estudos de Casos e Controles , Sistema Livre de Células , Hibridização Genômica Comparativa , Feminino , Humanos , Reação em Cadeia da Polimerase , Gravidez , Manejo de Espécimes
19.
Paediatr Drugs ; 16(3): 199-211, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24574047

RESUMO

Nausea and vomiting of pregnancy (NVP) affects up to 85 % of all pregnancies. Effective treatment can greatly improve a woman's quality of life, reduce the risk for maternal and fetal complications, and reduce healthcare costs. Unfortunately, many women receive either no pharmacological treatment or are recommended therapies for which fetal safety and efficacy have not been established. First-line treatment of NVP, as recommended by several leading healthcare and professional organizations, is the combination of doxylamine and pyridoxine. This combination, formulated as a 10 mg/10 mg delayed-release tablet, was approved by the US Food and Drug Administration (FDA) for the treatment of NVP in April 2013 under the brand name Diclegis(®), and has been on the Canadian market since 1979, currently under the brand name Diclectin(®). The efficacy of Diclegis(®)/Diclectin(®) has been demonstrated in several clinical trials, and, more importantly, studies on more than 200,000 women exposed to doxylamine and pyridoxine in the first trimester of pregnancy have demonstrated no increased fetal risk for congenital malformations and other adverse pregnancy outcomes. The present review aims to present the scientific evidence on the effectiveness and fetal safety of Diclegis(®)/Diclectin(®) for the treatment of NVP to justify its use as first-line treatment for NVP.


Assuntos
Antieméticos/uso terapêutico , Diciclomina/uso terapêutico , Doxilamina/uso terapêutico , Náusea/tratamento farmacológico , Complicações na Gravidez/tratamento farmacológico , Piridoxina/uso terapêutico , Vômito/tratamento farmacológico , Antieméticos/farmacologia , Preparações de Ação Retardada/farmacologia , Preparações de Ação Retardada/uso terapêutico , Diciclomina/farmacologia , Doxilamina/farmacologia , Combinação de Medicamentos , Feminino , Humanos , Gravidez , Piridoxina/farmacologia , Resultado do Tratamento
20.
Acta Haematol ; 132(1): 15-21, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24356203

RESUMO

BACKGROUND: Fanconi anemia (FA) is a rare autosomal recessive disorder clinically characterized by developmental abnormalities, progressive bone marrow failure (BMF) and profound cancer predisposition. Approximately 65% of all affected individuals have mutation in the FANCA (Fanconi anemia complementation group A) gene. The mutation spectrum of the FANCA gene is highly heterogeneous. FA-A is usually associated with private FANCA mutations in individual families. METHODS: We describe 3 unrelated patients with FA with a similar clinical presentation: BMF, renal anomalies and café-au-lait pigmentation without major skeletal abnormality. The molecular analysis of the FANCA gene using the FA MLPA kit P031-A2/P032 FANCA, showed homozygous deletion of exon 3 in all 3 patients. Molecular analysis of the flanking regions of exon 3 precisely defined unique deletion of 2,040 bp and duplication of C (1788_3828dupC). DISCUSSION/CONCLUSIONS: These are the first 3 patients homozygous for deletion of FANCA exon 3 described to date. Although not related, the patients originated from the same Gypsy-like ethnic population. We conclude that c.190-256_283 + 1680del2040 dupC mutation in the FANCA gene is a founder mutation in Macedonian FA patients of Gypsy-like ethnic origin. Our finding has very strong implications for these patients in formulating diagnostic and carrier-screening strategy for BMF and FA and to enable comprehensive genetic counseling.


Assuntos
Proteína do Grupo de Complementação A da Anemia de Fanconi/genética , Anemia de Fanconi/genética , Efeito Fundador , Mutação , Adolescente , Criança , Análise Mutacional de DNA , Éxons , Feminino , Homozigoto , Humanos , Masculino , Linhagem , República da Macedônia do Norte , Roma (Grupo Étnico)/genética , Deleção de Sequência , Adulto Jovem
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