Association Of Aneuploidy Research Articles

Aneuploidy in oocytes from women of advanced maternal age: analysis of the causal meiotic errors and impact on embryo development.

In oocytes of advanced maternal age (AMA) women, what are the mechanisms leading to aneuploidy and what is the association of aneuploidy with embryo development? Known chromosome segregation errors such as precocious separation of sister chromatids explained 90.4% of abnormal chromosome copy numbers in polar bodies (PBs), underlying impaired embryo development. Meiotic chromosomal aneuploidies in oocytes correlate with AMA (>35 years) and can affect over half of oocytes in this age group. This underlies the rationale for PB biopsy as a form of early preimplantation genetic testing for aneuploidy (PGT-A), as performed in the 'ESHRE STudy into the Evaluation of oocyte Euploidy by Microarray analysis' (ESTEEM) randomized controlled trial (RCT). So far, chromosome analysis of oocytes and PBs has shown that precocious separation of sister chromatids (PSSC), Meiosis II (MII) non-disjunction (ND), and reverse segregation (RS) are the main mechanisms leading to aneuploidy in oocytes. Data were sourced from the ESTEEM study, a multicentre RCT from seven European centres to assess the clinical utility of PGT-A on PBs using array comparative genomic hybridization (aCGH) in patients of AMA (36-40 years). This included data on the chromosome complement in PB pairs (PGT-A group), and on embryo morphology in a subset of embryos, up to Day 6 post-insemination, from both the intervention (PB biopsy and PGT-A) and control groups. ESTEEM recruited 396 AMA patients: 205 in the intervention group and 191 in the control group. Complete genetic data from 693 PB pairs were analysed. Additionally, the morphology from 1034 embryos generated from fertilized oocytes (two pronuclei) in the PB biopsy group and 1082 in the control group were used for statistical analysis. Overall, 461/693 PB pairs showed abnormal segregation in 1162/10 810 chromosomes. The main observed abnormal segregations were compatible with PSSC in Meiosis I (MI) (n = 568/1162; 48.9%), ND of chromatids in MII or RS (n = 417/1162; 35.9%), and less frequently ND in MI (n = 65/1162; 5.6%). For 112 chromosomes (112/1162; 9.6%), we observed a chromosome copy number in the first PB (PB1) and second PB (PB2) that is not explained by any of the known mechanisms causing aneuploidy in oocytes. We observed that embryos in the PGT-A arm of the RCT did not have a significantly different morphology between 2 and 6 days post-insemination compared to the control group, indicating that PB biopsy did not affect embryo quality. Following age-adjusted multilevel mixed-effect ordinal logistic regression models performed for each embryo evaluation day, aneuploidy was associated with a decrease in embryo quality on Day 3 (adjusted odds ratio (aOR) 0.62, 95% CI 0.43-0.90), Day 4 (aOR 0.15, 95% CI 0.06-0.39), and Day 5 (aOR 0.28, 95% CI 0.14-0.58). RS cannot be distinguished from normal segregation or MII ND using aCGH. The observed segregations were based on the detected copy number of PB1 and PB2 only and were not confirmed by the analysis of embryos. The embryo morphology assessment was static and single observer. Our finding of frequent unexplained chromosome copy numbers in PBs indicates that our knowledge of the mechanisms causing aneuploidy in oocytes is incomplete. It challenges the dogma that aneuploidy in oocytes is exclusively caused by mis-segregation of chromosomes during MI and MII. Data were mined from a study funded by ESHRE. Illumina provided microarrays and other consumables necessary for aCGH testing of PBs. None of the authors have competing interests. Data were mined from the ESTEEM study (ClinicalTrials.gov Identifier NCT01532284).

O-307 Aneuploidy in oocytes of women of advanced maternal age: impact on embryo development and evidence for a novel mechanism of meiotic error

Abstract Study question In oocytes of advanced maternal age (AMA) women, what is the association of aneuploidy with embryo development and what are the mechanisms leading to aneuploidy? Summary answer We confirmed precocious separation of sister chromatids as the main cause of aneuploidy. Known chromosome segregation errors explained 90.4% of the observed abnormal chromosome copy numbers in polar bodies. What is known already Meiotic chromosomal aneuploidies in oocytes correlate with AMA (>35 years), and can affect over half of oocytes in this age group. This underlies the rationale for polar body (PB) biopsy as a form of early PGT-A, as performed in the ‘ESHRE STudy into the Evaluation of oocyte Euploidy by Microarray analysis’ (ESTEEM) randomised controlled trial (RCT). So far, chromosome analysis of oocytes and PBs has shown that precocious separation of sister chromatids (PSSC), meiosis II (MII) nondisjunction (ND) and reverse segregation (RS) are the main mechanisms leading to aneuploidy in oocytes. Study design, size, duration Data was sourced from the ESTEEM study, a multicentre RCT from seven European centres to assess the clinical utility of PGT-A on PBs using array comparative genomic hybridisation (aCGH) in patients of AMA (36-40 years). This included data on the chromosome complement in PB pairs, and on embryo morphology in a subset of embryos, up to day 6 post insemination (dpi), from both the intervention (PB biopsy for PGT-A) and from control (no biopsy) arms. Participants/materials, setting, methods ESTEEM recruited 396 AMA patients: 205 in the intervention group and 191 in the control group. Complete genetic data from 693 PB pairs were analysed. Additionally, the morphology from 1034 embryos generated from fertilized oocytes (2 pronuclei) in the PB biopsy group and 1082 in the control group were compared using age-adjusted multilevel mixed-effect ordinal logistic regression for each day of embryo scoring and summarized with adjusted odds ratios (aOR) and 95% confidence intervals (CI). Main results and the role of chance Overall, 461/693 PB pairs showed abnormal segregation in 1162/10810 chromosomes. The main observed abnormal segregations were compatible with PSSC in meiosis I (MI) (n = 568/1162; 48,9%), ND of chromatids in MII or RS (n = 417/1162; 35.9%) and less frequently ND in MI (n = 65/1162; 5.6%). For 112 chromosomes (112/1162; 9,6%), we observed a chromosome copy number in PB1 and PB2 that is not explained by any of the known mechanisms causing aneuploidy in oocytes. We observed that embryos in the PGT-A arm of the RCT did not have a significantly different morphology between 2 and 6 dpi compared to the control group, indicating that polar body biopsy did not affect embryo quality. Following age-adjusted multilevel mixed-effect ordinal logistic regression models performed for each embryo evaluation day (using the before-mentioned embryo scoring subcategories), aneuploidy was associated with a decrease in embryo quality on day 3 (aOR 0.62, 95% CI 0.43-0.90), day 4 (aOR 0.15, 95% CI 0.06-0.39) and day 5 (aOR 0.28, 95% CI 0.14-0.58). Limitations, reason for caution RS cannot be distinguished from normal segregation or MII ND using aCGH. The observed segregations were based on the detected copy number of PB1 and PB2 only and were not confirmed by the analysis of embryos. The embryo morphology assessment was static and single observer. Wider implications of the findings Our finding of frequent unexplained chromosome copy numbers in polar bodies indicates that our knowledge of the mechanisms causing aneuploidy in oocytes is incomplete. It challenges the dogma that aneuploidy in oocytes is exclusively caused by missegregation of chromosomes during meiosis I and II. Study funding/competing interest(s) Data was mined from a study funded by ESHRE. Illumina provided microarrays and other consumables necessary for aCGH testing of polar bodies. None of the authors have competing interests. This work is dedicated to the memory of Professor Joep Geraedts. Trial registration number NCT01532284 Study funding Yes Funding source Funding by national/international organization(s)

Which factors are correlated with embryo euploidy in PGT-A cycles? - A multivariate logistic regression analysis of 3054 embryos

Introduction Aneuploidy is generally accepted as one of the main causes of IVF/ICSI treatment failures worldwide. For this reason, increasing number of clinics & infertility specialists seek and promote preimplantation genetic testing for aneuploidy (PGT-A) to improve the clinical outcome for indications advanced maternal age (AMA), repeated implantation failure (RIF), recurrent miscarriage (RM), severe male factor infertility. Once an euploid embryo is found, elective single euploid blastocyst transfer (eSEBT) has been shown to improve the overall clinical outcome by providing high implantation rates and lower multiple pregnancy rates. Although It is well known that aneuploidy is strongly correlated with maternal biologic age, the possibility of finding an euploid embryo in a given cycle remains largely unknown and association of aneuploidy with many patient or cycle-related parameters remain controversial. The aim of this study was to evaluate which patient, cycle or laboratory factors have an impact on aneuploidy rate in euploidy for blastocyst stage human embryos. Material & Methods This retrospective study compiled 1362 ICSI cycles of 1136 patients undergoing PGT-A with clinical indications recurrent implantation failure (RIF), recurrent miscarriage (RM) and advanced maternal age (AMA) between January 2016 and December 2018. Trophectoderm(TE) biopsy was performed and next generation sequencing (NGS) was used for comprehensive chromosomal screening (CCS). We checked the possible influence of patient characteristics, treatment history, ovarian stimulation variables, embryo scores and day on aneuploidy. To identify characteristics that may be associated with the euploidy rate multivariate logistic regression analysis was performed. Generalized estimating equations were used after we had taken into account the patient effect. Results The mean age of patients was 38.1±4.7 years. Of these 1136 patients 187(16,5%) have male factor, 256(22,5%) diminished ovarian reserve, 83(7,3%) PCOS, (9,4%) 131(11.5%) endometriosis, 89(7,8%) tubal factor, 277(24,3%) unexplained infertility, and 113(9,9%) male and female factor. Clinical indications of PGT-A these of 1136 patients were RIF in 552(48,5%), RM in 162(14,2%), AMA in 270(23,7%), RIF+AMA in 152(13,4%). Out of 3084 embryos that were biopsied at blastocyst stage, 3054 concluded with valid result (99.02%). After analysis of biopsied materials by NGS, 1327 (43.5%) were found to be euploid and 1727(56,5%) were aneuploid. Severe male factor infertility, duration of ovarian stimulation, total dose of gonadotropins used, number of oocytes retrieved, serum estradiol level on the day of hCG trigger, serum progesterone level on the day of trigger, trophectoderm and inner cell mass scores have been found to be the parameters which significantly affect euploidy in univariate analysis (p Conclusions Due to the limited number of embryos available for embryo biopsy, relative estimation of finding a euploid embryo, which is one of the most frustrating issue for both patients and clinicians, remains a challenge in PGT-A cycles. Our study and results shed light on the nature as well as the magnitude of the patient-, cycle- or laboratory-related parameters that can be used to predict the probability of euploidy before a PGT-A is planned.

Simultaneous, single cell, flow cytometric quantification of PD-L1/TILs/cell cycle in non-small cell lung cancer tissue biopsies: Concordance of PD-L1 expression detection between flow cytometry and immunohistochemistry.

159 Background: Tumor cell expression of PD-L1 leads to the inhibition of immune responses specifically inactivation of cytotoxic T-cells. PD-L1 expression on tumor cells by immunohistochemistry does not provide the complete picture of therapeutic (PD-L1) and prognostic (TILs, aneuploidy) markers. Here, we report a clinical assay that quantifies tumor infiltrating lymphocytes (TILs), cell cycle/DNA content as well as PD-L1 expression on tumor and aneuploid tumor cell populations. Methods: Punch biopsies were taken from 19 fresh tissues specimens and were processed into single cell suspensions using the IVD/CE-IVD IncellPREP Single Cell Preparation Kit. Cells were fixed and permeabilized in 1 mL IncellMax. Cells were tested with the OncoTect iO Lung Assay which contains antibodies directed against PD-L1 (clone 28-8), CD45, CD3, and CD8, and a cell cycle dye. Concordance to IHC was tested by the Dako PD-L1 IHC 28-8 PharmDx Kit. Immune cell populations were quantified and aneuploidy was determined using a DNA index > 1.05. Results: The number of CD8+ CTL were significantly increased (P = 0.02) in tumor compared to normal lung tissue (37.4% to 28.2%). The number of CTL in aneuploid tumors was twice the number of CTL in diploid tumors. The percentage of antigen presenting cells (CD45+, high SSC) was decreased in the tumor cell samples relative to the normal lung tissue (P = 0.01). PD-L1 expression varies in cells depending on the cell cycle. Conclusions: In this study, the concordance between Oncotect iO and IHC was 95% (NPA-97%, PPA-89%). PD-L1 expression varies with DNA content, PD-L1 expression decreases with increasing DNA content. Though CTL are increased in aneuploid tumors, PD-L1 expression decreasing with DNA content may contribute to the association of aneuploidy with distant metastases. PD-L1 expression is a powerful determinant of treatment and aneuploidy is a powerful prognostic factor that combined yield important clinical information.

Association of Aneuploidy and Flat Dysplasia With Development of High-Grade Dysplasia or Colorectal Cancer in Patients With Inflammatory Bowel Disease

There is controversy over how to best manage patients with inflammatory bowel disease and flat low-grade dysplasia (fLGD) in the colon. We performed a retrospective analysis of formalin-fixed paraffin-embedded colon tissues with fLGD from 37 patients undergoing surveillance colonoscopy for inflammatory bowel disease from 1990 to 2015 at the University of California at San Francisco Medical Center, to determine whether detection of aneuploidy is associated with later development of high-grade dysplasia (HGD) or colorectal cancer. Medical data were collected from the patients for a mean follow-up time of 37 months. Using flow cytometry analysis of paraffin-embedded colon tissue, we detected aneuploidy in 15 of 37 samples with fLGD (40.5%). By comparison, aneuploidy was detected in 14 of 15 samples with flat HGD (93.3%) and 2 of 45 samples that were negative for dysplasia (4.4%). The univariate hazard ratio for subsequent detection of HGD or colorectal cancer in patients with fLGD and aneuploidy was 5.3 (95% CI, 1.542-24.121) within a mean follow-up time of 37 months. The presence of aneuploidy therefore identifies patients with fLGD in colon tissue who have an increased risk for HGD or colorectal cancer and may provide supportive evidence to a morphologic impression or suspicion of flat HGD.

What is Wrong with the Human Reproductive System?

What is Wrong with the Human Reproductive System? Human reproduction is highly inferior by mammalian standards. This is manifest as: (i) poor semen quality; (ii) a lower probability of conception; (iii) an increased risk of aneuploidy embryos (e.g. Down syndrome); (iv) a high risk of embryonic loss (early miscarriage); and (v) the recent rise of germ-cell testicular cancer, which occurs in few non-human mammalian species. Review and synthesis of the evidence from several distinct research areas reveals a shared pathogenesis underlying at least some cases of all of them. This involves an intergenerational process, starting with DNA damage to round spermatids. DNA repair in the oocyte is faulty, leading to abnormal chromosome structure in the surviving embryos. This is amplified during meiosis due to delayed synapsis, leading to cellular disorganization. This mechanism also explains the association of aneuploidy with maternal age: structural mismatch between maternal and paternal chromosomes reduces the number of crossovers, making segregation errors more likely.

Association of aneuploidy with multiple neurodegenerative diseases and its possible involvement in the progression of cognitive decline

Association of aneuploidy with multiple neurodegenerative diseases and its possible involvement in the progression of cognitive decline

Securin predicts aneuploidy and survival in breast cancer

Securin is known to participate in maintaining chromosomal integrity during the cell cycle through regulation of metaphase-anaphase transition, DNA damage repair, and apoptosis. The aim of this study was to investigate the role of securin in aneuploidy and prognosis in human breast cancer. The study was based on 603 breast cancer patients with up to 20 years of follow-up. DNA content was determined by image cytometry on cell imprints, and securin immunohistochemistry was performed on tissue microarrays of breast cancer tissue. We show, for the first time in human breast cancer, that high-level securin expression predicts abnormal DNA content, with up to 9.8-fold odds for aneuploid DNA content (P = 0.0007). Securin also shows strong independent prognostic value for disease-specific survival, with a significant difference in survival time between patients with low-level and high-level securin expression. The main result of the present study is the association of aneuploidy and securin expression. According to our results, securin immunohistochemistry is also a potential new prognosticator for treatment decisions concerning breast cancer patients.

The relationship between blastocyst morphology, chromosomal abnormality, and embryo gender

To assess correlation between blastocyst morphology and chromosomal status. Observational research study. An IVF clinic and a specialist preimplanation genetic diagnosis (PGD) laboratory. Ninety-three couples undergoing IVF treatment in combination with chromosome screening of embryos. Five hundred blastocysts underwent trophectoderm biopsy and comprehensive chromosome screening using comparative genomic hybridization (CGH). The morphology of the embryos was evaluated using standard methods. Association of aneuploidy and morphologic score. A total of 56.7% of blastocysts were aneuploid. One-half of the grade 5/6 blastocysts were euploid, compared with only 37.5% of embryos graded 1/2, suggesting an effect of aneuploidy on blastocyst development. Aneuploidy also had a negative effect on inner cell mass and trophectoderm grades. Morphologically poor blastocysts had a higher incidence of monosomy and abnormalities affecting several chromosomes. The gender ratio was significantly skewed in relation to morphology. A total of 72% of blastocysts attaining the highest morphologic scores (5AA and 6AA) were found to be male, compared with only 40% of grade 3 embryos. Morphology and aneuploidy are linked at the blastocyst stage. However, the association is weak, and consequently, morphologic analysis cannot be relied on to ensure transfer of chromosomally normal embryos. A significant proportion of aneuploid embryos are capable of achieving the highest morphologic scores, and some euploid embryos are of poor morphology. Gender was associated with blastocyst grading, male embryos developing at a significantly faster rate than females.

The association of aneuploidy and mild fetal pyelectasis in an unselected population: the results of a multicenter study.

Mild pyelectasis is a common finding and there is some debate as to its association with aneuploidy. The results of a large, prospective, multicenter study of mild pyelectasis designed to determine the incidence and association with aneuploidy in an unselected population are reported. A large multicenter, prospective observational study of unselected fetuses with mild pyelectasis identified between 16 and 26 weeks' gestation in routine ultrasound departments. There were 737 fetuses with mild pyelectasis of which 12 had an abnormal karyotype. Pyelectasis was isolated in three fetuses with Down syndrome, but in one the mother was older (36 years). These data confirm the fact that the presence of mild fetal pyelectasis increases the risk for aneuploidy, in particular trisomy 21. However, other risk factors should be considered before embarking on fetal karyotyping, as for most pregnancies complicated by isolated mild pyelectasis, risks of aneuploidy will remain small.

Alphoid variant-specific FISH probes can distinguish autosomal meiosis I from meiosis II non-disjunction in human sperm

Over the past few years, several groups have used fluorescence in situ hybridization (FISH) to study aneuploidy in human sperm. Several important observations have derived from these studies, including the demonstration of chromosome-specific variation in non-disjunction frequencies, and the possible association of aneuploidy with environmental agents and with increasing paternal age. However, an important technical limitation of these studies has been the inability to distinguish between autosomal non-disjunction occurring at meiosis I and meiosis II. In the present report, we describe a simple FISH-based approach designed to overcome this limitation. Using oligonucleotide probes capable of distinguishing subtle differences in the alpha satellite sequences of chromosome 17, we demonstrate that (in appropriate heterozygotes) it is possible to simultaneously identify disomic sperm and to determine the meiotic stage of origin of the additional chromosome. This novel approach has important implications for future FISH sperm studies, since the ability to distinguish between meiosis I and meiosis II non-disjunction will make it possible to determine whether putative etiological agents affect chromosome segregation at both, or only one, of the two meiotic stages.

Flow cytometric DNA analysis of colorectal carcinoid

We analyzed the patterns of DNA ploidy in 31 colorectal carcinoid tumors from paraffin-embedded tissues by DNA flow cytometry and the relationship of the patterns of DNA ploidy to prognosis. Diploid DNA was found in 78% (24 of 31) of carcinoids and tetraploid in 6% (2 of 31). Five (16%) carcinoids were DNA aneuploid, and four of the patients with aneuploidy showing a near-hypertriploid pattern died during the first 5 years of follow-up. The association of aneuploidy with stage, size, and invasion of tumor was significant. However, our data indicated that DNA aneuploidy of a near-hypertriploid pattern was the most precise and reliable parameter for predicting the prognosis of colorectal carcinoid tumors.

DNA flow cytometry of epithelioid sarcoma.

DNA ploidy of paraffin-embedded tissue from 11 patients with epithelioid sarcoma and 20 control specimens (granuloma annulare and sarcoid) was studied. DNA aneuploidy was found in 64% of the epithelioid sarcomas and in none of the control samples. Aneuploidy did not correlate with necrosis, vascular invasion, or mitotic rate. There was also no association of aneuploidy with adverse outcome.

Suppression of differentiation phenotypes in myogenic cells: Association of aneuploidy and altered regulation of c- myc gene expression

We isolated 10 myoblast clones from Syrian/golden hamster embryo (SHE) cells irradiated with ultraviolet light. They were originally isolated as anchorage-independent clones. All clones showed characteristic morphology of myoblast in culture and formed swirled myofiber colonies specific to myoblast macrocolonies. However, in immunostaining experiments using anti-fast myosin antibody, we found that 4 of 10 myoblast clones were myosin negative (Myo −), while six were myosin positive (Myo +). Western blot analysis confirmed the disappearance of the 200-kDa myosin-specific band in the 4 Myo − clones. Furthermore, these 4 Myo − clones lost the ability to form multinucleated myotubes. Karyotype analysis revealed that all Myo − clones had trisomy of chromosome 7, while Myo + clones showed no apparent karyotypic change from normal SHE cells. Compared with that of SHE cells, the transcriptional level of the myc gene in Myo + clones was augmented, but there was no increase of myc gene expression in Myo − clones. Furthermore, the introduction of activated myc gene partially converted the Myo − phenotype to Myo +. These results suggest that trisomy of chromosome 7 and a deficiency in enhanced expression of the myc oncogene are associated with the suppression of both the production of myosin and the formation of multinucleated cells.