Aneuploidy Research Articles

Multiomic Analyses Reveal New Targets of Polycomb Repressor Complex 2 in Schwann Lineage Cells and Malignant Peripheral Nerve Sheath Tumors

Abstract Background Malignant peripheral nerve sheath tumors (MPNST) can arise from atypical neurofibromas (ANF). Loss of the polycomb repressor complex 2 (PRC2) is a common event. Previous studies on PRC2-regulated genes in MPNST used genetic addback experiments in highly aneuploid MPNST cell lines which may miss PRC2-regulated genes in NF1-mutant ANF-like precursor cells. A set of PRC2-regulated genes in human Schwann cells has not been defined. We hypothesized PRC2 loss has direct and indirect effects on gene expression resulting in MPNST, so we sought to identify PRC2-regulated genes in immortalized human Schwann cells (iHSCs). Methods We engineered NF1-deficient iHSCs with loss of function SUZ12 or EED mutations. RNA sequencing revealed 1,327 differentially expressed genes to define PRC2-regulated genes. To investigate MPNST pathogenesis we compared genes in iHSCs to consistent gene expression differences between ANF and MPNSTs. Chromatin immunoprecipitation sequencing was used to further define targets. Methylome and proteomic analyses were performed to further identify enriched pathways. Results We identified potential PRC2-regulated drivers of MPNST progression. Pathway analysis indicates many upregulated cancer-related pathways. We found transcriptional evidence for activated Notch and Sonic Hedgehog signaling in PRC2-deficient iHSCs. Functional studies confirm Notch signaling is active in MPNST cell lines, patient-derived xenografts, and transient cell models of PRC2 deficiency. A combination of MEK and γ-secretase inhibition shows synergy in MPNST cell lines. Conclusions We identified PRC2-regulated genes and potential drivers of MPNSTs. Our findings support the Notch pathway as a druggable target in MPNSTs. Our identification of PRC2-regulated genes and pathways could result in more novel therapeutic approaches.

Investigating the role of RNA-binding protein Ssd1 in aneuploidy tolerance through network analysis.

RNA-binding proteins (RBPs) play critical cellular roles by mediating various stages of RNA life cycles. Ssd1, an RBP with pleiotropic effects, has been implicated in aneuploidy tolerance in Saccharomyces cerevisiae but its mechanistic role remains unclear. Here we used a network-based approach to inform on Ssd1's role in aneuploidy tolerance, by identifying and experimentally perturbing a network of RBPs that share mRNA targets with Ssd1. We identified RBPs whose bound mRNA targets significantly overlap with Ssd1 targets. For 14 identified RBPs, we then used a genetic approach to generate all combinations of genotypes for euploid and aneuploid yeast with an extra copy of chromosome XII, with and without SSD1 and/or the RBP of interest. Deletion of 10 RBPs either exacerbated or alleviated the sensitivity of wild-type and/or ssd1∆ cells to chromosome XII duplication, in several cases indicating genetic interactions with SSD1 in the context of aneuploidy. We integrated these findings with results from a global over-expression screen that identified genes whose duplication complements ssd1∆ aneuploid sensitivity. The resulting network points to a sub-group of proteins with shared roles in translational repression and p-body formation, implicating these functions in aneuploidy tolerance. Our results reveal a role for new RBPs in aneuploidy tolerance and support a model in which Ssd1 mitigates translation-related stresses in aneuploid cells.

Coordination of cell division and chromosome segregation by iron and a sRNA in Escherichia coli

Iron is a vital metal ion frequently present as a cofactor in metabolic enzymes involved in central carbon metabolism, respiratory chain, and DNA synthesis. Notably, iron starvation was previously shown to inhibit cell division, although the mechanism underlying this observation remained obscure. In bacteria, the sRNA RyhB has been intensively characterized to regulate genes involved in iron metabolism during iron starvation. While using the screening tool MAPS for new RyhB targets, we found that the mRNA zapB, a factor coordinating chromosome segregation and cell division (cytokinesis), was significantly enriched in association with RyhB. To confirm the interaction between RyhB and zapB mRNA, we conducted both in vitro and in vivo experiments, which showed that RyhB represses zapB translation by binding at two distinct sites. Microscopy and flow cytometry assays revealed that, in the absence of RyhB, cells become shorter and display impaired chromosome segregation during iron starvation. We hypothesized that RyhB might suppress ZapB expression and reduce cell division during iron starvation. Moreover, we observed that deleting zapB gene completely rescued the slow growth phenotype observed in ryhB mutant during strict iron starvation. Altogether, these results suggest that during growth in the absence of iron, RyhB sRNA downregulates zapB mRNA, which leads to longer cells containing extra chromosomes, potentially to optimize survival. Thus, the RyhB-zapB interaction demonstrates intricate regulatory mechanisms between cell division and chromosome segregation depending on iron availability in E. coli.

The response to single-gene duplication implicates translation as a key vulnerability in aneuploid yeast.

Aneuploidy produces myriad consequences in health and disease, yet models of the deleterious effects of chromosome amplification are still widely debated. To distinguish the molecular determinants of aneuploidy stress, we measured the effects of duplicating individual genes in cells with different chromosome duplications, in wild-type cells (SSD1+) and cells sensitized to aneuploidy by deletion of RNA-binding protein Ssd1 (ssd1Δ). We identified gene duplications that are nearly neutral in wild-type euploid cells but significantly deleterious in euploids lacking SSD1 or in SSD1+ aneuploid cells with different chromosome duplications. Several of the most deleterious genes are linked to translation. In contrast, duplication of other genes benefits multiple ssd1Δ aneuploids over controls, and this group is enriched for translational effectors. Furthermore, both wild-type and especially ssd1Δ aneuploids with different chromosome amplifications show increased sensitivity to translational inhibitor nourseothricin. We used comparative modeling of aneuploid growth defects, based on the cumulative fitness costs measured for single-gene duplication. Our results present a model in which the deleterious effects of aneuploidy emerge from an interaction between the cumulative burden of many amplified genes on a chromosome and a subset of duplicated genes that become toxic in that context. These findings provide a perspective on the dual impact of individual genes and overall genomic burden, offering new avenues for understanding aneuploidy and its cellular consequences.

A Novel Unbalanced Chromosome 8 Translocation with Multiple Congenital Malformations

Unbalanced translocation is estimated to be responsible for about 1% of developmental delay and intellectual disabilities. It is inherited from parent with balanced translocation or appears as a new mutation “Denovo”. Offsprings of balanced translocation’s parents usually display 46 chromosomes including the derivative chromosome and usually partially trisomic and partially monosomic as well. Partial trisomy 8 is responsible for most of the phenotypic features in our case report.

Accurate identification of locally aneuploid cells by incorporating cytogenetic information in single cell data analysis

Single-cell RNA sequencing is a powerful tool to investigate the cellular makeup of tumor samples. However, due to the sparse data and the complex tumor microenvironment, it can be challenging to identify neoplastic cells that play important roles in tumor growth and disease progression. This is especially relevant for blood cancers, where neoplastic cells may be highly similar to normal cells. To address this challenge, we have developed partCNV and partCNVH, two methods for rapid and accurate detection of aneuploid cells with local copy number deletion or amplification. PartCNV uses an expectation-maximization (EM) algorithm with mixtures of Poisson distributions and incorporates cytogenetic information to guide the classification. PartCNVH further improves partCNV by integrating a hidden Markov model for feature selection. We have thoroughly evaluated the performance of partCNV and partCNVH through simulation studies and real data analysis using three scRNA-seq datasets from blood cancer patients. Our results show that partCNV and partCNVH have favorable accuracy and provide more interpretable results compared to existing methods. In the real data analysis, we have identified multiple biological processes involved in the oncogenesis of myelodysplastic syndromes and acute myeloid leukemia.

Unusual Lifespan of Edwards' Syndrome in COVID-19 Era

Edwards' syndrome is caused by an extra chromosome on the 18th chromosome. Patients with this diagnosis have median survival of only 3-14.5 days, with the majority of infants with full trisomy 18 type die before or shortly after they are born. There is only supportive treatment available for Edwards’ syndrome. During the COVID-19 pandemic, providing health services was challenging, leading to less than optimum treatments added by patients’ reluctance to visit healthcare facilities due to the fear of COVID-19 transmission. This case study presented an infant with Edwards’ syndrome with major malformation. The patient had several organ problems, was admitted to the NICU, and continued with home care for monitoring for 6 months. Patient then died at 9 months of age. Maintaining the best quality of life for this patient was very challenging for parents and healthcare providers. Since the management for this type of sydnome is more palliative, good education, information, and psychosocial supports for the parents are needed to prepare them with the worst conditions, which was especially more challenging during the COVID-19 pandemic.

Human embryos harbor complex mosaicism with broad presence of aneuploid cells during early development

Pre-implantation genetic testing for aneuploidy (PGT-A) is used in approximately half of in vitro fertilization cycles. Given the limited understanding of the genetics of human embryos, the current use of PGT-A is based on biologically uncertain assumptions and unvalidated guidelines, leading to the possibility of disposing of embryos with pregnancy potential. We isolated and sequenced all single cells (1133) from in vitro cultured 20 human blastocysts. We found that all blastocysts exhibited mosaicism with mitotic-induced aneuploid cells and showed an ~25% aneuploidy rate per embryo. Moreover, 70% (14/20) of blastocysts contained ‘chromosome-complementary’ cells, suggesting genetic mosaicism is underestimated in routine PGT-A. Additionally, the analysis of 20,945 single cells from day 8–14 embryos (in vitro cultured) and embryonic/fetal organs showed that 97% of the analyzed embryos/organs were mosaic. Over 96% of their aneuploid cells harbored ≤ 2 chromosome errors. Our findings have revealed a high prevalence of mosaicism in human embryos.

Stretchable Composite Films Are Prepared by Using Cellulose Pulp with Differential Substitution of Carboxymethyl Cellulose and Thymol with an Infrared Heating Process for Packaging Application.

Herein, cellulose pulp (CP), carboxymethyl cellulose (CMC), and thymol-based eco-friendly, transparent, and flexible composite films are prepared. These materials dissolved well in an environment-friendly process in N-methyl morpholine N-oxide (NMMO) ionic liquids using infrared heating. Fourier-transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), and scanning electron microscopy (SEM) analyses are used to study the structure, microstructure, and morphology of the composite films. The thermal properties of the CP/CMC/thymol composites are thoroughly studied by thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC). The thermal stability of the composites (T 5%-57.8-91.2 °C and T 10%-216.6-284.1 °C) significantly improves following the CMC addition. In each case, all the composite film exhibits unique T g (140.5-143.1 °C) values, as confirmed by DSC. The tensile strength of the cellulose pulp is 66.5 MPa, increasing to 78.4 MPa for the CP/CMC-1:0.5 composites and steadily increasing to 93.8 MPa with increasing CMC content. Similarly, CMC increases the EB of cellulose pulp from 9.3 to 7.4%. The water vapor permeability and swelling ratio values of the CP/CMC/thymol composites are in the range of 1.9-2.11 × 10-9 g/(m2·Pa·s) and 52.5-50.8° respectively. The CP, CMC, and thymol-based composite do not exhibit cytotoxicity to the aneuploid immortal keratinocyte cell line and demonstrate excellent antioxidant properties, for promising packaging and biomedical applications.

Comprehensive analysis of clinicopathological characteristics in oral squamous cell carcinoma arising from oral submucous fibrosis: a retrospective study

BackgroundOral squamous cell carcinoma arising from oral submucous fibrosis is recognized as a clinicopathologically distinct disease. This study aims to summarize and compare the population and clinicopathologic characteristics of OSCC-OSF with other OSCC in Hunan Province, China. The objective is to formulate treatment strategies more in line with regional characteristics.MethodsA retrospective review of OSCC cases recruited from the Second Xiangya Hospital from 2010 to 2020 was conducted. A total of 1,413 OSCC patients were selected, including 481 patients with OSCC-OSF and 932 were other OSCC. Population characteristics, risk factors and clinicopathological manifestations were explored in OSCC-OSF and other OSCC, as well as thinprep cytologic test and DNA quantification.ResultsWe found that OSCC-OSF patients were younger than those with other OSCC. Both types of disease were predominantly observed in males compared to females. Tumor biopsy analysis indicated that tumor cells within OSCC-OSF patients were more likely to be well differentiated and showed a higher frequency of lymph node metastases. Clinicopathological factors, such as the chewing betel nuts habit and smoking, were more prevalent in OSCC-OSF patients in contrast to other OSCC. DNA quantification revealed that the number of DNA aneuploid cells was higher in OSCC-OSF compared to other OSCCs.ConclusionIn this study, OSCC-OSF is considered a clinicopathologically distinct disease. Compared with other OSCC, OSCC-OSF patients have a higher incidence of nodal metastases, an early clinical TNM stage, and a lower three-year survival rate. Therefore, early diagnosis and treatment are crucial to improving the prognosis of OSCC-OSF.

Increased RNA and protein degradation is required for counteracting transcriptional burden and proteotoxic stress in human aneuploid cells.

Aneuploidy results in a stoichiometric imbalance of protein complexes that jeopardizes cellular fitness. Aneuploid cells thus need to compensate for the imbalanced DNA levels by regulating their RNA and protein levels, but the underlying molecular mechanisms remain unknown. Here, we dissected multiple diploid vs. aneuploid cell models. We found that aneuploid cells cope with transcriptional burden by increasing several RNA degradation pathways, and are consequently more sensitive to the perturbation of RNA degradation. At the protein level, aneuploid cells mitigate proteotoxic stress by reducing protein translation and increasing protein degradation, rendering them more sensitive to proteasome inhibition. These findings were recapitulated across hundreds of human cancer cell lines and primary tumors, and aneuploidy levels were significantly associated with the response of multiple myeloma patients to proteasome inhibitors. Aneuploid cells are therefore preferentially dependent on several key nodes along the gene expression process, creating clinically-actionable vulnerabilities in aneuploid cells.

Human aneuploid cells depend on the RAF/MEK/ERK pathway for overcoming increased DNA damage

Aneuploidy is a hallmark of human cancer, yet the molecular mechanisms to cope with aneuploidy-induced cellular stresses remain largely unknown. Here, we induce chromosome mis-segregation in non-transformed RPE1-hTERT cells and derive multiple stable clones with various degrees of aneuploidy. We perform a systematic genomic, transcriptomic and proteomic profiling of 6 isogenic clones, using whole-exome DNA, mRNA and miRNA sequencing, as well as proteomics. Concomitantly, we functionally interrogate their cellular vulnerabilities, using genome-wide CRISPR/Cas9 and large-scale drug screens. Aneuploid clones activate the DNA damage response and are more resistant to further DNA damage induction. Aneuploid cells also exhibit elevated RAF/MEK/ERK pathway activity and are more sensitive to clinically-relevant drugs targeting this pathway, and in particular to CRAF inhibition. Importantly, CRAF and MEK inhibition sensitize aneuploid cells to DNA damage-inducing chemotherapies and to PARP inhibitors. We validate these results in human cancer cell lines. Moreover, resistance of cancer patients to olaparib is associated with high levels of RAF/MEK/ERK signaling, specifically in highly-aneuploid tumors. Overall, our study provides a comprehensive resource for genetically-matched karyotypically-stable cells of various aneuploidy states, and reveals a therapeutically-relevant cellular dependency of aneuploid cells.

Psychiatric disorders and comorbidity in women with Turner Syndrome: a retrospective national cohort study

Turner syndrome (TS) is a genetic condition characterized by partial or complete monosomy X. A reduced life expectancy has been shown in TS, depending on an increased risk of aortic dissection, and ischemic heart disease. Studies covering the occurrence of psychiatric conditions are sparse within TS. Several case reports describe concomitant TS and neuropsychiatric abnormalities that may represent a pathogenetic link to genetics, as well as feature correlates of TS. The aim of this study was to determine the presence, and the frequency of psychiatric diagnosis in women with TS in a Swedish cohort followed during 25 years’ time. Statistics from the entire female population in Sweden of corresponding age was used as reference. Data were retrieved from clinical examinations and validated from the National Board of Health and Welfare registries for women with TS (n = 487), aged 16 to 84 years, with respect to mental health disorders. The most common diagnoses in TS were mood and anxiety disorders. There was no increase in psychiatric diagnosis within the group with time, nor correlation to specific karyotype or somatic comorbidity as congenital heart disease and hypothyroidism, hormonal treatment, or childbirth. In addition, the frequency of psychiatric diagnosis in TS was lower than in the population-based data. Further investigations are needed in the view of the fact that women with Turner syndrome should not be burdened with more severe diagnoses.

Imprinting as Basis for Complex Evolutionary Novelties in Eutherians.

The epigenetic phenomenon of genomic imprinting is puzzling. While epigenetic modifications in general are widely known in most species, genomic imprinting in the animal kingdom is restricted to autosomes of therian mammals, mainly eutherians, and to a lesser extent in marsupials. Imprinting causes monoallelic gene expression. It represents functional haploidy of certain alleles while bearing the evolutionary cost of diploidization, which is the need of a complex cellular architecture and the danger of producing aneuploid cells by mitotic and meiotic errors. The parent-of-origin gene expression has stressed many theories. Most prominent theories, such as the kinship (parental conflict) hypothesis for maternally versus paternally derived alleles, explain only partial aspects of imprinting. The implementation of single-cell transcriptome analyses and epigenetic research allowed detailed study of monoallelic expression in a spatial and temporal manner and demonstrated a broader but much more complex and differentiated picture of imprinting. In this review, we summarize all these aspects but argue that imprinting is a functional haploidy that not only allows a better gene dosage control of critical genes but also increased cellular diversity and plasticity. Furthermore, we propose that only the occurrence of allele-specific gene regulation mechanisms allows the appearance of evolutionary novelties such as the placenta and the evolutionary expansion of the eutherian brain.

THE COMPARATIVE STUDY OF THE EFFECT OF LOW-INTENSITY BROADBAND AND LOW-INTENSITY PULSED ULTRASOUND ON B16 MELANOMA CELLS IN VITRO

Aim. To evaluate the effect of low-intensity broadband ultrasound (UMUS) and low-intensity pulsed ultrasound (LIPUS) on B16 melanoma cells in vitro. Methods. The research was conducted on B16 melanoma cells under two modes of low-intensity ultrasonic sounding of cells, for which an SDG 2082 X Siglent generator (USA) with an ultrasonic emitter diameter of 20 mm was used; with a frequency for UMUS of 1-7 MHz, intensity of 30 mW/cm, pulse duration of 50 μs, and for LIPUS with a frequency of 1.5 MHz, intensity of 30 mW/cm, pulse duration of 200 μs. Both modes of cell sounding were carried out with threefold irradiation, once a day after which the distribution of cells by cycle phases, the level of apoptosis and proliferative activity was determined by the method of ductal cytofluorimetry. Results. Under the influence of UMUS ultrasonic sounding, the inhibition of the proliferation of melanoma cells by 1.6 times (P<0.05), a 2-fold decrease in the content of aneuploid cells, compared to the control, was found, however, cytostatic and proapoptotic effects were not recorded. Under the influence of LIPUS sounding, an anti-proliferative effect, an increase in the content of dead and apoptotic cells and a decrease in the DI-index compared to the corresponding control were found. Conclusions. Under the action of low-intensity broadband ultrasound (UMUS) and low-intensity pulsed ultrasound (LIPUS), a cytotoxic/cytostatic and pro-apoptotic effect and a decrease in the content of the aneuploid cell population were revealed.

The Role of Diet and Physical Activity in Obesity and Overweight in Children with Down Syndrome in Developed Countries.

Down's syndrome (DS), or Trisomy 21, represents the most common chromosomal abnormality in live births, characterized by an extra chromosome 21. Children affected by Down's syndrome are more susceptible to the development of obesity and of becoming overweight compared with other children. Furthermore, they seem to present a more unfavorable lipid profile than the non-DS obese pediatric population. Diet and physical activity are closely related to the development of overweight and obesity, and they can be assessed using questionnaires such as the Mediterranean Diet Quality Index in children and adolescents (KIDMED) and the Godin-Shephard Leisure-Time Physical Activity Questionnaire. This review aims to undertake a comprehensive analysis of the intricate interplay between diet and physical activity in children affected by Down's syndrome. Specifically, it seeks to deepen understanding regarding the question of how diet and exercise can influence and prevent the development of overweight and obesity in that special pediatric population.

Energy‑stress‑mediated activation of AMPK sensitizes MPS1 kinase inhibition in triple‑negative breast cancer.

Monopolar spindle 1 kinase (Mps1, also known as TTK protein kinase) inhibitors exert marked anticancer effects against triple‑negative breast cancer (TNBC) by causing genomic instability and cell death. As aneuploid cells are vulnerable to compounds that induce energy stress through adenosine monophosphate‑activated protein kinase (AMPK) activation, the synergistic effect of Mps1/TTK inhibition and AMPK activation was investigated in the present study. The combined effects of CFI‑402257, an Mps1/TTK inhibitor, and AICAR, an AMPK agonist, were evaluated in terms of cytotoxicity, cell‑cycle distribution, and in vivo xenograft models. Additional molecular mechanistic studies were conducted to elucidate the mechanisms underlying apoptosis and autophagic cell death. The combination of CFI‑402257 and AICAR showed selective cytotoxicity in a TNBC cell line. The formation of polyploid cells was attenuated, and apoptosis was increased by the combination treatment, which also induced autophagy through dual inhibition of the PI3K/Akt/mTOR and mitogen‑activated protein kinase (MAPK) signaling pathways. Additionally, the combination therapy showed strongly improved efficacy in comparison with CFI‑402257 and AICAR monotherapy in the MDA‑MB‑231 xenograft model. The present study suggested that the combination of CFI‑402257 and AICAR is a promising therapeutic strategy for TNBC.

67. An undiagnosed chronic myeloid leukemia (CML) with p190 BCR::ABL1 transcript, an extra Philadelphia chromosome, and IKARO

67. An undiagnosed chronic myeloid leukemia (CML) with p190 BCR::ABL1 transcript, an extra Philadelphia chromosome, and IKARO

Investigating the role of RNA-binding protein Ssd1 in aneuploidy tolerance through network analysis.

RNA-binding proteins (RBPs) play critical cellular roles by mediating various stages of RNA life cycles. Ssd1, an RBP with pleiotropic effects, has been implicated in aneuploidy tolerance in Saccharomyces cerevisiae but its mechanistic role remains unclear. Here we used a network-based approach to inform on Ssd1's role in aneuploidy tolerance, by identifying and experimentally perturbing a network of RBPs that share mRNA targets with Ssd1. We identified RBPs whose bound mRNA targets significantly overlap with Ssd1 targets. For 14 identified RBPs, we then used a genetic approach to generate all combinations of genotypes for euploid and aneuploid yeast with an extra copy of chromosome XII, with and without SSD1 and/or the RBP of interest. Deletion of 10 RBPs either exacerbated or alleviated the sensitivity of wild-type and/or ssd1 Δ cells to chromosome XII duplication, in several cases indicating genetic interactions with SSD1 in the context of aneuploidy. We integrated these findings with results from a global over-expression screen that identified genes whose duplication complements ssd1 Δ aneuploid sensitivity. The resulting network points to a sub-group of proteins with shared roles in translational repression and p-body formation, implicating these functions in aneuploidy tolerance. Our results reveal a role for new RBPs in aneuploidy tolerance and support a model in which Ssd1 mitigates translation-related stresses in aneuploid cells.

HIF1A contributes to the survival of aneuploid and mosaic pre-implantation embryos.

Human fertility is suboptimal, partly due to error-prone divisions in early cleavage-stages that result in aneuploidy. Most human pre-implantation are mosaics of euploid and aneuploid cells, however, mosaic embryos with a low proportion of aneuploid cells have a similar likelihood of developing to term as fully euploid embryos. How embryos manage aneuploidy during development is poorly understood. This knowledge is crucial for improving fertility treatments and reducing developmental defects. To explore these mechanisms, we established a new mouse model of chromosome mosaicism to study the fate of aneuploid cells during pre-implantation development. We previously used the Mps1 inhibitor reversine to generate aneuploidy in embryos. Here, we found that treatment with the more specific Mps1 inhibitor AZ3146 induced chromosome segregation defects in pre-implantation embryos, similar to reversine. However, AZ3146-treated embryos showed a higher developmental potential than reversine-treated embryos. Unlike reversine-treated embryos, AZ3146-treated embryos exhibited transient upregulation of Hypoxia Inducible-Factor-1A (HIF1A) and lacked p53 upregulation. Pre-implantation embryos develop in a hypoxic environment in vivo, and hypoxia exposure in vitro reduced DNA damage in response to Mps1 inhibition and increased the proportion of euploid cells in the mosaic epiblast. Inhibiting HIF1A in mosaic embryos also decreased the proportion of aneuploid cells in mosaic embryos. Our work illuminates potential strategies to improve the developmental potential of mosaic embryos.