Unbalanced Chromosome Research Articles

Consequences of Chromosome Loss: Why Do Cells Need Each Chromosome Twice?

Aneuploidy is a cellular state with an unbalanced chromosome number that deviates from the usual euploid status. During evolution, elaborate cellular mechanisms have evolved to maintain the correct chromosome content over generations. The rare errors often lead to cell death, cell cycle arrest, or impaired proliferation. At the same time, aneuploidy can provide a growth advantage under selective conditions in a stressful, frequently changing environment. This is likely why aneuploidy is commonly found in cancer cells, where it correlates with malignancy, drug resistance, and poor prognosis. To understand this “aneuploidy paradox”, model systems have been established and analyzed to investigate the consequences of aneuploidy. Most of the evidence to date has been based on models with chromosomes gains, but chromosome losses and recurrent monosomies can also be found in cancer. We summarize the current models of chromosome loss and our understanding of its consequences, particularly in comparison to chromosome gains.

Prenatal diagnosis of a novel 7q31.31q31.33 microduplication with a favorable outcome

BackgroundCopy number variants (CNVs) are an important source of normal and pathogenic genome variations. Especially CNVs identified in prenatal cases need careful considerations and correct interpretation if those are harmless or harmful variants from the norm.Case presentationHerein, we reported a paternally inherited duplication of 7.6 Mb in 7q31.3 with, surprisingly, a favorable outcome. GTG-banding and CMA on the DNA derived from uncultured amniocytes revealed a karyotype: 46,XX.arr[GRCh37] 7q31.31q31.33(118,601,001_126,177,044) × 3. Ultrasound examination showed no dysmorphisms or intrauterine growth restriction in the fetus and the father was clinically normal as well.ConclusionPrenatal detection of a 7.6 Mb in 7q31.31 to 7q31.33 duplication in a female fetus turned out to be a yet unreported unbalanced chromosome abnormality. This is another example that parental testing and GTG-banding are necessary additional tests to be done in prenatal cases, before a reliable conclusion on the meaning of an aberration can be drawn.

EP112: Septo-optic dysplasia associated with unbalanced chromosome translocation resulting in trisomy 11q13 and monosomy 12q24.3

Distal trisomy 11q syndrome is rare and most often associated with a translocation involving chromosome 22 (Emanuel syndrome). Patients with isolated distal trisomy 11q present with intellectual disability, short stature and microcephaly. Birth defects commonly reported with the syndrome include brain malformations, cardiac defects, diaphragmatic hernia, hip dysplasia, and micrognathia or Pierre-Robin sequence. Distal 12q deletion syndrome is even more rare, and associated with developmental delay and intellectual disability.

MACS-annexin V cell sorting of semen samples with high TUNEL values decreases the concentration of cells with abnormal chromosomal content: a pilot study.

We question whether, in men with an abnormal rate of sperm DNA fragmentation, the magnetic-activated cell sorting (MACS) could select spermatozoa with lower rates of DNA fragmentation as well as spermatozoa with unbalanced chromosome content. Cryopreserved spermatozoa from six males were separated into nonapoptotic and apoptotic populations. We determined the percentages of spermatozoa with (i) externalization of phosphatidylserine (EPS) by annexin V-Fluorescein isothiocyanate (FITC) labeling, (ii) DNA fragmentation by TdT-mediated-dUTP nick-end labeling (TUNEL), and (iii) numerical abnormalities for chromosomes X, Y, 13, 18, and 21 by fluorescence in situ hybridization (FISH), on the whole ejaculate and selected spermatozoa in the same patient. Compared to the nonapoptotic fraction, the apoptotic fraction statistically showed a higher number of spermatozoa with EPS, with DNA fragmentation, and with numerical chromosomal abnormalities. Compared to the whole ejaculate, we found a significant decrease in the percentage of spermatozoa with EPS and decrease tendencies of the DNA fragmentation rate and the sum of disomy levels in the nonapoptotic fraction. Conversely, we observed statistically significant higher rates of these three parameters in the apoptotic fraction. MACS may help to select spermatozoa with lower rates of DNA fragmentation and unbalanced chromosome content in men with abnormal rates of sperm DNA fragmentation.

Detection of aneuploidy using multiplex ligation–dependent probe amplification in fetal tissues from aborted pregnancies

Purpose: About 10-15% of all pregnancies terminate as spontaneous miscarriages. In the first trimester, ≈50% of spontaneous miscarriages are the result of chromosomal aberrations, mostly chromosomal aneuploidies. Cytogenetic analyses are used to confirm aneuploidy in failed pregnancies. Culture failure or poor-quality chromosomes are often problems in those cases. In such situations, methods that are independent of tissue culture are used, and we employed multiplex ligation-dependent probe amplification (MLPA). We determined if MLPA is an appropriate and compatible method compared with classical cytogenetic analyses on fetal tissues.
 Methods: All fetal samples received from spontaneous abortions were cultured, karyotyped (if possible) and genomic DNA extracted. MLPA analyses were undertaken using subtelomeric probe kits. Additionally, comparative genomic hybridization (CGH) was used to confirm aneuploidy detected by MLPA in cases of failed culture growth.
 Results: MLPA analyses confirmed an unbalanced chromosome abnormality identified by cytogenetic analyses in all cases in which tissue culture was successful, and provided data in cases of failed culture growth. Several common numeric chromosome aberrations were detected, as well as rare trisomies and other unbalanced chromosome rearrangements.
 Conclusions: MLPA analyses can provide information about the karyotype of a DNA sample if cytogenetic analyses are not possible because of a lack of viable cells or if only a small amount of genomic DNA is available. Th

A Case-Control Study on Chromosomal Anomalies in Parents Experiencing Repeated Spontaneous Abortions From Northern India

ObjectivesMany women lose their fetuses through miscarriage due to a variety of causes. The incidence of three or more consecutive pregnancy losses is often classified as repeated spontaneous abortion (RSA) and is considered the most frustrating and complex area in reproductive medicine. Parental chromosomal abnormalities, underlying medical condition, heritable or acquired thrombophilias, immunologic abnormalities, infections, and environmental factors are reported to be possible etiologies responsible for RSA. Gametes with unbalanced chromosomes, which are formed when abnormalities exist in parent chromosomes, are one such cause and are responsible for about 50-60% of first-trimester pregnancy loss. This paper aims to identify whether there is an association between chromosomal anomalies in parents and RSA.MethodA case-control study was performed on a total sample size of 600 individuals, including 150 couples with a history of RSA and 150 fertile couples as control. The participants were cytogenetically analyzed using G-banding. Associations between variables were tested using Chi-square and Fisher’s exact test (a p-value<0.05 was considered significant). Informed consent from participants and institutional ethical clearance was obtained before the research began.ResultsChromosomal anomalies were detected in 21 individuals (7%) with a history of RSA. Female preponderance was observed with a female to male ratio of 2.5:1. Structural chromosomal aberrations (SCAs) were detected in 17 patients, with nine (53%) cases showing balanced reciprocal translocation (involving chromosomes 1,3,6,8,12,13,15,16,18,22 and X) and three (17.65%) cases of Robertsonian translocation (exclusively in males). Mosaicism was observed in four (19.05%) cases. A statistically significant positive association (p-value <0.05) was observed between the presence of parental chromosomal anomalies and RSA.ConclusionThese results support an association between RSA and parental chromosomal abnormalities. Currently, clinicians treating cases of RSA face challenging clinical conditions. Identifying a cytogenetic cause for RSA may be of great help to clinicians who manage affected couples.

Abdominal ectopia cordis in an aborted calf without chromosomal aberrations

Ectopia cordis is a rare congenital heart disease characterized by partial or complete displacement of the heart out of the thoracic cavity. Apart from cattle, the condition has also been described in humans and is frequently associated with Cantrell’s pentalogy. It is classified into five types: cervical, cervicothoracic, thoracic, abdominal and thoracoabdominal. The prognosis is poor and ectopia cordis may be linked to the presence of unbalanced chromosome alterations. In this report, a case of abdominal ectopia cordis is described in an aborted calf, in which no unbalanced structural chromosomal variants could be identified.

Novel TNC-PDGFD fusion in fibrosarcomatous dermatofibrosarcoma protuberans: a case report

BackgroundDermatofibrosarcoma protuberans (DFSP) is a superficial fibroblastic tumor characterized by high rate of local recurrence and low metastatic potential. Fibrosarcomatous transformation can rarely arise in DFSP either de novo or as recurrent, which represents a form of tumor progression and carries an increased risk of metastasis over classic DFSP. Cytogenetically, DFSP is characterized by a recurrent unbalanced chromosome translocation t (17;22)(q22;q13), leading to the formation of COL1A1-PDGFB fusion transcript that is present in more than 90% of cases. Alternative fusions involving the PDGFD with partners of COL6A3 or EMILIN2 have recently been documented in less than 2% of cases. Herein, we report a DFSP with fibrosarcomtous morphology harboring a novel TNC-PDGFD fusion.Case presentationA 54-year-old female presented with a slowly growing mass in the right thigh. Excision demonstrated a 2-cm ovoid, well-circumscribed, gray-white, mass. Microscopic examination revealed a partially encapsulated subcutaneous nodule without dermal connection. The neoplasm was composed of cellular and fairly uniform spindle cells with brisk mitoses, arranged in elongated fascicles and herringbone patterns, with focal collagenized stroma. The neoplastic cells were positive for CD34 and smooth muscle actin. Fluorescence in-situ hybridization analyses showed negative for COL1A1-PDGFB fusion as well as NTRK1/2/3 rearrangements. A subsequent RNA sequencing detected an in-frame fusion between exon 15 of TNC and exon 6 of PDGFD. This fusion was further confirmed by nested reverse transcription polymerase chain reaction amplification followed by Sanger sequencing. A diagnosis of fibrosarcomatous DFSP was rendered and the patient was in good status at a follow-up of 12 months after the operation.ConclusionsWe report a fibrosarcomatous DFSP with novel TNC-PDGFD fusion, which adds to the pathologic and genetic spectrum of PDGFD-rearranged DFSP.

Loss of obligate crossovers, defective cytokinesis and male sterility in barley caused by short-term heat stress.

Short-term heat stress during male meiosis causes defects in crossover formation, meiotic progression and cell wall formation in the monocot barley, ultimately leading to pollen abortion. High temperature conditions cause a reduction of fertility due to alterations in meiotic processes and gametogenesis. The male gametophyte development has been shown to be particularly sensitive to heat stress, and even short-term and modest temperature shifts cause alterations in crossover formation. In line with previous reports, we observed that male meiosis in the monocot barley exposed for 24-45h to heat stress (32-42°C) partially or completely eliminates obligate crossover formation and causes unbalanced chromosome segregation and meiotic abortion. Depending on the severity of heat stress, the structure and organization of the chromosomes were altered. In addition to alterations in chromosome structure and dynamics, heat treatment abolished or reduced the formation of a callose wall surrounding the meiocytes and interrupted the cell cycle progression leading to cytokinesis defects and microspore cell death.

Detection of cytogenomic abnormalities by OncoScan microarray assay for products of conception from formalin-fixed paraffin-embedded and fresh fetal tissues

BackgroundThe OncoScan microarray assay (OMA) using highly multiplexed molecular inversion probes for single nucleotide polymorphism (SNP) loci enabled the detection of cytogenomic abnormalities of chromosomal imbalances and pathogenic copy number variants (pCNV). The small size of molecular inversion probes is optimal for SNP genotyping of fragmented DNA from fixed tissues. This retrospective study evaluated the clinical utility of OMA as a uniform platform to detect cytogenomic abnormalities for pregnancy loss from fresh and fixed tissues of products of conception (POC).ResultsFresh specimens of POC were routinely subjected to cell culture and then analyzed by karyotyping. POC specimens with a normal karyotype (NK) or culture failure (CF) and from formalin-fixed paraffin-embedded (FFPE) tissues were subjected to DNA extraction for OMA. The abnormality detection rate (ADR) by OMA on 94 cases of POC-NK, 38 cases of POC-CF, and 35 cases of POC-FFPE tissues were 2% (2/94), 26% (10/38), and 57% (20/35), respectively. The detected cytogenomic abnormalities of aneuploidies, triploidies and pCNV accounted for 50%, 40% and 10% in POC-CF and 85%, 10% and 5% in POC-FFPE, respectively. False negative result from cultured maternal cells and maternal cell contamination were each detected in one case. OMA on two cases with unbalanced structural chromosome abnormalities further defined genomic imbalances and breakpoints.ConclusionOMA on POC-CF and POC-FFPE showed a high diagnostic yield of cytogenomic abnormalities. This approach circumvented the obstacles of CF from fresh specimens and fragmented DNA from fixed tissues and provided a reliable and effective platform for detecting cytogenomic abnormalities and monitoring true fetal result from maternal cell contamination.

UV-induced DNA damage disrupts the coordination between replication initiation, elongation and completion.

Replication initiation, elongation and completion are tightly coordinated to ensure that all sequences replicate precisely once each generation. UV-induced DNA damage disrupts replication and delays elongation, which may compromise this coordination leading to genome instability and cell death. Here, we profiled the Escherichia coli genome as it recovers from UV irradiation to determine how these replicational processes respond. We show that oriC initiations continue to occur, leading to copy number enrichments in this region. At late times, the combination of new oriC initiations and delayed elongating forks converging in the terminus appear to stress or impair the completion reaction, leading to a transient over-replication in this region of the chromosome. In mutants impaired for restoring elongation, including recA, recF and uvrA, the genome degrades or remains static, suggesting that cell death occurs early after replication is disrupted, leaving partially duplicated genomes. In mutants impaired for completing replication, including recBC, sbcCD xonA and recG, the recovery of elongation and initiation leads to a bottleneck, where the nonterminus region of the genome is amplified and accumulates, indicating that a delayed cell death occurs in these mutants, likely resulting from mis-segregation of unbalanced or unresolved chromosomes when cells divide.

Characterization of Chromosomal Abnormalities in Acute Myeloid Leukaemia Patients at the University Teaching Hospital, Lusaka, Zambia

Introduction: Acute myeloid leukaemia (AML) is a highly malignant clonal hematopoietic disease caused by both inherited and acquired genetic alterations (Song et al, 1999). Current AML classification and prognostic systems incorporate genetic information but are limited to known abnormalities that have previously been identified with the use of cytogenetics, array comparative genomic hybridization (CGH), gene-expression profiling, and the resequencing of candidate genes. At diagnosis, most patients with AML harbour at least 1 chromosome aberration in their marrow blasts. Numerous recurrent structural and numeric cytogenetic aberrations have been identified and many of them not only are diagnostic markers for specific AML subtypes but also constitute independent prognostic factors for attainment of complete remission (CR), relapse risk, and overall survival (OS) (Mro´zeket al, 2007). With the targeted cytogenetic therapy, 30% of the patients achieve long-term cure. At University Teaching Hospital(UTH) however, the current diagnostic approach of acute leukaemia involves mainly cytomorphology and occasional flow cytometry. The cytomorphological blast characterization is not enough to provide a critical determination of prognosis and developing a treatment plan. Most of the AML patients at the UTH die within few months after diagnosis despite being put on chemotherapy. Cytogenetic analysis is not done despite the cytogenetic abnormalities being the major predictors of favourable, intermediate or adverse prognosis. Aim: To characterize acute myeloid leukaemia (AML) according to WHO 2008 revised classification in patients at the University Teaching Hospital. Design and Methods: This was a descriptive cross-sectional study conducted to characterize acute myeloid leukaemia (AML) according to WHO 2008 revised classification in patients at the UTH. Patients with AML were simultaneously analyzed for the presence of 4 genetic abnormalities, PML/RARα for t(15;17), AML1/ETO for t(8;21), CBFβ/MYH11 for inv(16)/t(16;16) and rearrangements of the MLL gene for 11q23 abnormalities. AML was classified using the new World Health Organization (WHO) classification for haematologic malignancies. The techniques used were standardized according to the recommendations of the European BIOMED-1 Concerted Action. Results: The overall frequency of leukemia displaying one of the four recurrent cytogenetic translocations were 13 cases (46.5%) of which PML/RARα transcript was present in six(6) patients (21.4%) (3 were bcr1, 1 bcr2 and 2 bcr3). The AML1/ETO fusion transcript was detected in only one(1) case (3.6%) with M2 morphology, but other cases with M2 morphology were negative. CBFβ/MYH11 transcript was present in 2 cases (7.1%) and some of them displaying M4Eo morphology. Finally, 4 cases (14.3%) showed rearrangements of the MLL gene. By contrast, the frequency of AML not otherwise characterized which was 15 cases (53.6%) increased with age (13% for 6-35years age group, 20% for 36-65years age group and 67% for above 66years age group). Our results differ from those reported from the United States and North/Central Europe, particularly regarding the incidence of t(15;17) and t(8;21) translocations. In Zambia the frequency of t(15;17) is higher while that of t(8;21) is lower. This supports the view that geographic variations in tumor-associated aberrations in hematologic malignancies exist. Conclusions: Our study showed that chromosomal alteration PML/RAR t(15,17) which was 21.4% ,was the commonest, whereas AML1/ETO t(8,21) which was 3.6%,was the least common among patients presenting at UTH, Lusaka, Zambia. Our study showed that chromosomal aberration detected in our patients make them less responsive to cytotoxic drugs. The use of molecular technique at point of diagnosis would assist in identifying AML with better prognosis by administering appropriate treatment. The results support the existence of chromosomal abnormalities of AML in our Zambian patients. Awareness of these chromosomal abnormalities and morphology could contribute to the design of cost-effective screening strategies, adapted by our National Health systems according to the prevalence of locally detected genetic aberrations. Acquired genetic alterations such as balanced and unbalanced chromosome aberrations and submicroscopic gene mutations and changes in gene expression strongly affect pre-treatment features and prognosis of patients with acute myeloid leukemia (AML).

Three Offspring with Cri-du-Chat Syndrome from Phenotypically Normal Parents

Cri-du-chat syndrome is characterized by facial dysmorphism, intellectual disability, and multiple congenital anomalies. Most cases occur de novo. Here, we report 3 siblings with cri-du-chat syndrome born to healthy parents. The proband was admitted to our clinic at the age of 6.5 years due to severe intellectual disability, facial dysmorphism, and heart defect. His karyotype showed a deletion of chromosome 5p. Microarray analysis revealed a 29-Mb deletion in chromosome 5p and a 4.7-Mb duplication in chromosome 19q. FISH analysis indicated an unbalanced translocation between 5p13.3 and 19q13.4. During follow-up, the second and the third child of the family were born with the same chromosome abnormality. Parental peripheral blood and skin fibroblast karyotypes as well as the FISH results using chromosome 5p- and 19q-specific subtelomeric probes were normal. FISH analysis of the father's sperm detected a 5p deletion in 12.8% of 200 cells, and microarray analysis confirmed the same unbalanced chromosome abnormality in a mosaic pattern. Uncultured peripheral blood and buccal smear of the father were also studied by FISH to exclude low-level mosaicism and in vitro culture effect. This is the first study that provides molecular evidence of paternal gonadal mosaicism of an unbalanced translocation detected in 3 siblings with cri-du-chat syndrome.

Characterization of a Rare Mosaicism in Autosomal Translocation of t(5;21) Using Conventional Cytogenetics and FISH Methods

Background:Mosaicism of a normal cell population and an unbalanced autosomal chromosome rearrangement is rarely seen. If the abnormal cell line contributes to a minor part of soma, the phenotype is expected to be normal. Case Report:We report a 29-year-old woman who had balance chromosomal translocation of 46,XX,t(5;21) with a two-year-old affected girl, characterized by mental retardation, dystrophia, hearing impartment, and dysphagia. Methods and Results:Cytogenetic investigation revealed a low mosaic unbalanced translocation of 46,XX,t(5;21)/ 46,XX, which was confirmed by FISH analysis. Studying 200 metaphases and interphases of peripheral blood sample revealed 70% partial monosomy of 21q22 and partial trisomy of 5q(35.3) and 30% of normal pattern. Conclusion:In rare cases such as this study, parents with balanced translocation with no phenotypes may lead to a mosaic unbalanced translocation with abnormal phenotypes in offspring, which underscores the need for prenatal karyotyping and genetics counseling.

Pericentric inversion (Inv) 9 variant\u2014reproductive risk factor or benign finding?

PurposeTo report the unbalanced chromosome rearrangement rate and overall aneuploidy rate in day 5/6 embryos from a series of patients who underwent in vitro fertilization (IVF) with preimplantation genetic testing for structural rearrangements (PGT-SR) for the pericentric inversion 9 variant, inv(9)(p11q13) or inv(9)(p12q13), with concurrent 24 chromosome preimplantation genetic testing for aneuploidy (PGT-A).MethodsThis was a retrospective cohort analysis. IVF cycles and embryo biopsies were performed by referring clinics. Fifty-two trophectoderm biopsy samples from seven couples were sent to a single lab for PGT-SR for an inversion 9 variant with concurrent 24 chromosome PGT-A using single-nucleotide polymorphism (SNP) microarrays with bioinformatics.ResultsThe unbalanced rearrangement rate for this embryo cohort was 0/52 (0.0%); mean maternal age per embryo was 33.3 years (range 21–39 years). The overall euploid rate was 61.5% and aneuploidy rate was 38.5%.ConclusionsChromosome 9 pericentric inversions did not result in unbalanced structural rearrangements in day 5/6 embryo samples, supporting that this population variant is not associated with increased reproductive risks.

Selective autophagy maintains centrosome integrity and accurate mitosis by turnover of centriolar satellites

The centrosome is the master orchestrator of mitotic spindle formation and chromosome segregation in animal cells. Centrosome abnormalities are frequently observed in cancer, but little is known of their origin and about pathways affecting centrosome homeostasis. Here we show that autophagy preserves centrosome organization and stability through selective turnover of centriolar satellite components, a process we termed doryphagy. Autophagy targets the satellite organizer PCM1 by interacting with GABARAPs via a C-terminal LIR motif. Accordingly, autophagy deficiency results in accumulation of large abnormal centriolar satellites and a resultant dysregulation of centrosome composition. These alterations have critical impact on centrosome stability and lead to mitotic centrosome fragmentation and unbalanced chromosome segregation. Our findings identify doryphagy as an important centrosome-regulating pathway and bring mechanistic insights to the link between autophagy dysfunction and chromosomal instability. In addition, we highlight the vital role of centriolar satellites in maintaining centrosome integrity.

Breakpoint mapping uncovering about 1.32% of apparent balanced reciprocal translocation (ABRT) carriers existing cryptic complex chromosomal structural variations in preimplantation genetic testing (PGT)

To identify precise breakpoints, evaluate the reproduction-related risks and guide the following PGT treatment, high resolution breakpoint mapping was performed in ABRT carriers indicated by G-banding. A single-center, descriptive research. A large sample of 833 cases with ABRT who planned to accept PGT treatment were recruited in this study. For these patients, the approach of the next-generation sequencing following microdissection (MiroSeq) of the junction region in the derivative chromosomes, and linkage analysis of the adjacent single nucleotide polymorphisms (SNPs) were performed to distinguish the carriers from noncarriers in balanced embryos. For some cases with unbalanced chromosome rearrangement in the breakpoint region, SNP-array and fluorescence in situ hybridization (FISH) techniques were further used to determine the accurate karyotype. In the 833 cases with ABRT, we found 11 cases (1.32%) carried cryptic complex chromosomal structural variation, including 3 unbalanced chromosome rearrangements and 8 balanced ones in which 2 cases carried both inversion and translocation. In these 11 cases, 5 cases related to 3 chromosomes with 4 to 21 breakpoints and 6 cases involved 2 chromosomes with 3 to 6 breakpoints. It is noteworthy that there were two cases exhibited rare chromoanagenesis, including chromothripsis and chromoplexy. Fortunately, two couples have been both successfully transplanted a normal embryo and given birth to a healthy child, and the remaining nine cases are undergoing PGT treatment. In this large-scale analysis of ABRT, high resolution breakpoint mapping precisely characterized these breakpoints and uncovered 1.32% of the ABRT carriers existed cryptic complex chromosomal structural rearrangement. These data suggests that high resolution breakpoint mapping used in PGT can improve the accuracy of evaluating the reproduction-related risks and avoid genetic risks for the ABRT carriers.

Decrease of spermatozoa with an unbalanced chromosome content after cell sorting in men carrying a structural chromosomal abnormality.

We showed that in men with a constitutional chromosomal abnormality, DNA fragmentation was significantly higher in chromosomally unbalanced spermatozoa than in spermatozoa with a normal or balanced chromosomal content. These results could be explained by a phenomenon already described in infertile men: abortive apoptosis. To determine whether magnetic-activated cell separation could select spermatozoa with lower levels of DNA fragmentation and unbalanced chromosome content in men carrying a structural chromosomal abnormality. The spermatozoa of ten males with a chromosomal rearrangement were separated into two populations using magnetic-activated cell separation (annexin V (-) and annexin V (+) fractions), in order to study meiotic segregation by fluorescence insitu hybridization, the percentage of spermatozoa with an externalization of phosphatidylserine by annexin V staining and DNA fragmentation by TdT-mediated dUTP nick-end labeling on the whole ejaculate and on selected spermatozoa in the same patient. For all patients, the percentage of spermatozoa with externalization of phosphatidylserine decreased in the annexin V (-) fraction and increased in the annexin V (+) fraction as compared to the frozen-thawed semen sample. The rates of DNA fragmentation were statistically much lower in the annexin V (-) fraction when compared to the rate before magnetic-activated cell separation for all but one patient. Conversely, we observed a statistically significantly higher rate of DNA fragmentation in the annexin V (+) fraction for six patients. After magnetic-activated cell separation, there was a significant increase of normal/balanced spermatozoa in the fraction of annexin V (-) for all patients. Conversely, we observed a significant decrease in the fraction of annexin V (+) for seven patients. Magnetic-activated cell separation is a promising tool for increasing the selection of healthy spermatozoa, with a decrease in the number of spermatozoa with externalization of phosphatidylserine, DNA fragmentation, and chromosome unbalance, for use in assisted reproductive technologies such as intracytoplasmic sperm injection for males with a chromosomal structural abnormality.

Hypo-osmotic-like stress underlies general cellular defects of aneuploidy.

Aneuploidy, referring to unbalanced chromosome numbers, represents a class of genetic variation associated with cancer, birth defects and eukaryotic microbes1–4. Whereas it is known that each aneuploid chromosome stoichiometry can give rise to a distinct pattern of gene expression and phenotypic profile4,5, it has remained a fundamental question as to whether there are common cellular defects associated with aneuploidy. In this study, we designed a unique strategy that allowed for the observation of common transcriptome changes of aneuploidy by averaging out karyotype-specific dosage effects using aneuploid yeast cell populations with random and diverse chromosome stoichiometry. This analysis uncovered a common aneuploidy gene-expression (CAGE) signature suggestive of hypo-osmotic stress. Consistently, aneuploid yeast exhibited increased plasma membrane (PM) stress leading to impaired endocytosis, and this defect was also observed in aneuploid human cells. Thermodynamic modeling showed that hypo-osmotic-like stress is a general outcome of proteome imbalance caused by aneuploidy and predicted a ploidy-cell size relationship observed in yeast and aneuploid cancer cells. A genome-wide screen further uncovered a general dependency of aneuploid cells on a pathway of ubiquitin-mediated endocytic recycling of nutrient transporters. Loss of this pathway coupled with the aneuploidy-inherent endocytic defect leads to marked alteration of intracellular nutrient homeostasis.

Complex karyotype in de novo acute myeloid leukemia: typical and atypical subtypes differ molecularly and clinically.

Complex karyotype (CK) with ≥3 abnormalities is detected in 10-12% of patients with acute myeloid leukemia (AML) and associated with poor prognosis. The most common unbalanced abnormalities found in CK result in loss of material from the 5q, 7q and/or 17p chromosome arms. The presence of 5q, 7q and/or 17p abnormalities denotes typical CK and their absence denotes atypical CK. Since molecular features of CK-AML are not well-characterized, we investigated mutational status of 81 leukemia/cancer-associated genes in 160 clinically well-characterized patients. They included 136 patients with ≥3 exclusively unbalanced chromosome abnormalities, 96 of whom had a typical CK and 40 atypical CK, and 24 patients with ≥1 balanced abnormality in addition to ≥2 unbalanced ones. Patients with atypical CK-AML differed from those with typical CK-AML: they carried TP53 mutations less often (P<0.001) and more often PHF6 (P=0.008), FLT3-TKD (P=0.02), MED12 (P=0.02) and NPM1 (P=0.02) mutations. They were younger (P=0.007), had higher WBC (P=0.001) and percentages of marrow (P<0.001) and blood (P=0.006) blasts, higher complete remission rates (P=0.02) and longer overall survival (P<0.001), thus indicating that atypical and typical CK-AMLs constitute distinct disease subtypes. We also identified smaller patient subsets within both typical and atypical CK-AML that differed molecularly and clinically.