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Patients with advanced high-grade serous ovarian cancer (HGSOC) are usually treated with paclitaxel and carboplatin; however, predictive markers for this drug combination are unknown. Tumor samples from 71 consecutive HGSOC patients, who received neoadjuvant chemotherapy with paclitaxel and carboplatin, were subjected to molecular analysis. BRCA1/2 germline mutation carriers (n = 22) had longer treatment-free interval (TFI) than non-carriers (n = 49) (9.5 months vs. 3.8 months; P = 0.007). Fifty-one HGSOCs had sufficient quality of tumor DNA for the next-generation sequencing (NGS) analysis by the SeqCap EZ CNV/LOH Backbone Design panel. All 13 tumors obtained from BRCA1/2 germline mutation carriers and 12 sporadic HGSOCs showed a high number of evenly spread chromosomal breaks, which was defined as a BRCAness phenotype; median TFI for this combined group approached 9.5 months. The remaining 26 HGSOCs had similarly high global LOH score (above 20%); however, in contrast to BRCAness tumors, LOH involved large chromosomal segments; these patients had significantly lower TFI (3.7 months; P = 0.006). All patients with CCNE1 amplification (n = 7), TP53 R175H substitution (n = 6), and RB1 mutation (n = 4) had poor response to paclitaxel plus carboplatin. This study describes a cost-efficient method of detecting the BRCAness phenotype, which is compatible with the laboratory-scale NGS equipment. Some molecular predictors allow the identification of potential non-responders to paclitaxel plus carboplatin, who may need to be considered for other treatment options.

The highly successful career of Stephen T. Warren was very well described in the PNAS Profile Series in 2015 (1). We write this remembrance of Steve’s life on behalf of his many friends and scientific colleagues. His death on June 6, 2021 came far too soon and he is sorely missed. Stephen T. Warren. Image credit: Karen Warren (photographer). Steve’s attraction to the field of genetics and his early scientific development came through his work with James Higgins at Michigan State University. It was a time when human genetics was transitioning from basic science to medical utility, with the application of laboratory testing to individual patients serving as the primary stimulus. Tay Sachs carrier screening was developed in Detroit out of the Henry Ford Hospital with Steve’s involvement even as an undergraduate. He recognized the importance of genetic testing to families through his early experience and elected to direct his career to original discovery, where he was very successful. Steve’s interest in syndromes with chromosomal breaks directed his initiative with two very different discovery outcomes. The first disorder he studied was a rare multiple chromosomal breakage disorder, Bloom syndrome. This research, as a graduate student with James Trosko at Michigan State University, seasoned him in somatic genetic methods. Bloom syndrome was eventually found to be caused by mutations in a single gene, a RecQ helicase, causing additional mutations at many sites, often involving chromosome breaks. This rare recessive disease was found to be caused by homozygous loss-of-function of this gene critical to DNA repair. During his postdoctoral training, Steve’s focus turned to a second chromosomal breakage syndrome, at the time termed marker X (fragile X) syndrome. The chromosome break could be elicited in lymphocytes … [↵][1]1To whom correspondence may be addressed. Email: nelson{at}bcm.edu. [1]: #xref-corresp-1-1

Genotoxic impurities can be described as impurities that can induce genetic mutations and chromosomal breaks, or that damage the genetic information within a cell, which may lead to cancer. The European Medical Agency (EMA) and the United States Food and Drug Administration (US FDA) have set a threshold of toxicological concern (TTC) of genotoxic impurities 1.5 μg per day. In a continuous effort to develop an analytical method for the estimation of genotoxic impurities in quetiapine fumarate, a sensitive, simple, and precise high-performance thin-layer chromatography method has been developed and validated for the determination of 2-nitrophenyl( phenyl)sulfane as a genotoxic impurity at trace levels. The limits of detection (LOD) for quetiapine fumarate and 2-nitrophenyl( phenyl)sulfane were found to be 5.11 and 15.5 ng per band, whereas the limits of quantification (LOQ) were observed 0.09 and 0.3 ng per band, respectively. The calibration curve for 2-nitrophenyl( phenyl)sulfane was linear over the concentration range of 10 to 50 ng per band. The method was found to be specific, precise, linear, and accurate for the estimation of 2-nitrophenyl(phenyl)sulfane at trace levels in quetiapine fumarate.

Thiacloprid, a neonicotinoid insecticide, is widely used to control various species of pests in the current agriculture of today. The potential genotoxic effects of thiacloprid on human peripheral blood lymphocytes were investigated in vitro by chromosome aberrations (CA), and double-strand breaks (DSB), which were detected by the phosphorylation of γH2AX histone. Human peripheral blood lymphocytes were exposed to 30, 60, 120, 240, 480 µg.ml−1 doses for the last 24 and 48 hours of culture. Thiacloprid increased CA at the concentrations of 240, 480 μg.ml−1 (P < 0.05), but these results did not confirm genotoxicity. The mitotic index (MI) was important to us; it served as a basis for the confirmation of the cytotoxicity of this insecticide. During 48 hours of culture, at the concentration of 480 µg.ml−1, its value rapidly decreased (0.42) (P < 0.001), which did not allow us to analyse the results because of the high cytotoxic response.

Genetic mutations, chromosomal breaks, and chromosomal rearrangements, which are induced due to organic impurities, are considered as potential genotoxic impurities. The European Medicines Agency (EMA) and the United States Food and Drug Administration (US FDA) have set a threshold of toxicological concern (TTC) of 1.5 µg per person per day for each impurity. A sensitive and simple high-performance thin-layer chromatography (HPTLC) method has been developed and validated for determination of the potential genotoxic impurity, namely, 2-chloroaniline, at trace levels in quetiapine fumarate. The method was found to be specific and selective for the application. The limit of detection (LOD) and limit of quantification (LOQ) for quetiapine fumarate were found to be 1.27 and 3.87 ng per band. The LOD and LOQ values for 2-chloroaniline were found 0.018 and 0.054 ng per band, respectively. The calibration curve for 2-chloroaniline was linear over a concentration range from 2.5 to 12.5 ng. The method was found to be specific, precise, linear, and accurate and can be employed for monitoring and estimation of levels of 2-chloroaniline in quetiapine fumarate.

DNA double‐strand breaks (DSBs) are highly cytotoxic lesions, and unrepaired or misrepaired DSBs can lead to various human diseases, including immunodeficiency, neurological abnormalities, growth retardation, and cancer. Nonhomologous end joining (NHEJ) is the major DSB repair pathway in mammals. Ku70 and Ku80 are DSB sensors that facilitate the recruitment of downstream factors, including protein kinase DNA‐dependent protein kinase, catalytic subunit (DNA‐PKcs), structural components [X‐ray repair cross‐complementing protein 4 (XRCC4), XRCC4‐like factor (XLF), and paralogue of XRCC4 and XLF (PAXX)], and DNA ligase IV (LIG4), which complete DNA repair. DSBs also trigger the activation of the DNA damage response pathway, in which protein kinase ataxia‐telangiectasia mutated (ATM) phosphorylates multiple substrates, including histone H2AX. Traditionally, research on NHEJ factors was performed using in vivo mouse models and murine cells. However, the current knowledge of the genetic interactions between NHEJ factors in human cells is incomplete. Here, we obtained genetically modified human HAP1 cell lines, which lacked one or two NHEJ factors, including LIG4, XRCC4, XLF, PAXX, DNA‐PKcs, DNA‐PKcs/XRCC4, and DNA‐PKcs/PAXX. We examined the genomic instability of HAP1 cells, as well as their sensitivity to DSB‐inducing agents. In addition, we determined the genetic interaction between XRCC4 paralogues (XRCC4, XLF, and PAXX) and DNA‐PKcs. We found that in human cells, XLF, but not PAXX or XRCC4, genetically interacts with DNA‐PKcs. Moreover, ATM possesses overlapping functions with DNA‐PKcs, XLF, and XRCC4, but not with PAXX in response to DSBs. Finally, NHEJ‐deficient HAP1 cells show increased chromosomal and chromatid breaks, when compared to the WT parental control. Overall, we found that HAP1 is a suitable model to study the genetic interactions in human cells. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.

Recently, the concerns about micro- and nano-plastics (NPs) toxicity have been increasing constantly, however the investigations are quiet meager. The present study provides evidences on the toxicological prospectives of virgin-, coronated- and isolated-NPs on human blood cells and Allium cepa root tip, respectively. Several plasma proteins displayed strong affinity towards NPs and produced multi-layered corona of 13 nm to 600 nm size. The coronated-NPs often attracted each other via non-specific protein-protein attraction which subsequently induced protein-induced coalescence in NPs. In the protein point of view, the interaction caused conformational changes and denaturation of protein thereby turned it as bio-incompatible. The coronated-NPs with increased protein confirmation changes caused higher genotoxic and cytotoxic effect in human blood cells than the virgin-NPs. On the other hand, virgin-NPs and the NPs isolated from facial scrubs hindered the root growth and caused chromosome aberration (ring formation, C-mitotic and chromosomal breaks, etc.) in root of Allium cepa. At the outset, the present study highlights the urgent need of scrutinization and regulation of NPs use in medical applications and pre-requisition of additional studies for assessing the bio-accumulation and bio-magnification of NPs.

Miroslav Radman’s far-sighted ideas have penetrated many aspects of our study of the repair of broken eukaryotic chromosomes. For over 35 years my lab has studied different aspects of the repair of chromosomal breaks in the budding yeast, Saccharomyces cerevisiae. From the start, we have made what we thought were novel observations that turned out to have been predicted by Miro’s extraordinary work in the bacterium Escherichia coli and then later in the radiation-resistant Dienococcus radiodurans. In some cases, we have been able to extend some of his ideas a bit further.

The aim of research: Studing cariotype conducted cows of Ukrainian Black-and-White dairy breeds and installed corelation conection between cariotype instability and reproduction ability. Methods of research: Сytogenetic – analysis of metaphase plates, peripheral blood cells cows of Ukrainian Black-and-White dairy breeds, micronucleus test-reaction of the organism on mutagenic factors of different nature, zootechnical – analysis of reproductive properties of animals Ukrainian Black-and-White dairy breeds, statistical. Research result: Cariotype analis of Ukrainian Black-and-White dairy breeds (30 heads) farm husbandry “Progress +” showed the following results: control of spontaneous mytagenesis in cows wich was reserch showing quantitative and structural chromosomal aberation. Quantative chromosomal aberationwas find as aneuploidy. Aneuploidy was equal to 7,4% which does not exceed the limits of spontaneous chromosomal variability(1,5–8,3%), what is normal for animals of milk production breeds. Cellular frequency with asynchronous difference centroid chromosomal areas i population of cows at husbandry “Progress +” was 1,05% and did not exceed the spontaneous cytogenetic variability characteristic for the species Bostaurus. The share of metaphase plates with structural chromosomal abnormalities (chromosomal breaks) 2,6% equal to the level of spontaneous chromosomal variability for cows (0,17–11,1%). Cellular cell with micro nucleus and dual core lymphocytes was within the spontaneous level, characteristic of mammals, which were kept in the absence of direct genotoxic effects and amounted to 3.7‰ and 2.3‰, respectively. To install associate conection between chromosomal abnormalities and the reproductive ability of the studied cows was carried out a correlation analysis. Received coefficient between dual-core lymphocytes and age 1 calving (r = -0,9585) indicates a negative correlation with the strength of communication according to the Student's criteria isreliable at P > 0.99. Conclusions: Quantitative chromosomal aberrations (aneuploidy) were 7.4% which does not exceed the limit of spontaneous chromosomal variability, characteristic for animals breeds of milk production. The frequency of cells with asynchronous difference is established centroid chromosomal areas – 1,05% and a fraction of metaphase plates – 2,6% which corresponds to the level of spontaneous chromosomal variability in cattle. Detected сell frequency with micro nucleus and dual core lymphocytes which were within the spontaneous level, characteristic of mammals which are kept in the absence of direct genotoxic effects and amounted to 3.7‰ and 2.3‰. Was detected a negative correlation relationship between dual-core lymphocytes and age of the 1 calving (r = -0.9585, P > 0.99).

The purpose. To study participation of individual chromosomes in constitutive cytogenetic abnormalities of cattle. Methods. Cultivation of lymphocytes, preparation of cytogenetic specimens. Grading and the record-keeping aberrations of chromosomes was realized under practical standards. Results. The frequency of participation of individual chromosomes in chromosomal rearrangement were studied. It was found that the most frequently happening chromosomal breaks were the following: 20.0% for the first chromosome, 16.0%, for the second one, 8.5% and 7.0% for the thirteenth and the twenty-first chromosomes accordingly. The breaks occurred in places between euchromatin and heterochromatin chromosome segments, as it was displayed by the results of differently colored samples analysis. Conclusions. The gained experimental data testify to the following: frequency of participation of individual chromosomes in constitutive cytogenetic abnormalities can be used as an index of non-specific destabilization of the chromosomal apparatus of animals.

Both copy number variations (CNVs) and chromothripsis are phenomena that involve complex genomic rearrangements. Chromothripsis results in CNVs and other structural changes. CNVs are frequently observed in the human genome. Studies on CNVs have been increasing exponentially; the Database of Genomic Variants shows an increase in the number of data published on structural variations added to the database in the last 15 years. CNVs may be a result of replicative and non-replicative mechanisms, and are hypothesized to serve important roles in human health and disease. Chromothripsis is a phenomena of chromosomal rearrangement following chromosomal breaks at multiple locations and involves impaired DNA repair. In 2011, Stephens et al coined the term chromothripsis for this type of fragmenting event. Several proposed mechanisms have been suggested to underlie chromothripsis, such as p53 inactivation, micronuclei formation, abortive apoptosis and telomere fusions in telomere crisis. Chromothripsis gives rise to normal or abnormal phenotypes. In this review, constitutional chromothripsis, which may coexist with multiple de novo CNVs are described and discussed. This reviews aims to summarize recent advances in our understanding of CNVs and chromothripsis, and describe the effects of these phenomena on human health and birth defects.

To elucidate important cellular and molecular interactions that regulate patterning and skeletal development, vertebrate limbs served as a model organ. A growing body of evidence from detailed studies on a subset of limb regulators like the HOXD cluster or SHH, reveals the importance of enhancers in limb related developmental and disease processes. Exploiting the recent genome-wide availability of functionally confirmed enhancer dataset, this study establishes regulatory interactions for dozens of human limb developmental genes. From these data, it appears that the long-range regulatory interactions are fairly common during limb development. This observation highlights the significance of chromosomal breaks/translocations in human limb deformities. Transcriptional factor (TF) analysis predicts that the differentiation of early nascent limb-bud into future territories entail distinct TF interaction networks. Conclusively, an important motivation for annotating the human limb specific regulatory networks is to pave way for the systematic exploration of their role in disease and evolution.

Background and Aims Acute lymphoblastic leukemia (ALL) is the most common cancer in the pediatric population; ∼80% of the cases show some translocation. Translocations that result in ALL are due to chromosome breaks. However, the exact mechanisms that cause these breaks have not been well studied. A detailed search of the breakpoints associated with ALL reported in the NCBI database shows that some are concentrated in limited regions of the chromosome, whereas others are scattered throughout. Therefore, the objective of this study was to identify the structural factors involved in chromosomal breaks in ALL. Methods We performed several bioinformatic studies on the sequences where chromosomal breakpoints have been reported in search of rearrangements: areas of high similarity, thermodynamic stability, composition and conformation of the DNA. Results Certain factors may influence chromosome breaks and are capable of predicting the propensity towards these types of events. Conclusions These findings may be useful in the design of molecular techniques able to detect these changes in ALL.

The frequency of pathogenic large chromosome rearrangements detected in patients with different Mendelian diseases is truly diverse and can be remarkably high. Chromosome breaks could arise through different known mechanisms. Congenital PAX6-associated aniridia is a hereditary eye disorder caused by mutations or chromosome rearrangements involving the PAX6 gene. In our recent study, we identified 11p13 chromosome deletions in 30 out of 91 probands with congenital aniridia or WAGR syndrome (characterized by Wilms’ tumor, Aniridia, and Genitourinary abnormalities as well as mental Retardation). The loss of heterozygosity analysis (LOH) was performed in 10 families with de novo chromosome deletion in proband. In 7 out of 8 informative families, the analysis revealed that deletions occurred at the paternal allele. If paternal origin is not random, chromosome breaks could arise either (i) during spermiogenesis, which is possible due to specific male chromatin epigenetic program and its vulnerability to the breakage-causing factors, or (ii) in early zygotes at a time when chromosomes transmitted from different parents still carry epigenetic marks of the origin, which is also possible due to diverse and asymmetric epigenetic reprogramming occurring in male and female pronuclei. Some new data is needed to make a well-considered conclusion on the reasons for preferential paternal origin of 11p13 deletions.

Resumen Objetivo: evaluar efecto genotóxico y mutagénico en linfocitos humanos expuestos a nanotubos de carbono (NTC) prístinos y dopados con Nitrógeno. Métodos: linfocitos humanos fueron expuestos a NTC dopados y prístinos (0.08, 0.09, 0.1mg/mL), se evaluó alteraciones cromosómicas e intercambio de cromátidas hermanas (ICH). Resultados: se presentaron rupturas cromatídicas y cromosómicas en linfocitos expuestos a 0.1mg/mL y 0.08mg/mL de NTC prístinos. NTC dopados, indujeron cromosomas dicéntricos y anillos cromosómicos. Se presentó diferencia significativa en el porcentaje de ICH de células tratadas con NTC dopados versus control negativo y NTC prístinos (p

This Expression of Concern relates to the article ‘Chromosomal breaks during mitotic catastrophe trigger γH2AX–ATM–p53-mediated apoptosis’ by Gabriela Imreh, Helin Vakifahmetoglu Norberg, Stefan Imreh and Boris Zhivotovsky. J. Cell Sci. (2011) 124 , [2951-2963][1] ([doi: 10.1242/

This Publisher's Note updates and replaces the Expression of Concern ([doi: 10.1242/jcs.205559][1]) relating to the article ‘Chromosomal breaks during mitotic catastrophe trigger γH2AX–ATM–p53-mediated apoptosis’ by Gabriela Imreh, Helin Vakifahmetoglu Norberg, Stefan Imreh and Boris

Nowadays, it has been appeared that there are several advantages for the medical use of hibiscus, which showed the ability to reduce cholesterol level and lipids in animals at laboratory tests in addition to antibiotic oxidation. Thus, the aim of this research is to study its role as anticlastogenic agent upon the chromosomes damage. The calyx and sub-calyx of the Roselle plant has long been recognized as a source of antioxidants. The objective of this study was to investigate the capability of Hibiscus sabdariffa juice to act as anticlastogenic agent by preventing or decreasing chromosomal breaks. In order to achieve such a purpose the genetic material of Mouse (Mus musculus, 2n = 40) and roottip cells of Onion (Allium cepa, 2n = 16) were selected and used employing a variety of shortterm genotoxic bioassays that recommended by EPAUS.The obtained result revealed that Roselle cold extract or syrup treatment had anticlastogenic effect. While hot extract has not. How does this suggested repair system play its role? by activation of cell proliferation, apoptosis; or by interfering with cellular repair system or by all these assumptions. Further research is needed in order to precisely answer this question.

Nine Fanconi anemia patients complementation group A (FA-A), age 2-6 years, have been infused with autologous hematopoietic cells after genetic correction with the therapeutic PGK-FANCA.Wpre* lentiviral vector. In all instances patients underwent CD34+ cell mobilization with G-CSF and plerixafor and were subsequently infused in the absence of any pre-conditioning regimen, in order to avoid genotoxic side effects in a population characterized by DNA repair defects and cancer predisposition. The first four patients were treated between January 2016 and March 2017 and were infused with an estimated number of 170,000 and 410,000 transduced CD34+ cells/Kg. The other five patients were treated more recently with cell numbers that ranged between 50,000 to 1.6x106 corrected CD34+ cells/kg. The analyses of the first four patients showed the presence of corrected cells both in BM and PB after six months post-infusion and progressive increases of gene marking were observed thereafter in all these patients until the most recent follow-up (2 to >3 years post-infusion). Gene marking in BM CD34+ cells correlated with the survival of the CFCs to mitomycin-C, with levels up to 70% at 3 years post-infusion. Additionally, progressive decreases in the percentage of PB T cells with diepoxybutane-induced chromosomal breaks were observed in the patients with higher levels of gene marking. Similarly, stabilized PB cell counts have been observed in patients with higher percentages of gene corrected cells. Insertion site analyses revealed the absence of genotoxic events, and demonstrated the engraftment of pluripotent HSCs and a pattern of oligoclonal reconstitution, consistent with the number of infused corrected CD34+ cells and the absence of conditioning. In the five additional patients treated more recently, the presence of gene corrected PB cells has been confirmed; levels of gene marking have been consistent with data observed in the first four treated patients and with the number of infused CD34+ cells. Our results confirm the engraftment of gene corrected HSCs in non-conditioned FA-A patients, in some cases through more than 3 years of follow-up, suggesting the relevance of this therapeutic approach in FA. Disclosures Rio: Rocket Pharmaceuticals, Inc.: Equity Ownership, Patents & Royalties: Inventor on patents on lentiviral vectors filled by CIEMAT, CIBERER and F.J.D and may be entitled to receive financial benefits from the licensing of such patents, Research Funding. Navarro:Rocket Pharmaceuticals, Inc.: Patents & Royalties: Inventor on patents on lentiviral vectors filled by CIEMAT, CIBERER and F.J.D and may be entitled to receive financial benefits from the licensing of such patents. Segovia:Rocket Pharmaceuticals, Inc.: Equity Ownership, Honoraria, Patents & Royalties: Inventor on patents on lentiviral vectors filled by CIEMAT, CIBERER and F.J.D and may be entitled to receive financial benefits from the licensing of such patents, Research Funding. Wang:GeneWerk: Employment. Casado:Rocket Pharmaceuticals, Inc.: Patents & Royalties: Inventor on patents on lentiviral vectors filled by CIEMAT, CIBERER and F.J.D and may be entitled to receive financial benefits from the licensing of such patents. Galy:Genethon: Employment. Cavazzana:SmartImmune: Other: Founder. Schwartz:Rocket Pharmaceuticals: Employment, Equity Ownership. Schmidt:GeneWerk GmbH, Heidelberg, Gemrany: Equity Ownership; German Cancer Research Center, Heidelberg, Germany: Employment. Díaz de Heredia:Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Sevilla:Rocket Pharmaceuticals, Inc.: Honoraria, Patents & Royalties: Inventor on patents on lentiviral vectors filled by CIEMAT, CIBERER and F.J.D and may be entitled to receive financial benefits from the licensing of such patents; NOVARTIS: Honoraria, Membership on an entity's Board of Directors or advisory committees; Rocket: Membership on an entity's Board of Directors or advisory committees; Amgen: Honoraria, Membership on an entity's Board of Directors or advisory committees; Sobi: Membership on an entity's Board of Directors or advisory committees; Miltenyi Biotech: Honoraria. Bueren:Rocket Pharmaceuticals, Inc.: Consultancy, Equity Ownership, Patents & Royalties: Inventor on patents on lentiviral vectors filled by CIEMAT, CIBERER and F.J.D and may be entitled to receive financial benefits from the licensing of such patents, Research Funding.

In order to evaluate cytogenetic effect of nanocomposite materials used in traumatology and orthopedics an investigation of chromosomal aberration ins in the culture of lymphocytes short-term contacted with artificial material (nanostructured titanium samples of implants bearing bilateral calcium phosphate coating) in patients with dysplasia of connective tissue (DCT) and osteogenesis imperfecta (OI) has been conducted. A reduction of percent of aberrant cells having single or/and paired chromosomal breaks as compared with spontaneous mutagenesis (without model samples) was revealed under introduced biocompatible composites into blood cell cultures of patients with DCT or OI. Analysis of frequency and spectrum of chromosomal aberrations in blood cells can be considered as potential prognostic criterion and a predictor of implant’s efficacy in the surgical treatment of DCT and OI complications.

While a good safety for recombinant human growth hormone (rhGH) therapy at replacement doses is recognized, a possible link between high concentration of the GH-IGF-I axis hormones and side negative effect has been reported. The aim of this pilot study was to assess whether a short-term exposure to supra-physiological doses of rhGH may affect DNA integrity in human lymphocytes (PBL). Eighteen healthy Caucasian female (24.2 ± 3.5 years) were randomly included in a Control (n = 9) and rhGH administration group (n = 9, 3-week treatment). DNA damage (comet assay), chromosomal breaks, and mitotic index in phytohemagglutinin-stimulated PBL were evaluated before (PRE), immediately (POST), and 30 days (POST30) after the last rhGH administration (0.029 mg kg− 1 BW; 6 days/week), together with serum IGF-1 and IGFBP-3 concentrations. rhGH administration increased IGF-I, without evidence of persisting IGF-I and IGFBP-3 changes 30 days after withdrawal. Total DNA breakage (% DNA in tails) was not significantly different in subjects treated with rhGH in comparison with controls, although the rhGH-treated subjects showed an higher percentage of heavily damaged nuclei immediately after the treatment (POST30 vs. PRE: p = 0.003), with a lower mitogenic potential of lymphocytes, detectable up to the POST30 (PRE vs. POST: p = 0.02; PRE vs. POST30: p = 0.007). This pilot study showed that 3 weeks of short-term supra-physiological rhGH administration in healthy women induce a transient DNA damage and mitogenic impairment in PBL. The analysis of DNA damage should be explored as useful tool in monitoring the mid to long-term effects of high rhGH treatment or abuse.

We have developed an easy, reliable two-step method for the insertion of large DNA fragments into any desired location in the E. coli chromosome. The method is based on the recombineering of a small (∼1.3 kbp) “Landing Pad” into the chromosome at the insertion site, to which the large construct is subsequently delivered via I-SceI endonuclease excision from a donor plasmid. To demonstrate the power of this method, we here show the insertion of a fragment containing the entire lac operon (∼9 kbp) into four predefined novel locations in the E. coli chromosome, a feat not possible with existing technologies. In addition, the chromosomal breaks induced by landing pad excision provide sufficient selective pressure that positive selection by antibiotics is unnecessary, making precise, exact insertion without extraneous sequence possible.

Cytogenetic assays in peripheral blood lymphocytes (PBL) have been done to assess the incidences of chromosomal aberrations among Meos and Sunni Muslims of Haryana. Baseline frequencies of chromosome aberrations (CA) were assessed in 28 subjects from each cast. The mean frequency of chromosomal gaps in Meos was found to be 0.786 ± 0.686 and the frequency of chromosomal break was 0.429 ± 0.504. Mean frequency of the satellite association in Meos was found to be 1.357 ± 1.821. Among the Sunni Muslims the mean frequency of chromosomal gaps was found to be 1.250 ±0.701 and the frequency of chromosomal breaks was 0.643 ± 0.488. Mean frequency of satellite in Sunni Muslims was found to be 1.250 ± 1.435. Statistically non-significant differences were observed for the gaps, break and satellite associations, whereas values for total aberrations were found to be statistically significant among Meos and Sunni Muslims. The mean values for chromosomal aberrations and satellite associations were higher in Sunni Muslims (1.893) than in Meos (1.214).

Three patients with Fanconi's anemia were analyzed for chromosome breaks. T and B cells were separated and grown in tissue culture with PHA and pokeweed antigen to ascertain the rates of breakage in these lymphocytic subpopulations. It has been found that there is no statistically significant difference in breakage rates in T and B lymphocytes. It is postulated that both T and B cells could be involved in the development of leukemia in Fanconi's anemia patients, assuming that chromosome breaks constitute a factor predisposing to the development of malignancy.

The prevalence of squamous cell carcinoma of the skin in albinos reaches approximately 90 % in patients over 20 years of age in the vicinity of Enugu, Nigeria. Chromosome breaks and sister chromatid exchanges (SCE) were evaluated in tyrosinase-positive oculocutaneous albinos and pigmented controls of Ibo extraction who were life-long residents of Nigeria. No difference in the frequency of chromosomal breaks in 14 albinos compared to 6 pigmented controls, and no differences in the frequency of SCE in 9 albinos compared to 3 controls could be detected. Increased chromosomal abnormalities in lymphocytes do not appear to be associated with albinism or fulminating skin cancer present in albinos in the tropics.

We aimed to investigate the association between chronic exposure to benzene and genotoxicity in the lymphocytes of workers removed from exposure. The study included 20 workers with hematological disorders who had previously worked in the petrochemical industry of Salvador, Bahia, Brazil; 16 workers without occupational exposure to benzene served as the control group. Chromosomal analysis was performed on lymphocytes from peripheral blood, to assess chromosomal breaks and gaps and to identify aneuploidy. The Kruskal-Wallis test was used to compare the mean values between two groups, and Student’s t test for comparison of two independent means. The frequency of gaps was statistically higher in and the exposed group than in the controls (2.13 ± 2.86 vs. 0.97 ± 1.27, p = 0.001). The frequency of chromosomal breaks was significantly higher among cases (0.21 ± 0.58) than among controls (0.12 ± 0.4) (p = 0.0002). An association was observed between chromosomal gaps and breaks and occupational exposure to benzene. Our study showed that even when removed from exposure for several years, workers still demonstrated genotoxic damage. Studies are still needed to clarify the long-term genotoxic potential of benzene after removal from exposure.

Chromosomal structural changes can be induced by the addition of specific alien chromosomes called ‘gametocidal (Gc) chromosomes’. In the monosomic addition of the Gc chromosome to common wheat, chromosomal breaks occur in gametes receiving no Gc chromosome, and the broken ends heal and stabilize in the subsequent generations. Thus, by the so-called Gc system, deficiencies and translocations can be induced in common wheat and also in alien chromosome addition and substitution lines of common wheat. Deficiencies of wheat and alien chromosomes were cytologically identified by the chromosome banding and in situ hybridization techniques. The plants carrying deletions or wheat-alien translocations were established as new aneuploid lines of common wheat with sub-arm aneuploidy. Those for wheat chromosomes are called deletion stocks and those for alien chromosomes are called dissection lines. The new aneuploids have been used for cytological chromosome mapping and have corrected some mistakes in genetic mapping.

The diagnosis of Fanconi anemia (FA) is based on the association of congenital malformations, bone marrow failure syndrome, and hypersensitivity to chromosomal breaks induced by cross-linking agents. In the absence of typical features, the diagnosis is not easy to establish because there is no simple and cost-effective test; thus, investigators must rely on specialized analyses of chromosomal breaks. Because we observed elevated serum alpha-fetoprotein (sAFP) levels in FA patients, we investigated this parameter as a possible diagnostic tool. Serum AFP levels from 61 FA patients and 27 controls with acquired aplastic anemia or other inherited bone marrow failure syndromes were analyzed using a fluoroimmunoassay based on the TRACE technology. Serum AFP levels were significantly more elevated (P

Patients with Fanconi's anaemia (FA) present with a wide spectrum of congenital abnormalities and develop hypoplastic anaemia. There is also at least a 10% risk of developing acute myeloid leukaemia (Alter 1987). Phenotypically normal patients have also been described who have only shown an increased number of chromosomal breaks (Auerbach, Rogatko & Schroeder-Kurth, 1989). We describe a 7-year-old girl presenting with myelodysplasia (MDS) in whom a diagnoses of FA was made by chromosome analysis after treatment had commenced.

Chromosomal breaks destabilize the genome and can cause developmental defects and diseases such as cancer. New work suggests that, shortly after DNA damage, dissociation of the histone binding protein HP1β from chromatin facilitates restoration of genome integrity.

Summary About one half of Philadelphia (Ph)-positive acute leukaemia (AL) patients and rare cases of Ph-positive chronic myelogenous leukaemia (CML) show rearrangements within the first intron of the BCR gene on chromosome 22. We studied breakpoints within the first BCR intron in 22 adult patients with Ph-positive leukaemia; 21 with AL and one with CML, which lacked rearrangements within the major bcr (M-bcr). With a series of genomic probes from this intronic region, we detected chromosomal breaks in all 22 patients within the 35 kb area, corresponding almost to the 3’half portion of the intron. The breakpoints were distributed throughout this region but we could not identify any special cluster of breakpoints in this area. Our data support consistent involvement of the 3’half part of the first BCR intron in Ph-positive leukaemias without M-bcr rearrangement, and indicate relatively wide scattering of breakpoints in this portion of the intron.

The DNA helicase RECQL5 has a role in moderating transcription stress levels and consequently in suppressing chromosomal breaks.

Occupational exposure to low doses of ionizing radiation is a particularly delicate subject for investigation, due to the cumulative effects of chronic exposure. It is extremely important to consider and to measure the biological response to given conditions of exposure. The aim of this study was to establish possible recovery from DNA damage in subjects professionally exposed to radiation in their working area by examinations for chromosomal aberrations (CA) at two different times. The first group (I) was composed of 30 professionally exposed subjects in whom unstable CA (dicentrics, ring, acentric fragments, chromatid, chromosomal breaks, and chromatid interchanges) were identified at time zero. After removal from the radiation area, they were re-examined 9 months later. The second group (II) contained 64 healthy individuals, not professionally exposed to ionizing radiation or other known mutagenic agents. In the group of exposed individuals, five (16.67%) subjects exhibited permanent unstable CAs, even after 9 months absence from the radiation. When the nonexposed and exposed groups were compared, an increase of unstable aberrations (p < 0.05) was observed in the exposed group. Nevertheless, a statistically significant decrease of dicentrics, acentric fragments, and ring frequencies was observed in exposed individuals after 9 months away from the radiation area. However, chromatid and isochromatid break frequencies increased slightly but not significantly after 9 months. The detected CAs corresponded to the total effective doses of radiation measured in our subjects. The existence of CAs in some individuals even after absence from the radiation area suggests that the time necessary for the damaged DNA to recover is extremely variable and indicates interindividual differences in radiosensitivity as well as differences in the cellular-reparation response.

DNA double-strand breaks (DSBs) are considered the most important type of DNA damage inflicted by ionizing radiation. The molecular mechanisms of DSB repair by nonhomologous end joining (NHEJ) have not been well studied in live mammalian cells, due in part to the lack of suitable chromosomal repair assays. We previously introduced a novel plasmid-based assay to monitor NHEJ of site-directed chromosomal I-SceI breaks. In the current study, we expanded the analysis of chromosomal NHEJ products in murine fibroblasts to focus on the error-prone rejoining of DSBs with noncomplementary ends, which may serve as a model for radiation damage repair. We found that noncomplementary ends were efficiently repaired using microhomologies of 1-2 nucleotides (nt) present in the single-stranded overhangs, thereby keeping repair-associated end degradation to a minimum (2-3 nt). Microhomology-mediated end joining was disrupted by Wortmannin, a known inhibitor of DNA-PKcs. However, Wortmannin did not significantly impair the proficiency of end joining. In contrast to noncomplementary ends, the rejoining of cohesive ends showed only a minor dependence on microhomologies but produced fivefold larger deletions than the repair of noncomplementary ends. Together, these data suggest the presence of several distinct NHEJ mechanisms in live cells, which are characterized by the degree of sequence deletion and microhomology use. Our NHEJ assay should prove a useful system to further elucidate the genetic determinants and molecular mechanisms of site-directed DSBs in living cells.

The significance in the high incidence of thyroid nodules is that some of these nodules may harbour thyroid tumors. Still, the molecular mechanisms underlying the tumor formation are not fully understood. In this study, the positional cloning approach was used to characterize the chromosomal breakpoints specific for the two main cytogenetic subgroups in benign follicular thyroid tumors which habor chromosomal alterations in 19q13 and 2p21, respectively. In the cytogenetic subgroup with 19q13 aberrations, a candidate gene ZNF331 was identified at a 25 kbp distance proximal to the breakpoint region. ZNF331 consists of 15 exons and encodes for a 463 aa protein with a 42 aa KRAB-A domain and a 332 aa zinc finger region. The normal expression pattern of ZNF331 shows 3 transcripts of 2.1, 4.0 und 4.8 kbp. Exclusively in cell lines established from thyroid adenomas with 19q13 aberration, the expression of a 3.4 kbp transcript was detected. Hidden translocations involving chromosome 19 may mask a higher rate of clonal aberrations in benign thyroid tumors. FISH analyses did not confirm the existence of such translocations in a subset of 38 thyroid adenomas with apparently normal karyotype. The characterization of the canine orthologue of ZNF331 showed a high percentage of sequence identity with its human counterpart at the nucleic acid level as well as at the amino acid level. By FISH, canine ZNF331 was mapped to the terminal region of canine chromosome 1, i.e. CFA1q33.In the cytogenetic subgroup with 2p21 aberrations, a candidate gene, THADA, covering the 450-kbp breakpoint region was identified and first characterized. In two cell lines with 2p21 aberrations, the chromosomal breakpoint was narrowed down to intron 28 of THADA. Analyses of fusion transcripts resulting from the aberrations pointed to truncation of THADA rather than fusion to particular coding sequences as the critical effect of the chromosomal breaks.

Art. original(p.185-191) Submitted by ROBERTO PAULO CORREIA DE ARAÚJO (ppgorgsistem@ufba.br) on 2016-09-13T13:52:10Z No. of bitstreams: 1 R. Ci. méd. biol., v.2, n. 2-2003.pdf: 131643 bytes, checksum: 7d7045e42c32854634911a0c6e540fcc (MD5) Made available in DSpace on 2016-09-13T13:52:10Z (GMT). No. of bitstreams: 1 R. Ci. méd. biol., v.2, n. 2-2003.pdf: 131643 bytes, checksum: 7d7045e42c32854634911a0c6e540fcc (MD5) Previous issue date: 2003-07 O envelhecimento é influenciado por fatores genéticos, nutricionais, exposição a radicais livres, radiação e substâncias genotóxicas. A sensibilidade da célula às substâncias mutagênicas aumenta com a idade e pode contribuir para a aceleração do processo de envelhecimento. Esta investigação analisa o efeito do Metotrexato (MTX), agente indutor de quebras cromossômicas na inibição da reprodução celular em grandes longevos e em um grupo controle. A amostra foi constituída por dez idosos com idades entre 92 e 114 anos e doze controles de 20 a 25 anos. As células para análise foram obtidas por cultura temporária de linfócitos, com adição de MTX. A análise estatística foi realizada com as provas de Kruskal-Wallis e Friedman. O Metotrexato mostrou-se eficaz na redução da divisão celular com resultados estatisticamente significativos. O índice mitótico apresentou valores baixos nos idosos, mesmo sem o uso do MTX, mostrando que o processo de envelhecimento está relacionado com o decréscimo na capacidade de reprodução da célula. Salvador

Discovered three new proteins involved in DNA damage assessment. Interestingly they are all proteins involved in recombination, but they have very different roles in that process and other proteins that might be expected to be equivalently involved are not. This is developing into a very significant area of research.

Several laboratory surveys have shown that transposable elements (TEs) can cause chromosomal breaks and lead to inversions, as in dysgenic crosses involving P –elements. However, it is not presently clear what causes inversions in natural populations of Drosophila . The only direct molecular studies must be taken as evidence against the involvement of mobile elements. Here, in Drosophila lines transformed with the hobo transposable element, and followed for 100 generations, we show the appearance of five different inversions with hobo inserts at breakpoints. Almost all breakpoints occurred in hobo insertion sites detected in previous generations. Therefore, it can be assumed that such elements are responsible for restructuring genomes in natural populations.

The ORMO-48 is a new indigenous material for dental applications, developed by the Dental Products Laboratory of our Institute. The aim of the present study was to evaluate the genotoxic effect of an indigenously developed dental material in Swiss albino mice. The genotoxic effect was evaluated by micronucleus and chromosomal aberration tests. Two grams of dental material was extracted in 10.0 ml of physiological saline at 70°C for 24 h. The extract was cooled to room temperature and was used for the experiment. The experimental designed had three groups each (six mice in each group) for micronucleus and chromosomal aberration tests. The first, second, and third groups were given a single exposure of physiological saline alone (control), dental material's extract (test), and cyclophosphamide (positive control) respectively for micronucleus and chromosomal aberration tests. The result of the study indicated that, the percentage of micronucleated PCE (polychromatic erythrocytes) and NCE (normochromatic erythrocytes) induced by the dental material (extract) treated group was well comparable with control group, whereas the positive control induced significantly high (P < 0.001) micronucleated PCE when compared to control. The PCE and NCE ratio of the dental material extract treated group was similar to that of control group. The chromosomal anomalies such as chromatid/chromosomal breaks, centric rings, exchanges, dicentric, and acentric fragments were evaluated. The result showed that the anomalies of the dental material extract treated group were similar to control group, however, significant anomalies were observed in the cyclophosphamide treated group. Hence, the present study concluded that the indigenously developed biocompatible dental material, ORMO-48 is non genotoxic at our laboratory conditions.

Inhibitory effects of corn oil and its constituents have been studied against 7,12-dimethylbenz[a]anthracene (DMBA)-induced chromosomal breaks in B6C3F1 female mice using the in vivo bone-marrow micronucleus assay. We tested propyl gallate, alpha-tocopherol and beta-sitosterol as constituents of corn oil. In addition, sunflower oil was tested also to check whether corn oil differs from any other vegetable oil. Corn oil, propyl gallate, beta-sitosterol or sunflower oil were injected i.p. to mice for 2 days at 24-h intervals, prior to injecting DMBA i.p. alpha-Tocopherol was mixed in powdered food and the mice were fed on it for 4 days before receiving DMBA. Bone-marrow samples were collected at various 24-h intervals. About 50-70% reduction in number of micronucleated polychromatic erythrocytes (MNPCE)/500 PCEs were observed in all the treatments wherever corn oil was used. Significant inhibitory effects were noted in treatments with alpha-tocopherol and beta-sitosterol. Sunflower oil also showed an inhibitory effect, similar to that with corn oil.