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At least 181 records · Page 10

Evaluation of an automated karyotyping system for chromosome aberration analysis

Chromosome aberration analysis is a promising complement to conventional radiation dosimetry, particularly in the complex radiation fields encountered in the space environment. The capabilities of a recently developed automated karyotyping system were evaluated both to determine current capabilities and limitations and to suggest areas where future development should be emphasized. Cells exposed to radiometric chemicals and to photon and particulate radiation were evaluated by manual inspection and by automated karyotyping. It was demonstrated that the evaluated programs were appropriate for image digitization, storage, and transmission. However, automated and semi-automated scoring techniques must be advanced significantly if in-flight chromosome aberration analysis is to be practical. A degree of artificial intelligence may be necessary to realize this goal.

Prichard, Howard M.↗

Capillary electrophoresis: Biotechnology for separation of DNA and chromosomes

Electrophoresis has been used for the separation of particles, ions, and molecules for a number of years. The technology for separation and detection of the results has many applications in the life sciences. One of the major goals of the scientific community is to separate DNA molecules and intact chromosomes based upon their different lengths or number of base pairs. This may be achieved by using some of the commercially available and widely used methods, but these processes require a considerable amount of time. The challenge is to achieve separation of intact chromosomes in a short time, preferably in a matter of minutes.

Williams, George O., Jr.↗

Automated Chromosome Breakage Assessment

An automated karyotyping machine was built at JPL in 1972. It does computerized karyotyping, but it has some hardware limitations. The image processing hardware that was available at a reasonable price in 1972 was marginal, at best, for this job. In the meantime, NASA has developed an interest in longer term spaceflights and an interest in using chromosome breakage studies as a dosimeter for radiation or perhaps other damage that might occur to the tissues. This uses circulating lymphocytes as a physiological dosimeter looking for chromosome breakage on long-term spaceflights. For that reason, we have reactivated the automated karyotyping work at JPL. An update on that work, and a description of where it appears to be headed is presented.

Castleman, Kenneth↗

Painting Analysis of Chromosome Aberrations Induced by Energetic Heavy Ions in Human Cells

This viewgraph presentation reviews some of the techniques used to analyze the damage done to chromosome from ion radiation. Fluorescence in situ hybridization (FISH), mFISH, mBAND, telomere and centromereprobes have been used to study chromosome aberrations induced in human cells exposed to low-and high-LET radiation in vitro. There is some comparison of the different results from the various techniques. The results of the study are summarized.

Wu, Honglu↗

Distribution of Chromosome Breakpoints in Human Epithelial Cells Exposed to Low- and High-LET Radiation

Low-and high-LET radiations produced distinct breakpoint distributions. The difference of the breakpoint distributions between low-and high-LET only appeared in break ends involved in interchromosome exchanges. The breakpoint distributions for break ends participating in intrachromosome exchanges were similar. Gene-rich regions do not necessarily have more chromosome breaks. High-LET appeared to produce long live (data not shown) or longer live breaks that can migrate a longer distance before rejoining with other breaks. Domains occupied by different segments of the chromosomes may be responsible for the breakpoint distribution. The dose responses for interchromosomal exchanges were linear in all four exposures. However, the dose response for intrachromosomal exchanges were none linear. Increasing dose of high dose rate exposure (Fe-ions or -rays) increase the fraction of cells with intrachromosome aberrations, whereas increasing dose of low dose rate exposure (neutrons or -rays) does not affect the fraction of cells with intrachromosome aberrations.

Hada, Megumi↗

Space Radiation Effects on Human Cells: Modeling DNA Breakage, DNA Damage Foci Distribution, Chromosomal Aberrations and Tissue Effects

Future long-tem space travel will face challenges from radiation concerns as the space environment poses health risk to humans in space from radiations with high biological efficiency and adverse post-flight long-term effects. Solar particles events may dramatically affect the crew performance, while Galactic Cosmic Rays will induce a chronic exposure to high-linear-energy-transfer (LET) particles. These types of radiation, not present on the ground level, can increase the probability of a fatal cancer later in astronaut life. No feasible shielding is possible from radiation in space, especially for the heavy ion component, as suggested solutions will require a dramatic increase in the mass of the mission. Our research group focuses on fundamental research and strategic analysis leading to better shielding design and to better understanding of the biological mechanisms of radiation damage. We present our recent effort to model DNA damage and tissue damage using computational models based on the physics of heavy ion radiation, DNA structure and DNA damage and repair in human cells. Our particular area of expertise include the clustered DNA damage from high-LET radiation, the visualization of DSBs (DNA double strand breaks) via DNA damage foci, image analysis and the statistics of the foci for different experimental situations, chromosomal aberration formation through DSB misrepair, the kinetics of DSB repair leading to a model-derived spectrum of chromosomal aberrations, and, finally, the simulation of human tissue and the pattern of apoptotic cell damage. This compendium of theoretical and experimental data sheds light on the complex nature of radiation interacting with human DNA, cells and tissues, which can lead to mutagenesis and carcinogenesis later in human life after the space mission.

Ponomarev, A. L.↗

A Bi-Exponential Repair Algorithm for Radiation-Induced Double-Strand Breaks: Application to Chromosome Aberrations

Chromosome aberrations (CAs) are one of the effects of radiation exposure and are used as a biomarker. A new simulation program, named RITCARD (Radiation induced tracks, chromosome aberrations, repair, and damage) was developed to simulate radiation-induced CA. RITCARD is used with the program RITRACKS (Relativistic Ion Tracks), which simulates the radiation tracks. The restitution kinetics algorithm presented here is a significant improvement over the one used in the first version. Simulations of radiation-induced CA were performed for several ion types and mixed irradiation fields. These simulations will be useful to help interpreting experiments of galactic cosmic rays (GCR) simulator.

Plante, Ianik↗

Chromosome aberrations among atomic-bomb survivors exposed in utero: updated analysis accounting for revised radiation doses and smoking

A previous study of peripheral blood lymphocyte translocations around age 40 among atomic-bomb survivors exposed in utero revealed no overall association with radiation dose—despite a clear association between translocations and dose among their mothers—but the data suggested an increase at doses below 100 mGy with a definite peak. That analysis of the in utero-exposed survivors did not adjust for their subsequent smoking behavior, an established cause of chromosomal aberrations, or their subsequent exposures to medical irradiation, a potential mediator. In addition, atomic-bomb survivor radiation dose estimates have subsequently been updated and refined. We therefore re-estimated the dose response using the latest DS02R1 dose estimates and adjusting for smoking as well as for city and proximal–distal location at the time of exposure to the atomic bomb. Sex of the survivor, mother’s age around the time of conception, and approximate trimester of gestation at the time of exposure were also considered as explanatory variables and modifiers. Precision of the estimated dose response was slightly lower due to greater variability near zero in the updated dose estimates, but there was little change in evidence of a low-dose increase and still no suggestion of an overall increase across the entire dose range. Adjustment for smoking behavior led to a decline in background number of translocations (the dose–response intercept), but smoking did not interact with dose overall (across the entire dose range). Adjustment for medical irradiation did not alter the association between dose and translocation frequency. Sex, mother’s age, and trimester were not associated with number of translocations, nor did they interact with dose overall. Interactions with dose in the low-dose range could not be evaluated because of numerical instability.

59 BASIC BIOLOGICAL SCIENCES↗

Chromosome-level genome assembly of Quercus variabilis provides insights into the molecular mechanism of cork thickness

Quercus variabilis is a deciduous woody species with high ecological and economic value and is a major source of cork in East Asia. Cork from thick softwood sheets have higher commercial value than those from thin sheets. It is extremely difficult to genetically improve Q. variabilis to produce high quality softwood due to the lack of genomic information. Here, we present a high-quality chromosomal genome assembly for Q. variabilis with length of 791,89 Mb and 54,606 predicted genes. Comparative analysis of protein sequences of Q. variabilis with 11 other species revealed that specific and expanded gene families were significantly enriched in the "fatty acid biosynthesis" pathway in Q. variabilis, which may contribute to the formation of its unique cork. Additionally, based on weighted correlation network analysis of time-course (i.e., five important developmental ages) gene expression data in thick-cork versus thin-cork genotypes of Q. variabilis, we identified one co-expression gene module associated with the thick-cork trait. Within this co-expression gene module, 10 hub genes were associated with suberin biosynthesis. Furthermore, we identified a total of 198 suberin biosynthesis-related new candidate genes that were up-regulated in trees with a thick cork layer relative to those with a thin cork layer. Also, we found that some genes related to cell expansion and cell division were highly expressed in trees with a thick cork layer. Collectively, our results revealed that two metabolic pathways (i.e., suberin biosynthesis, fatty acid biosynthesis), along with other genes involved in cell expansion, cell division, and transcriptional regulation, were associated with the thick-cork trait in Q. variabilis, providing insights into the molecular basis of cork development and knowledge for informing genetic improvement of cork thickness in Q. variabilis and closely related species.

59 BASIC BIOLOGICAL SCIENCES↗

Simultaneous Exposure of Cultured Human Lymphoblastic Cells to Simulated Microgravity and Radiation Increases Chromosome Aberrations

During space travel, humans are continuously exposed to two major environmental stresses, microgravity (μG) and space radiation. One of the fundamental questions is whether the two stressors are interactive. For over half a century, many studies were carried out in space, as well as using devices that simulated μG on the ground to investigate gravity effects on cells and organisms, and we have gained insights into how living organisms respond to μG. However, our knowledge on how to assess and manage human health risks in long-term mission to the Moon or Mars is drastically limited. For example, little information is available on how cells respond to simultaneous exposure to space radiation and μG. In this study, we analyzed the frequencies of chromosome aberrations (CA) in cultured human lymphoblastic TK6 cells exposed to X-ray or carbon ion under the simulated μG conditions. A higher frequency of both simple and complex types of CA were observed in cells exposed to radiation and μG simultaneously compared to CA frequency in cells exposed to radiation only. Our study shows that the dose response data on space radiation obtained at the 1G condition could lead to the underestimation of astronauts’ potential risk for health deterioration, including cancer. This study also emphasizes the importance of obtaining data on the molecular and cellular responses to irradiation under μG conditions.

59 BASIC BIOLOGICAL SCIENCES↗

High-LET radiation-induced aberrations in prematurely condensed G2 chromosomes of human fibroblasts

PURPOSE: To determine the number of initial chromatid breaks induced by low- or high-LET irradiations, and to compare the kinetics of chromatid break rejoining for radiations of different quality. MATERIAL AND METHODS: Exponentially growing human fibroblast cells AG1522 were irradiated with gamma-rays, energetic carbon (290MeV/u), silicon (490MeV/u) and iron (200 and 600 MeV/u). Chromosomes were prematurely condensed using calyculin A. Chromatid breaks and exchanges in G2 cells were scored. PCC were collected after several post-irradiation incubation times, ranging from 5 to 600 min. RESULTS: The kinetics of chromatid break rejoining following low- or high-LET irradiation consisted of two exponential components representing a rapid and a slow time constant. Chromatid breaks decreased rapidly during the first 10min after exposure, then continued to decrease at a slower rate. The rejoining kinetics were similar for exposure to each type of radiation. Chromatid exchanges were also formed quickly. Compared to low-LET radiation, isochromatid breaks were produced more frequently and the proportion of unrejoined breaks was higher for high-LET radiation. CONCLUSIONS: Compared with gamma-rays, isochromatid breaks were observed more frequently in high-LET irradiated samples, suggesting that an increase in isochromatid breaks is a signature of high-LET radiation exposure.

Non-NASA Center↗

Dose reconstruction for individuals exposed to ionizing radiation using chromosome painting

To be most useful, a biomarker for dose reconstruction should employ an end point that is highly quantitative, stable with time and easily measured. Reciprocal translocations have been shown to be a promising biomarker that is linked to both prior exposure and risk, and they can be measured easily and quantitatively using fluorescence in situ hybridization. In contrast to other biomarkers that are available, the frequency of reciprocal translocations in individuals exposed to whole-body radiation is stable with time after exposure, has rather small interindividual variability and can be measured accurately at low levels of exposure. Results from recent studies demonstrate that measurements of reciprocal translocation frequencies, facilitated by chromosome painting, can be used to reconstruct radiation dose for individuals exposed in the distant past.

NASA Discipline Radiation Health↗

Protective effect of apigenin on radiation-induced chromosomal damage in human lymphocytes

The potential use of flavonoids as a radioprotector is of increasing interest because of their high antioxidant activity and abundance in the diet. The aim of this study is to examine genotoxic and radioprotective effects of one of the most common flavonoids, apigenin, on radiation-induced chromosome aberrations in human lymphocytes. The cytokinesis-block micronucleus (CBMN) assay was used to evaluate such effects of apigenin. Blood samples were collected from two non-smoking healthy male volunteers who had no history of previous exposure to other clastogenic agents. Isolated lymphocytes were cultured. There were two tubes per concentration for all treatments. To evaluate the genotoxicity of apigenin, cells were first treated with different concentrations of apigenin (0, 2.5, 5, 10 and 25 microg/mL) at 24 h after culture initiation, followed by cytochalasin-B (Cyt-B) treatment (3 microg/mL) and cell harvest at 44 and 72 h, respectively. Secondly, to investigate the radioprotective effect, cell cultures were exposed to different concentrations of apigenin as described above for 30 min before being irradiated to 2 Gy of 137Cs gamma rays (at a dose rate of 0.75 Gy/min). In all instances, the frequency of MN was scored in binucleated (BN) cells. The nuclear proliferation index also was calculated. We did not detect an increase in the frequency of MN in non-irradiated human lymphocyte cultures treated with 2.5, 5.0 or 10 microg/mL apigenin; although, we did observe an increase in cultures treated with 25 microg/mL apigenin (the highest concentration of apigenin used in our study). We also observed a significant increase in the frequency of MN in irradiated cells overall; however, the frequency was decreased as the concentration of apigenin increased, suggesting a radioprotective effect. These findings provide a basis for additional studies to help clarify the potential use and benefit of apigenin as a radioprotector.

NASA Discipline Radiation Health↗

Discovery, characterization, and application of chromosomal integration sites in the hyperthermophilic archaeon Sulfolobus islandicus

Sulfolobus islandicus , an emerging archaeal model organism, offers unique advantages for metabolic engineering and synthetic biology applications owing to its ability to thrive in extreme environments. Although several genetic tools have been established for this organism, the lack of well-characterized chromosomal integration sites has limited its potential as a cellular factory. Here, in this work, we systematically identified and characterized 13 artificial CRISPR RNAs targeting eight integration sites in S. islandicus using the CRISPR-COPIES pipeline and a multi-omics-informed computational workflow. We leveraged the endogenous CRISPR-Cas system to integrate the reporter gene lacS and validated heterologous expression through a β-galactosidase assay, revealing significant positional effects. As a proof of concept, we utilized these sites to genetically manipulate lipid ether composition by overexpressing glycerol dibiphytanyl glycerol tetraether (GDGT) ring synthase B (GrsB). This study expands the genetic toolbox for S. islandicus and advances its potential as a robust platform for archaeal synthetic biology and industrial biotechnology.

59 BASIC BIOLOGICAL SCIENCES↗