RESUMEN
Premature infants or neonates in need of advanced clinical care must be transported to specialized hospitals. Past studies have examined vibrations experienced by patients during transport; however, multiple confounding factors limit the utility of on-road data. Hence, the development of a standardized test environment is warranted. The overall purpose of this project is to characterize vibrations during neonatal patient transport and develop mitigation strategies to reduce exposure. This paper focusses on the development of a laboratory test environment and procedure that enables studying the equipment vibration in a comprehensive and repeatable manner. For the first time, a complete neonatal patient transport system, including a stretcher, has been mounted on an industrial shaker. Results largely validate the system's ability to simulate on-road vibrations with high repeatability.
Asunto(s)
Hospitales Especializados , Vibración , Humanos , Lactante , Recién NacidoRESUMEN
Interfacility transport to tertiary care for high-risk neonates has become an integral part of equitable access to optimal perinatal healthcare. Excellence in clinical care requires expertise in transport medicine and the coordination of safe transport processes. However, concerns remain regarding environmental stressors involved in the transportation of sick high-risk neonates, including noise and vibration. In order to mitigate the potential deleterious effects of these physical stressors during transport, further knowledge of the burden of exposure, injury mechanisms and engineering interventions/modifications as adjuncts during transport would be beneficial. We reviewed the current literature with a focus on the contribution of new and emerging technologies in the transport environment with particular reference to whole-body vibration. This review intends to highlight what is known about vibration as a physical stressor in neonates and areas for further research; with the goal to making recommendations for minimizing these stressors during transport.
Asunto(s)
Incubadoras para Lactantes , Transporte de Pacientes , Vibración/efectos adversos , Ambulancias , Diseño de Equipo , Humanos , Lactante , Recién Nacido , Recien Nacido Prematuro , Transporte de Pacientes/métodosRESUMEN
The goals of this study were to assess three biomarkers of genetic effect for their individual and collective ability to detect and estimate radiation exposure in Russian Chernobyl clean-up workers. Work assignments were planned to limit dose to 0.25 Gy. The three biomarkers employed were chromosome translocations detectcd in lynmphocytes by florescence in situ hybridisation (FISH), and mutation at two genes, glycophorin A (GPA) in red blood cells detected by flow cytometry and hypoxanthine phosphoribosyltransferase (HPRT) in lymphocytes detected by selective cell culture. Samples were Obtained from 1992 to 2000. The time between exposure at Chernobyl and sample acquisition was > or =5 years. The lymphocyte assays detected an elevation over controls in average outcomes it clean-up workers: translocation rates were 46% higher when adjusted for age and smoking and HPRT mutant frequencies were were 16% higher when adjusted for age. The G PA assay did not detect an exposure effect. The results indicate that measuring frequency of translocations by FISH is preferred for low dose radiation, retrospective biochemistry.
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Glicoforinas/genética , Hipoxantina Fosforribosiltransferasa/genética , Linfocitos/efectos de la radiación , Enfermedades Profesionales/diagnóstico , Traumatismos por Radiación/diagnóstico , Translocación Genética , Adulto , Análisis Mutacional de ADN , Relación Dosis-Respuesta en la Radiación , Marcadores Genéticos , Humanos , Hibridación Fluorescente in Situ , Mutación/efectos de la radiación , Exposición Profesional , Reacción en Cadena de la Polimerasa , Centrales Eléctricas , Traumatismos por Radiación/genética , Liberación de Radiactividad Peligrosa , UcraniaRESUMEN
Werner syndrome (WRN) is an uncommon autosomal recessive disease in which progeroid features are associated with genetic instability and an elevated risk of neoplasia. We have used the glycophorin A (GPA) somatic cell mutation assay to analyze genetic instability in vivo in WRN patients and heterozygotes. GPA variant frequencies were determined for 11 WRN patients and for 10 heterozygous family members who collectively carry 10 different WRN mutations. Genetic instability as measured by GPA O/N allele loss variant frequency was significantly increased, and this increase was strongly age-dependent in WRN patients. GPA O/N allele loss variants were also significantly elevated in heterozygous family members, thus providing the first evidence for in vivo genetic instability in heterozygous carriers in an autosomal recessive genetic instability syndrome. Our results and comparable data on other human genetic instability syndromes allow an estimate of the level of genetic instability that increases the risk of human bone marrow dysfunction or neoplasia.
Asunto(s)
Enfermedades Hematológicas/genética , Heterocigoto , Síndrome de Werner/genética , Adolescente , Adulto , Factores de Edad , Anciano , Alelos , Estudios de Casos y Controles , ADN Helicasas/genética , Exodesoxirribonucleasas , Salud de la Familia , Femenino , Citometría de Flujo , Genotipo , Glicoforinas/metabolismo , Humanos , Masculino , Persona de Mediana Edad , Mutación , RecQ Helicasas , Factores de Riesgo , Helicasa del Síndrome de WernerRESUMEN
BACKGROUND: We previously reported a new optical configuration, in which both the side scatter and the fluorescence are collected using the index-guided, total internal reflection of a flow stream in air (the flow-stream waveguide). METHODS: Using a mixture of 0.202-microm and 0.093-microm diameter polystyrene beads, we have characterized the side scatter (SSC) sensitivity of a custom-built flow cytometer (miniFlo) which incorporates a flow-stream waveguide. RESULTS: The SSC-triggered SSC signal of 0.093-microm polystyrene beads in water was almost baseline resolved from the background. We also measured the SSC-triggered SSC signal of the same beads in water on our FACScan, which is a commercial unit with the conventional optical arrangement that uses a custom imaging objective to collect light from a sheath flow cuvette in perpendicular direction-the signal from 0.093-microm beads was not resolved from the background. CONCLUSIONS: The SSC sensitivity of miniFlo is one of the best reported in the literature. Cytometry 37:160-163, 1999. Published 1999 Wiley-Liss, Inc.
Asunto(s)
Citometría de Flujo/métodos , Citometría de Flujo/instrumentación , Sensibilidad y EspecificidadRESUMEN
Hereditary genetic defects in DNA repair lead to increased risk of cancer. Polymorphisms in several DNA repair genes have been identified; however, the impact on repair phenotype has not been elucidated. We explored the relationship between polymorphisms in the DNA repair enzyme, XRCC1 (codons 194, 280, and 399), and genotoxic end points measured in two populations: (a) placental aflatoxin B1 DNA (AFB1-DNA) adducts in a group of Taiwanese maternity subjects (n = 120); and (b) somatic glycophorin A (GPA) variants in erythrocytes from a group of North Carolina smokers and nonsmokers (n = 59). AFB1-DNA adducts were measured by ELISA, and erythrocyte GPA variant frequency (NN and NO) was assessed in MN heterozygotes with a flow cytometric assay. XRCC1 genotypes were identified by PCR-RFLPs. The XRCC1 399Gln allele was significantly associated with higher levels of both AFB1-DNA adducts and GPA NN mutations. Individuals with the 399Gln allele were at risk for detectable adducts (odds ratio, 2.4; 95% confidence interval, 1.1-5.4; P = 0.03). GPA NN variant frequency was significantly higher in 399Gln homozygotes (19.6 x 10(-6)) than in Gln/Arg heterozygotes (11.4 x 10(-6); P < 0.05) or Arg/Arg homozygotes (10.1 x 10(-6); P = 0.01). No significant effects were observed for other XRCC1 polymorphisms. These results suggest that the Arg399Gln amino acid change may alter the phenotype of the XRCC1 protein, resulting in deficient DNA repair.
Asunto(s)
Aflatoxina B1/sangre , Aductos de ADN/sangre , Daño del ADN , Reparación del ADN , Proteínas de Unión al ADN/genética , Glicoforinas/genética , Polimorfismo Genético , Femenino , Marcadores Genéticos , Genotipo , Humanos , Masculino , Proteínas/genética , Proteína 1 de Reparación por Escisión del Grupo de Complementación Cruzada de las Lesiones por Rayos XRESUMEN
11 persons, who had been irradiated chronically at low dose rate under occupational conditions in 1950s in doses 220-581 cGy according data of individual film dosimeters, and 5 control persons were examined regarding the level of glycophorin A (GPA) mutation type NO and NN in blood erythrocytes. Significantly higher level of GPA mutations type NO was registered in average in the group of exposed persons (23.2 +/- 4.6 x 10(-6)) compared with the control group (10.2 +/- 2.1 x 10(-6)) through the dose dependence was expressed slightly. The coefficient of the linear regression has equaled (2.3 +/- 1.2 x 10(-6)) Gy. The outlook on GPA assay usage in retrospective biodosimetry is discussed.
Asunto(s)
Glicoforinas/genética , Glicoforinas/efectos de la radiación , Mutación/genética , Enfermedades Profesionales/genética , Traumatismos por Radiación/genética , Anciano , Enfermedad Crónica , Relación Dosis-Respuesta en la Radiación , Femenino , Citometría de Flujo , Humanos , Masculino , Persona de Mediana Edad , Enfermedades Profesionales/sangre , Traumatismos por Radiación/sangre , Análisis de Regresión , Estudios Retrospectivos , Factores de TiempoRESUMEN
Blood samples were collected from 192 exposed workers who participated in the cleanup after the April 26, 1986, nuclear reactor accident at Chernobyl, Ukraine. These samples, together with samples from 73 individuals living in Russia but not involved in Chernobyl cleanup activities, were collected during September 1991 to May 1996 and shipped to the U.S. for evaluation by three bioassays: cytogenetic analysis based on chromosome painting, HPRT mutation analysis and glycophorin A (GPA) variant analysis. Univariate statistical analyses of the results of each bioassay (including adjustments for age, smoking status and estimated precision of the bioassay) found greater frequencies of chromosome translocations and HPRT mutant T lymphocytes among the exposed individuals compared to the controls (P < or = 0.01). GPA analyses showed no significant difference for exposed compared to controls for either hemizygous, N/O, or homozygous, N/N, variant cell frequency. Multivariate analysis of variance of the subset of 44 exposed and 14 unexposed individuals with measurements from all three bioassays found elevated frequencies of chromosomal translocations and HPRT mutants, and reduced frequencies for both GPA end points among the exposed persons compared to the controls. However, none of these differences, considered singly or in combination, was statistically significant (although statistical power is low due to small sample sizes). Mean estimated dose, based on cytogenetic response, for those exposed was 9 cGy (range 0 to 51 cGy) and was less than that estimated by physical dosimetry (25 cGy). Correlation between the end points of the bioassays and estimated physical dosimetry was low (r < 0.2); the only significant correlation found was for physical dose estimate and dates worked at Chernobyl (r = 0.4, P < 0.01), with those working soon after the accident receiving greater estimated doses.
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Traumatismos por Radiación/diagnóstico , Liberación de Radiactividad Peligrosa , Adolescente , Adulto , Factores de Edad , Anciano , Análisis de Varianza , Bioensayo , Aberraciones Cromosómicas , Glicoforinas/análisis , Humanos , Hipoxantina Fosforribosiltransferasa/genética , Sistema del Grupo Sanguíneo MNSs/genética , Masculino , Persona de Mediana Edad , Exposición Profesional , Centrales Eléctricas , Fumar , UcraniaRESUMEN
The frequency of peripheral blood erythrocyte variants exhibiting allelic loss of glycophorin A (N/M antigen) has been used previously as a biological dosimeter to assess somatic mutations in bone marrow cells from external whole-body irradiation. The aim of the present study was to determine whether this marker could be used as a measure of bone marrow genotoxicity induced by 131I in the treatment of thyroid cancer. Flow cytometry of immunolabeled erythrocytes was performed to enumerate glycophorin A variants before and after eight therapy doses of 131I administered to five patients with differentiated thyroid carcinoma. Bone marrow radiation exposure from each dose was calculated from the integrated retention of 131I in the whole body and in the blood. In addition, the accumulated dose to the bone marrow received from earlier 131I therapy was calculated for each patient. Regression analysis was performed on the frequency of two glycophorin A variant cell types (N/O and N/N) as a function of accumulated dose to the bone marrow. Frequency of N/O variant cells showed a significant dose-related increase with a slope of 10.9 x 10(-6) per sievert. This dose effect is about one-half that previously observed after whole-body external irradiation at high dose rate. This decreased response could be explained by the low dose rate of the radiation to the bone marrow from 131I.
Asunto(s)
Médula Ósea/efectos de la radiación , Glicoforinas/farmacología , Radioisótopos de Yodo/efectos adversos , Neoplasias de la Tiroides/radioterapia , Relación Dosis-Respuesta en la Radiación , Femenino , Humanos , Masculino , Dosis de RadiaciónRESUMEN
The 'spontaneous' frequency of genetic damage (normal background) and the possible relationship of this damage to nutritional variables in humans were investigated in 22 subjects using several indices of genetic damage. The subjects were chosen, out of 122 initially analyzed, for being at the extremes of the highest and lowest values of one index of genetic damage, the frequency of micronucleated erythrocytes in peripheral blood. This index reflects chromosomal damage and loss in bone marrow erythropoietic cells. The assay for micronuclei is convenient but is restricted to splenectomized individuals because the human spleen removes micronucleated cells. The initial 122 subjects were splenectomized, but all were normal and healthy at the time of this study and none had a previous history of neoplastic disease. Factors investigated were stability of micronucleus frequency as a function of time, correlations among multiple markers of genetic damage, and influence on damage indices of nutritional variables, including blood levels of folate, B12 and antioxidant vitamins. Among different individuals, the range of values was 10-fold or more in the erythrocyte micronucleus, glycophorin A, plasma ascorbate and urinary 8-hydroxydeoxyguanosine (oxo8dG) assays, was approximately 6-fold in the lymphocyte micronucleus assay, and was 2-fold in the lymphocyte sister chromatid exchange (SCE) assay. Red blood cell folate and plasma folate, B12 and alpha-tocopherol values varied by up to 10-fold among individuals. Micronucleus frequencies in erythrocytes and peripheral blood lymphocytes ranged from < 0.3 to 16.9/1000 in mature red blood cells, < 1 to 33/1000 in reticulocytes, and 2.5 to 15/1000 in binucleate lymphocytes. Frequencies of glycophorin A variant erythrocytes ranged from 5.6 to 77.3 x 10(6) N/0 cells and 3.2 to 16.2 x 10(6) N/N cells, and oxo8dG excretion varied from 32 to 397 pmol/kg/day. Although a wide range of values was observed in each genetic endpoint, the extreme values for various endpoints of genetic damage were not observed in the same individuals. The frequency of micronucleated erythrocytes varied over time within individuals and indicated that individuals with the highest levels of damage exhibit greater variability than those with lower levels. In some subjects, frequencies of micronucleated erythrocytes changed dramatically over an interval of 2-3 years: four subjects with initial micronucleated reticulocyte frequencies of 20.4, 5.9, 6.4 and 33/1000 changed to 2.5, 20.5, 18.5 and 12/1000, respectively. Among more than 150 individuals we have studied, including the 64 individuals studied by Everson et al. [(1988) J. Natl. Cancer Inst., 80, 525-529] and Smith et al. [(1990) Cancer Res., 50, 5049-5054], the seven individuals with the highest observed frequencies of micronucleated erythrocytes all had exceptionally low values of plasma folate, red cell folate, or plasma B12, suggesting that folate and B12 status are the major determinants of the types of damage that lead to spontaneous micronucleus formation in erythrocytic cells.
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Aberraciones Cromosómicas , Eritrocitos/ultraestructura , Micronúcleos con Defecto Cromosómico/genética , 8-Hidroxi-2'-Desoxicoguanosina , Análisis de Varianza , Ácido Ascórbico/sangre , Desoxiguanosina/análogos & derivados , Desoxiguanosina/orina , Ácido Fólico/sangre , Marcadores Genéticos , Glicoforinas/genética , Humanos , Linfocitos/citología , Estado Nutricional , Reticulocitos/citología , Intercambio de Cromátides Hermanas , Esplenectomía , Vitamina B 12/sangre , Vitamina E/sangreRESUMEN
In this article, we address the issue of persistence of chromosome exchanges following acute in vitro exposure of rat peripheral blood to 137Cs. Irradiation occurred 24 hr after culture initiation, and metaphase chromosomes were prepared 2, 3, 4, and 5 days later. Chromosomes 1, 2, and 4 were painted in unique colors and scored for structural aberrations. Dicentric chromosomes and acentric fragments diminished rapidly with time, as expected. Translocations exhibited greater persistence, but still showed a reduction in frequency, reaching a plateau of approximately 65 and 55% of their initial values, 4 days after exposure to 1 and 2 Gy, respectively. An exponentially declining model was fit to the combined dicentric, acentric fragment, and translocation frequencies, which showed that all three aberration types declined at equivalent rates. The frequencies of dicentrics and fragments declined to a plateau of zero, while translocations reached a plateau at frequencies significantly greater than zero. The decline in translocations with time is inconsistent with prevailing theoretical expectations, but is consistent with a model where some translocations are fully stable (persistent) and some are unstable (not persistent) through cell division. These results may have implications for radiation biodosimetry in humans.
Asunto(s)
Radioisótopos de Cesio/efectos adversos , Hibridación Fluorescente in Situ , Linfocitos/efectos de la radiación , Translocación Genética , Animales , Células Cultivadas , Rayos gamma/efectos adversos , Masculino , Índice Mitótico , Probabilidad , Ratas , Ratas Wistar , Factores de TiempoRESUMEN
Eighty individuals (55 adults and 25 children) who were residents of four cities (Kiev, Mozyr, Gomel and Bobrujsk) located 100-200 km from Chernobyl at the time of the accident in 1986 were tested after immigrating to the US from 1989-1991. A whole-body counter was employed to quantitate radiocesium content. In addition, two biological measures of radiation effects, namely, chromosomal integrity using the micronucleus assay and somatic mutation analysis of erythrocytes at the glycophorin A (GPA) locus, were applied to this group. Radiocesium activity in the body ranged from 0 to 56.8 Bq/kg with a mean and standard deviation of 5.0 +/- 8.2 and a median value of 2.0 Bq/kg. Mean radiocesium content by groups was highest in adult males (9.0 +/- 11.7; range 0.21-56.8 Bq/kg) followed by adult females (3.3 +/- 4.5; range 0-21.3 Bq/kg), male children (3.0 +/- 5.7; range 0-20.2 Bq/kg) and lowest in female children (1.6 +/- 3.5; range 0-12.7 Bq/kg). Individuals with the highest radiocesium content in each group belonged to one family that lived in Mozyr (100 km from Chernobyl) until emigrating in 1989. The frequency of lymphocyte micronuclei and erythrocyte GPA allele-loss (O/N) somatic mutations were both significantly correlated with radiocesium content (r=0.57, p=0.002; r=0.75, p=0.002, respectively). The micronucleus frequency also correlated with the estimated internal absorbed dose from radiocesium in a subset of 20 immigrants for whom this calculation was possible (r=0.71, p=0.0005). Altogether, the biomonitoring data indicate that some subjects had radiation doses sufficient to produce gene and chromosomal mutations in blood cells, although these effects cannot be attributed solely to radiocesium exposure.
Asunto(s)
Centrales Eléctricas , Liberación de Radiactividad Peligrosa , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Niño , Preescolar , Eritrocitos/química , Eritrocitos/efectos de la radiación , Femenino , Glicoforinas/análisis , Humanos , Linfocitos/efectos de la radiación , Linfocitos/ultraestructura , Masculino , Micronúcleos con Defecto Cromosómico/efectos de la radiación , Persona de Mediana Edad , Reactores Nucleares , Ucrania/etnología , Irradiación Corporal TotalRESUMEN
We have used the glycophorin A (GPA) in vivo somatic cell mutation assay to assess the genotoxic potential of styrene exposure in 47 reinforced plastics workers occupationally exposed to styrene and 47 unexposed controls matched for age, gender, and active smoking status. GPA variant erythrocyte frequencies (Vf), reflecting GPA allele loss (phi/N) and allele loss and duplication (N/N) somatic mutations arising in vivo in the erythroid progenitor cells of individuals of GPA M/N heterozygous genotype, were flow cytometrically determined in peripheral blood samples from these subjects. Measurements of styrene exposure of the workers at the time of blood sampling showed a mean 8-h time-weighted average (TWA8-h) styrene concentration of 155 mg/m3 (37 ppm) in the breathing zone. Mean urinary concentrations of the styrene metabolites mandelic acid (MA) and mandelic acid plus phenyl glyoxylic acid (MA+PGA) were 4.4 mmol/liter (after workshift) and 2.1 mmol/liter (next morning), respectively. Multivariate analysis of covariance on log-transformed GPA Vf data with models allowing adjustment for age, gender, smoking status, and styrene exposure showed that N/N Vf were nearly significantly increased among all of the exposed workers (adjusted geometric mean, 6.3 per million versus 5.0 in the controls; P = 0.058) and were statistically significantly elevated (adjusted geometric mean, 6.8 versus 5.0 in the controls; P = 0.036) among workers classified into a high-exposure group according to personal TWA8-h concentration of styrene in the breathing zone of > or = 85 mg/m3 (20 ppm; Finnish threshold limit value). Women in this high exposure group showed especially elevated N/N Vf (adjusted geometric mean 8.5 versus 5.3 in control women; P = 0.020); this elevation was also significant if urinary MA+PGA of > or = 1.2 mmol/liter was used as the basis of classification (adjusted geometric mean, 8.3; P = 0.030). The occupational exposure could not be shown to influence phi/N Vf. Cigarette smoking was associated with significantly elevated GPA Vf among active smokers (P = 0.042 for phi/N and P = 0.020 for N/N) and among active and ex-smokers combined (P = 0.014 for N/N). Its influence on phi/N Vf was especially clear among active smokers in the control group (P = 0.005). An effect of smoking, nearly statistically significant, was also observed for the phi/N Vf of control ex-smokers (P = 0.055) and of all active and ex-smokers combined (P = 0.050). Thus, the two characterized chemical exposures experienced by this group of workers and controls appear to produce differential effects on the two independent classes of GPA variants enumerated in the assay. This result suggests that the genotoxicity of these agents is mediated, at least in part, by different genetic mechanisms. Styrene exposure is associated with a specific increase in GPA N/N Vf; these allele loss and duplication variants reflect predominantly somatic recombination mechanisms in erythroid progenitor cells. Tobacco smoke exposure in active and ex-smokers is also associated not only with an increase in N/N Vf but also with an increase in phi/N Vf, reflecting the induction of GPA gene-inactivating mutations, including point mutations and deletions. This finding is consistent with a broad mechanistic spectrum of tobacco smoke genotoxicity associated with this complex mixture of chemical mutagens. Finally, there was no detectable effect of age on phi/N Vf; however, a highly significant (P = 0.0002) increase in N/N Vf with age, even after adjustment for other variables, was observed.
Asunto(s)
Glicoforinas/genética , Mutación , Exposición Profesional/efectos adversos , Estirenos/efectos adversos , Adulto , Células Precursoras Eritroides/citología , Células Precursoras Eritroides/efectos de los fármacos , Femenino , Finlandia , Humanos , Modelos Lineales , Masculino , Análisis Multivariante , Pruebas de Mutagenicidad , Mutación/genética , Plásticos , Fumar , EstirenoRESUMEN
Three human chromosome 2-specific clone libraries were constructed and characterized. Chromosome 2-specific cosmid and fosmid clone libraries were constructed using flow-sorted DNA from the monochromosomal hybrid cell line GM10826. The cosmid and fosmid libraries consist of 38,496 and 26,400 arrayed clones, respectively, with an average size of 40 kb. Colony hybridization of a representative number of clones with both human and hamster genomic DNA probes demonstrates that between 58 and 66% of the clones in the flow-sorted libraries contain human inserts. Approximately 5% of the cosmid and fosmid clones are nonrecombinants. A chromosome 2-specific PAC library was also produced from the hybrid cell line GM10826. DNA from the hybrid cell line was cloned, and the human chromosome 2-specific clones were identified by colony hybridization. Approximately 5800 chromosome 2-specific PAC clones with an average insert size of approximately 85 kb were arrayed. Based on the size of the clones, the cosmid, fosmid, and PAC libraries are approximately 3.6x, approximately 2.5x, and approximately 1.9x, respectively in chromosomal coverage. The chromosome 2 coverage of each of the three libraries was further determined by PCR screening clone pools with 82 chromosome 2-specific STSs. The average number of clones identified for each STS in the library indicates the cosmid, fosmid, and PAC libraries to be approximately 3.2x, approximately 2.1x, and approximately 1.5x, respectively, in chromosome coverage. All except one of the 82 STSs were represented in the portions of the libraries screened.
Asunto(s)
Cromosomas Humanos Par 2/genética , Clonación Molecular , Biblioteca de Genes , Animales , Mapeo Cromosómico , Cósmidos , Cricetinae , ADN/genética , Marcadores Genéticos , Humanos , Células HíbridasRESUMEN
In 1986, when an explosion accident occurred at the Chernobyl, Ukraine nuclear power plant, a large number of people were exposed to significant amounts of ionizing radiation. During the time between 1986 and 1992, peripheral blood samples were obtained from 102 people who either were on site during the emergency or were brought to Chernobyl shortly thereafter to assist in the cleanup of radioactive contaminants and isolate the damaged reactor from the environment. These blood samples plus samples from 13 unexposed Soviet individuals were analyzed by flow cytometry using the allele-loss somatic mutation assay for glycophorin A. Results of these assays show that the frequency of N/O variant red cells increased in proportion to the estimated radiation exposure of each individual. The radiation dose-response function derived from this population closely resembles that determined previously for atomic bomb survivors whose blood samples were obtained and analyzed 40 years after their exposure. This suggests comparable mutation induction per unit dose for these two populations and long-term persistence of the mutational damage. In addition, measurements on multiple blood samples from each of 10 donors taken over a 7-year period showed no significant changes in N/O variant cell frequencies, confirming the persistence of radiation-induced somatic mutations in long-lived bone marrow stem cells.
Asunto(s)
Eritrocitos/efectos de la radiación , Glicoforinas/genética , Mutación , Centrales Eléctricas , Liberación de Radiactividad Peligrosa , Relación Dosis-Respuesta en la Radiación , Eritrocitos/metabolismo , Femenino , Humanos , Masculino , UcraniaRESUMEN
Glycophorin A (GPA) is an erythroid-lineage-specific membrane sialoglycoprotein which occurs in two allelic forms, M and N, which form the antigens of the MN blood group. Purified cDNAs and RNAs isolated from peripheral blood and erythroleukemia cell lines, HEL and K562, were used to develop an RT-PCR technique for amplifying GPA gene transcripts (GYPA). The relative expression of transcripts from the M and N alleles was determined using restriction analysis of these amplified products with four allele-specific restriction endonucleases. The use of this method permits the sensitive identification of GYPA transcripts in these cells and confirms GPA protein expression in the erythroleukemia cell lines and the MN phenotypes of individuals determined by immunolabeling with GPA allele-specific monoclonal antibodies. A novel restriction pattern was obtained using peripheral blood RNA from two individuals with a rare inherited variant allele, GPA Mg. Sequencing of the cDNA obtained using this method revealed a single C to A transversion in the fourth codon in the mature GYPA N coding sequence is responsible for the difference between GYPA Mg and GYPA N.
Asunto(s)
Células Precursoras Eritroides/metabolismo , Glicoforinas/genética , ARN Mensajero/sangre , Alelos , Secuencia de Aminoácidos , Secuencia de Bases , Humanos , Leucemia Eritroblástica Aguda/metabolismo , Datos de Secuencia Molecular , Fenotipo , Reacción en Cadena de la Polimerasa , Polimorfismo de Longitud del Fragmento de Restricción , ARN Mensajero/aislamiento & purificación , Reticulocitos/metabolismo , Células Tumorales CultivadasRESUMEN
To assess the potential effect of maternal environments on human embryonic/fetal somatic mutation, we measured the frequencies of hypoxanthine-guanine phosphoribosyltransferase (HPRT, hprt gene), mutant T lymphocytes (Mf), and glycophorin A (GPA) variant erythrocytes (Vf) of both allele-loss (phi/N) and allele-loss-and-duplication (N/N) phenotypes in umbilical cord blood. The mean hprt Mf (1.40 +/- 1.11 x 10(-6), N = 66) and GPA Vf (phi/N 4.0 +/- 2.2 x 10(-6), N = 114; N/N 2.7 +/- 2.0 x 10(-6), N = 91) were significantly lower than those previously reported for adult populations. In addition, the hprt Mf was significantly higher than that of a published study of newborn cord blood samples from a geographically distant population (0.64 +/- 0.41 x 10(-6), N = 45, P < 0.01; t test, P < 0.01, Mann-Whitney U test). An examination of the demographic data from these two populations led to the sampling of 10 additional newborns specifically matched to the published study for maternal socioeconomic status. The hprt Mf (0.70 +/- 0.49 x 10(-6)) of this selected population was consistent with the published report and significantly lower than that of our initial population (P < 0.03, t test; P < 0.01, Mann-Whitney U test). These results indicate that there is an environmental effect related to maternal socioeconomic status on the frequency of embryonic/fetal somatic mutations. Molecular analyses of hprt mutants from this cohort with elevated Mf revealed a significant decrease in the relative contribution of gross structural mutations to the overall Mf (25 of 38, 66% vs. 34 of 41, 83%, P = 0.024, chi 2 test), suggesting that the higher Mf resulted from an elevated level of "point" mutations. No individual maternal demographic or environmental factor was identified as contributing more significantly than other any factor to the observed variability in hprt Mf or GPA Vf.
Asunto(s)
Contaminantes Ambientales , Sangre Fetal , Glicoforinas/genética , Hipoxantina Fosforribosiltransferasa/genética , Mutación , Adulto , Análisis de Varianza , Clonación Molecular , Colorado , ADN/sangre , Eritrocitos/enzimología , Etnicidad , Femenino , Eliminación de Gen , Humanos , Hipoxantina Fosforribosiltransferasa/sangre , Recién Nacido , Masculino , Embarazo , Factores de Riesgo , Caracteres Sexuales , Fumar , Linfocitos T/citologíaRESUMEN
The glycophorin A (GPA) assay for in vivo somatic cell mutations was performed on blood samples from 39 survivors of the atomic bomb at Hiroshima. Parallel analyses were performed at two laboratories using three different GPA assay methods to enumerate cells lacking expression of either the M- or N-allele of GPA. All assay methods yielded significant dose-dependent increases in hemizygous GPA variant cell frequencies (VFs) and smaller increases in homozygous VFs. The slopes of the fitted linear dose-response functions did not differ significantly among assay methods used in the present study, or from slopes obtained in a study reported previously. The version of the assay described most recently (BR6) appears best suited for future studies because the assay has a higher precision than earlier methods. Variant frequencies from different assay methods measuring the same variant cell type agreed with each other better than with the estimated dose, suggesting that the imprecision in the assay is not primarily responsible for VFs that differ from the fitted dose response. Consistent deviations from the dose response were seen for some individuals, suggesting either errors in dose estimates for these individuals or interindividual differences in susceptibility or other exposures. For the study population as a whole, however, discrepancies between assays for M-loss and N-loss variants suggest stochastic factors may have an important effect on individual VFs for A-bomb survivors.
Asunto(s)
Glicoforinas/genética , Mutación , Guerra Nuclear , Mapeo Cromosómico , Pruebas Enzimáticas Clínicas/métodos , Relación Dosis-Respuesta en la Radiación , Glicoforinas/metabolismo , Humanos , JapónRESUMEN
The glycophorin A (GPA) assay is a human mutation assay that is potentially useful for large epidemiological studies. The assay is rapid and requires a minimal amount of blood, which can be stored before analysis. The data presented here were collected from workers exposed to styrene in a boat manufacturing plant. This study was the first to apply the GPA assay to an occupationally exposed population. Subjects with a mean styrene exposure of 30 ppm had a higher frequency of GPA N phi variant cells than subjects with mean exposure of 1 ppm, but the subjects differed in respect to smoking and age distribution. Results indicate that the original 1-W-1 version of the assay may not be suitable for studies of small numbers of exposed subjects due to variability and artifacts. The newer BR6 version, however, has much lower variability and shows promise for use in the occupational setting.
Asunto(s)
Glicoforinas/genética , Pruebas de Mutagenicidad/métodos , Exposición Profesional , Estirenos/efectos adversos , Estudios de Evaluación como Asunto , Frecuencia de los Genes , Variación Genética , Humanos , EstirenoRESUMEN
Hemopoietic stem cells from human fetal liver were transplanted in utero into preimmune fetal sheep (48-54 days of gestation). The fate of donor cells was followed using karyotype analysis, by immunofluorescence labeling with anti-CD antibodies, and by fluorescent in situ hybridization using human-specific DNA probes. Engraftment occurred in 13 of 33 recipients. Of five live born sheep that exhibited chimerism, all expressed human cells in the marrow, whereas three expressed them in blood as well. Engraftment was multilineage (erythroid, myeloid, and lymphoid) and human hemopoietic progenitors (multipotent colony-forming units, colony-forming units-granulocyte, macrophage, and erythroid burst-forming units) capable of forming colonies in vitro were detected in all five lambs for greater than 2 yr. These progenitors responded to human-specific growth factors both in vitro and in vivo. Thus the administration of recombinant human IL-3 and granulocyte macrophage-colony-stimulating factor to chimeric sheep resulted in a 2.1-3.4-fold increase in the relative expression of donor (human) cells. These results demonstrate that the permissive environment of the preimmune fetal sheep provides suitable conditions for the engraftment and long-term multilineage expression of human hemopoietic stem cells in a large animal model. In this model, donor human cells appear to retain certain phenotypic and functional characteristics that can be used to manipulate the size of donor cell pool.