RESUMO
Large deletions and other gross forms of chromosome imbalance are known in man but have rarely been found in the mouse. By screening progeny of spermatogonially irradiated male mice for a combination of runting and other phenotypic effects, we have identified animals that have large deletions comprising from 2.5-30 percent of the length of individual chromosomes, or other major chromosome changes, which are compatible with viability and fertility. Certain chromosome regions appear particularly susceptible to the generation of viable deletions and this has implications for radiation mutagenesis studies. Correlations with human deletions are also indicated.
Assuntos
Aberrações Cromossômicas , Deleção Cromossômica , Transtornos Cromossômicos , Fertilidade/genética , Viabilidade Fetal/genética , Animais , Aberrações Cromossômicas/genética , Feminino , Cariotipagem , Masculino , Camundongos , Camundongos Endogâmicos C3H , FenótipoRESUMO
An approach designed to map and generate mutations in the region of the short arm of the mouse Y chromosome, known to be involved in sex determination and spermatogenesis, is described. This relies on homologous Yp-Sxra pairing and asymmetrical exchange which can occur at meiosis in XY males carrying Sxra on their X chromosome. Such exchange potentially generates deficiencies and duplications of Yp or Sxra. Three fertile XY females were found out of about 450 XY offspring from XSxra/Y x XX crosses. In all three, despite evidence for deletion of Y chromosomal material, the Sry locus was intact. Each deletion involved a repeat sequence, Sx1, located at a distance from Sry. Since expression of Sry was affected these results suggest that long range position effects have disrupted Sry action.
Assuntos
Deleção Cromossômica , Transtornos do Desenvolvimento Sexual , Proteínas Nucleares , Fatores de Transcrição , Cromossomo X , Cromossomo Y , Animais , Sequência de Bases , Primers do DNA , Proteínas de Ligação a DNA/genética , Feminino , Masculino , Camundongos , Camundongos Endogâmicos , Dados de Sequência Molecular , Diferenciação Sexual/genética , Proteína da Região Y Determinante do SexoRESUMO
Angelman syndrome (AS) is a human genetic disorder characterized by mental retardation, seizures, inappropriate laughter, abnormal galt, tremor and ataxia. There is strong genetic evidence that the disorder is associated with a maternally expressed, imprinted gene mapping to chromosome 15q11-13. Affected patients demonstrate varied molecular abnormalities, including large maternal deletions, uniparental paternal disomy (UPD). Imprinting mutations and loss of function mutations of E6-associated-protein (E6-AP) ubiquitin-protein ligase (UBE3A). All of these abnormalities are associated with loss of maternal expression of UBE3A. Although mutations in UBE3A cause AS, indicating that maternal-specific expression of UBE3A is essential for a normal phenotype, evidence for maternal-specific expression of UBE3A has been lacking. Using mice with partial paternal UPD encompassing Ube3a to differentiate maternal and paternal expression, we found by in situ hybridization that expression of Ube3a in Purkinje cells, hippocampal neurons and mitral cells of the olfactory bulb in UPD mice was markedly reduced compared to non-UPD littermates. In contrast, expression of Ube3a in other regions of the brain was only moderately or not at all reduced in UPD mice. The major phenotypic features of AS correlate with the loss of maternal-specific expression of Ube3a in hippocampus and cerebellum as revealed in the mouse model.
Assuntos
Síndrome de Angelman/genética , Hipocampo/metabolismo , Ligases/genética , Neurônios/metabolismo , Células de Purkinje/metabolismo , Síndrome de Angelman/metabolismo , Síndrome de Angelman/patologia , Animais , Cerebelo/metabolismo , Cerebelo/patologia , Mapeamento Cromossômico , Cromossomos Humanos Par 15 , Embrião de Mamíferos , Feminino , Expressão Gênica , Impressão Genômica , Hipocampo/patologia , Humanos , Ligases/biossíntese , Masculino , Camundongos , Camundongos Endogâmicos ICR , Dados de Sequência Molecular , Neurônios/patologia , Células de Purkinje/patologia , Caracteres Sexuais , Translocação Genética , Ubiquitina-Proteína LigasesRESUMO
The best examples of imprinting in humans are provided by the Angelman and Prader-Willi syndromes (AS and PWS) which are associated with maternal and paternal 15q11-13 deletions, respectively, and also with paternal and maternal disomy 15. The region of the deletions has homology with a central part of mouse chromosome 7, incompletely tested for imprinting effects. Here, we report that maternal duplication for this region causes a murine imprinting effect which may correspond to PWS. Paternal duplication was not associated with any detectable effect that might correspond with AS. Gene expression studies established that Snrpn is not expressed in mice with the maternal duplication and suggest that the closely-linked Gabrb-3 locus is not subject to imprinting. Finally, an additional new imprinting effect is described.
Assuntos
Autoantígenos/genética , Modelos Genéticos , Síndrome de Prader-Willi/genética , Ribonucleoproteínas Nucleares Pequenas/genética , Animais , Mapeamento Cromossômico , Feminino , Expressão Gênica , Humanos , Masculino , Camundongos , Família Multigênica , Translocação Genética , Proteínas Centrais de snRNPRESUMO
Albino mammals have abnormal visual pathways. "Flecked" mice have a variegated pigment distribution, with roughly half the cells normally pigmented and half albino. They do not show the albino abnormality in the visual pathways. The gene at the albino locus, which determines the course of the visual pathways must, thus, have an extracellular action.
Assuntos
Albinismo/genética , Vias Visuais/anormalidades , Albinismo/complicações , Animais , Axônios , Aberrações Cromossômicas , Cruzamentos Genéticos , Feminino , Lateralidade Funcional , Gânglios/patologia , Heterozigoto , Homozigoto , Masculino , Camundongos , Nervo Óptico/anormalidades , Nervo Óptico/patologia , Retina/anormalidades , Retina/patologia , Pigmentos da Retina , Transtornos da Visão/complicações , Transtornos da Visão/genéticaRESUMO
The mouse insulin-like growth factor II gene (Igf 2), located on distal chromosome 7, is parentally imprinted such that the paternal allele is expressed while the maternal allele is transcriptionally silent. We derived a cell line from a mouse embryo maternally disomic and paternally deficient for distal chromosome 7 (MatDi7) to determine the stability of gene repression in culture. MatDi7 cells maintained Igf2 in a repressed state even after immortalization, except for one randomly picked clone which spontaneously expressed the gene. Igf 2 was expressed in a cell culture derived from a normal littermate; this expression was growth regulated, with Igf 2 mRNA levels increasing in the stationary phase of growth. Analysis of the methylation status of 28 sites distributed over 10 kb of the gene did not show consistent differences associated with expression level in the normal and MatDi7 cell lines, and the CpG island in the Igf 2 promoter remained unmethylated in all of the cell lines. Only with an oncogenically transformed cell line did the promoter become extensively methylated. We attempted to derepress the imprinted gene in MatDi7 cells by treatments known to alter gene expression. Expression of the Igf 2 allele in MatDi7 cells was increased in a dose-dependent manner by treatment with 5-aza-2'-deoxycytidine or bromodeoxyuridine, agents known to change DNA methylation patterns or chromatin conformation. Treatment of the cells with 1-beta-D-arabinofuranosylcytosine, 2'-deoxycytidine, calcium ionophore, heat shock, cold shock, or sodium butyrate did not result in increases in the levels of Igf 2 expression. It seems likely that the mechanism of the Igf 2 imprint involves subtle changes in the methylation or chromatin conformation of the gene which are affected by 5-aza-2'-deoxycytidine and bromodeoxyuridine.
Assuntos
Regulação da Expressão Gênica , Genes , Fator de Crescimento Insulin-Like II/genética , Animais , Sequência de Bases , Células Cultivadas , Cromatina/ultraestrutura , Temperatura Alta , Técnicas In Vitro , Metilação , Camundongos , Dados de Sequência Molecular , Oligonucleotídeos/química , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , Mapeamento por RestriçãoRESUMO
Mice with uniparental disomies (uniparental duplications) for defined regions of certain chromosomes, or certain disomies, show a range of developmental abnormalities most of which affect growth. These defects can be attributed to incorrect dosages of maternal or paternal copies of imprinted genes lying within the regions involved. Combinations of certain partial disomies result in interactions between the imprinting effects that seemingly independently affect foetal and/or placental growth in different ways or modify neonatal and postnatal development. The findings are generally in accord with the 'conflict hypothesis' for the evolution of genomic imprinting but do not demonstrate common growth axes within which imprinted genes may interact. Instead, it would seem that any gene that favours embryonic/foetal development, at consequent cost to the mother, will have been subject to evolutionary selection for only paternal allele expression. Reciprocally, any gene that reduces embryonic/foetal growth to limit disadvantage to the mother will have been selected for only maternal allele expression. It is concluded that survival of the placenta is core to the evolution of imprinting.
Assuntos
Cromossomos Humanos Par 11 , Cromossomos Humanos Par 2 , Cromossomos Humanos Par 7 , Cromossomos Humanos Par 9 , Impressão Genômica , Mapeamento Cromossômico , HumanosRESUMO
The agouti locus on mouse chromosome 2 encodes a secreted cysteine-rich protein of 131 amino acids that acts as a molecular switch to instruct the melanocyte to make either yellow pigment (phaeomelanin) or black pigment (eumelanin). Mutations that up-regulate agouti expression are dominant to those causing decreased expression and result in yellow coat color. Other associated effects are obesity, diabetes, and increased susceptibility to tumors. To try to define important functional domains of the agouti protein, we have analyzed the molecular defects present in a series of recessive viable agouti mutations. In total, six alleles (amJ, au, ada, a16H, a18H, ae) were examined at both the RNA and DNA level. Two of the alleles, a16H and ae, result from mutations in the agouti coding region. Four alleles (amJ, au, a18H, and ada) appear to represent regulatory mutations that down-regulate agouti expression. Interestingly, one of these mutations, a18H, also appears to cause an immunological defect in the homozygous condition. This immunological defect is somewhat analogous to that observed in motheaten (me) mutant mice. Short and long-range restriction enzyme analyses of homozygous a18H DNA are consistent with the hypothesis that a18H results from a paracentric inversion where one end of the inversion maps in the 5' regulatory region of agouti and the other end in or near a gene that is required for normal immunological function. Cloning the breakpoints of this putative inversion should allow us to identify the gene that confers this interesting immunological disorder.
Assuntos
Alelos , Genes Recessivos , Peptídeos e Proteínas de Sinalização Intercelular , Camundongos Mutantes/genética , Proteínas/genética , Proteína Agouti Sinalizadora , Animais , Sequência de Bases , Inversão Cromossômica , Mapeamento Cromossômico , DNA/genética , Regulação da Expressão Gênica , Cor de Cabelo/genética , Doenças Pulmonares Intersticiais/genética , Transtornos Linfoproliferativos/genética , Masculino , Melaninas/biossíntese , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Mutagênese , Fenótipo , RNA Mensageiro/genética , Transcrição GênicaRESUMO
Insulin-like growth factor 2 (Igf 2) and H19 genes are oppositely imprinted and as such have been most extensively studied imprinted genes both genetically and at the molecular level. Imprints of the H19 gene, being established during spermatogenesis, are epigenetically transmitted to the somatic cells of the embryo. Current hypotheses attempting to explain the allele-specific silence of the H19 gene include DNA methylation and chromatin condensation. In order to understand the molecular basis of H19 epigenesis, it is crucial to identify the markings in the chromatin organising the imprinted domain in spermatozoa. Using Micrococcal nuclease (MNase), DNase I and Methidiumpropyl-EDTA. iron II (MPE.Fe(II)) as chromatin probes, we demonstrate that in mouse epididymal spermatozoa, at least 4kb DNA upstream of the H19 'cap' site, containing the imprinted and differentially methylated domain (DMD), is heterochromatic. The cleavage sites in this domain (-2 to -4kb) exhibit approximately 425bp periodicity. This structure is maintained in the paternal allele of normal embryos and is disrupted at -2.2, -2.65 and at -3.5kb in embryos maternally disomic for the distal end of chromosome 7 (MatDp 7). The hypersensitive sites in chromatin precisely register the MPE.Fe(II) cleavage sites in chromosomal DNA. Therefore, the DNA sequences in the imprinted domain constrain the chromatin structure in a way similar to that of 1.688g/cm(3) Drosophila satellite chromatin. In addition, we find that condensation of the paternal allele correlates with methylation-dependent alteration in the structure of DNA sequences in DMD. These results suggest that CpG-methylation induces localised changes in DNA conformation and these facilitate consequent remodelling of chromatin thereby allowing the paternal and maternal H19 alleles to be distinguished.
Assuntos
Impressão Genômica , Heterocromatina , Proteínas Musculares/genética , RNA não Traduzido , Espermatozoides , Alelos , Animais , Metilação de DNA , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Nucleossomos/metabolismo , RNA Longo não CodificanteRESUMO
The objective of this paper was to investigate by pedigree-based genetic means the origins and inheritance of arrhythmogenic right ventricular cardiomyopathy (ARVC) in UK Boxers and assess the role of the proposed causal mutation in the gene, Striatin (STRN). All ARVC cases traced back to a small number of imported American dogs deriving from the group of Boxers studied by Harpster (1983) to define the disease, strongly suggesting that the disease is the same in the two countries. Dogs with and without the STRN mutation were found in both ARVC affected and normal Boxers showing that the mutation is not responsible for the disease. Evidence was found that the STRN mutation is, however, genetically linked with the gene responsible on the same chromosome. The linkage implies that the two genes can separate by meiotic recombination such that both ARVC-affected and ARVC-unaffected lines of dogs may carry either the STRN mutation or its wild-type allele. These have been found. Homozygotes for the STRN mutation tended to be severely affected at early ages, suggesting that there is an interaction between the known effects of the STRN mutation on the cardiomyocyte and ARVC.
Assuntos
Displasia Arritmogênica Ventricular Direita/veterinária , Doenças do Cão/genética , Proteínas de Membrana/genética , Linhagem , Animais , Displasia Arritmogênica Ventricular Direita/genética , Cães , Feminino , Masculino , Mutação , Especificidade da Espécie , Reino UnidoRESUMO
31P magnetic resonance spectroscopy has been used to compare the effects of the vasoactive agents hydralazine and flunarizine on the oxygenation of the transplantable tumors, SCCVII/Ha and 16C, and a range of spontaneous mammary tumors arising in the breeding stock in the Genetics Division at the Radiobiology Unit. The vasodilator hydralazine, previously shown to increase the radiobiological hypoxic fraction of transplantable murine tumors, increased inorganic phosphate to total phosphate (Pi/total) in SCCVII/Ha and 16C tumors. However, only two spontaneous tumors responded to this agent (2/12). The calcium antagonist flunarizine, which sensitizes the SCCVII tumor to X rays, consistent with a reduction in hypoxic fraction, reduced Pi/total in this and the 16C tumor. Further, most spontaneous tumors tested (8/10) responded to this agent, as measured by a reduction in Pi/total. These results point to fundamental differences between transplantable and spontaneously arising tumors in mice in their response to vasoactive agents.
Assuntos
Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Experimentais/metabolismo , Vasodilatadores/farmacologia , Animais , Feminino , Flunarizina/administração & dosagem , Flunarizina/farmacologia , Hidralazina/administração & dosagem , Hidralazina/farmacologia , Injeções Intraperitoneais , Injeções Intravenosas , Espectroscopia de Ressonância Magnética , Camundongos , Camundongos Endogâmicos C3H , Transplante de Neoplasias , Oxigênio/metabolismo , Fosfatos/metabolismo , Vasodilatadores/administração & dosagemRESUMO
Evidence of an enhanced incidence of lung tumours (benign adenomas and adenocarcinomas) was sought in the BALB/cJ mouse following paternal germ cell X-irradiation. In a series of replicate studies spanning approximately 1 year, males were exposed to single, acute X-ray doses of 0, 250 and 500 cGy. In each of the 2 consecutive weeks immediately thereafter they were placed with two females to generate progeny that were derived from irradiated post-meiotic cells (spermatozoa to late spermatids). These animals were then examined at 8 or 12 months for lung tumours. While the proportion of fertile females and mean litter size was affected by the radiation, showing a dose-dependent, dominant lethal response, and while cases of mutant offspring were detected, the paternal radiation did not affect lung tumour incidence in the offspring. The incidence did not vary significantly between germ cell stages irradiated (week of mating), sex of offspring, or radiation dose. However, significant differences between lung tumour incidence (mostly representing benign adenomas) were found between different replicates, these being high at the start of the study, declining and then rising to yet higher levels at its close. The finding that lung tumour incidence in BALB/cJ mice is not affected by paternal germ cell irradiation does not accord with Nomura's reports using other strains of mice. This, in turn, weakens biological support for a causal association between the raised incidence of childhood leukaemia and non-Hodgkin lymphoma near Sellafield and the father's recorded radiation exposure during employment by the nuclear industry.
Assuntos
Adenoma/etiologia , Neoplasias Pulmonares/etiologia , Neoplasias Induzidas por Radiação/genética , Adenoma/genética , Animais , Relação Dose-Resposta a Droga , Feminino , Tamanho da Ninhada de Vivíparos/efeitos da radiação , Neoplasias Pulmonares/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Raios XRESUMO
Earlier studies have shown that the genetic response to X-irradiation of mouse spermatogonial stem-cell populations that are recovering from a previous radiation exposure may differ from that of a normal, unirradiated stem-cell population. Similar modified responses to X-irradiation have now been observed in stem spermatogonia that are recovering from treatment with the chemical mutagen, TEM. (1) In contrast to a normal response to 900 R, high translocation yields were obtained when this dose was administered 24 h after a TEM treatment. (2) A small but non-significant increase above a normal response to 500 R was obtained when this dose followed 24 h after TEM treatment and a normal response to 500 R was obtained from the reverse order treatment (500 R + TEM). (3) When given 4 days after a TEM treatment, the translocation yield from 500 R was only about half that normally obtained. Unexpectedly, a similar low response was obtained from the reverse order treatment which, if verified, would suggest selective cell killing by the TEM. THe chemical administered alone, was almost totally ineffective in producing recoverable translocations in stem spermatogonia. Since it is unlikely that TEM and X-rays should similarly synchronize the cell cycle of surviving stem spermatogonia it is concluded that the modified genetic responses obtained result from a different cause. Depletion of the stem-cell population is suggested as the common mediating factor. This may 'trigger' the radio-resistant, long cycling stem cells into a more active cycle such that, 24--48 h later, survivors may be highly 'synchronized' into either a sensitive cell-cycle stage or transient state preparatory to entering a shorter cell cycle to achieve repopulation. The low translocations yields obtained with longer intervals between treatments may typify the genetic response of the repopulating stem cells.
Assuntos
Cromossomos/efeitos dos fármacos , Cromossomos/efeitos da radiação , Espermatogônias/efeitos dos fármacos , Espermatogônias/efeitos da radiação , Espermatozoides/efeitos dos fármacos , Espermatozoides/efeitos da radiação , Trietilenomelamina/farmacologia , Animais , Relação Dose-Resposta à Radiação , Masculino , Camundongos , Tamanho do Órgão , Testículo/anatomia & histologia , Fatores de Tempo , Translocação Genética , Raios XRESUMO
Previous work has shown that a high yield of genetic damage can be recovered from stem spermatogonia exposed to a high (900 R) X-ray dose, despite extensive cell killing, when this follows 24 h after a smaller (100 R) radiation exposure. This differs from the response of the normal stem-cell population and has been interpreted to mean that the more radio-resistant cells surviving the first exposure become sensitive both to radiation-induced killing and genetic damage after this time interval and, as a consequence, lose the heterogeneity in radio-sensitivity that typifies a normal stem-cell population. Similar results have now been obtained with doses of 600 and 800 R given in fractions of 100 + 500 R and 100 + 700 R 24 h apart. Yields of translocations among spermatocytes were higher than obtained with the single doses and responses consistent with the fractions acting additively were obtained when the fractions were given in reverse order. Further analyses of the data provided support for the concept that 24 h after a radiation exposure there is a loss of heterogeneity in radio-sensitivity in the surviving stem-cell population.
Assuntos
Espermatogônias/efeitos da radiação , Espermatozoides/efeitos da radiação , Translocação Genética , Animais , Sobrevivência Celular , Relação Dose-Resposta à Radiação , Masculino , Camundongos , Tolerância a Radiação , Espermatócitos/efeitos da radiação , Espermatogônias/ultraestrutura , Testículo/efeitos da radiação , Fatores de Tempo , Raios XRESUMO
The spermatogonial stem cells of 101/H mice have been found to be more sensitive to killing by acute X-ray doses than those of the "standard" C3H/HeH X 101/H F1 hybrid. Duration of the sterile period was longer throughout the 0.5-8.0-Gy dose range tested and "recovered" testis weights, taken after recovery of fertility, were more severely reduced. The shapes of the sterile period dose-response curves were similar, but with the 101/H mice the plateau occurred at 3-5 Gy, rather than at 6 Gy. An equivalent observation was made with the testis weight data. The translocation dose-response curve was bell-shaped, as previously found with the hybrid, but yields were lower at all but the lowest doses. Notably, peak yields occurred at 3-5 Gy, rather than at 6 Gy. The altered stem cell killing and genetic responses may be explained either by a higher proportion of radiosensitive cells in the heterogeneous stem cell population or by a higher ratio of cell killing to recoverable chromosome damage which might imply a reduced repair capacity. The latter finds some support in other data. The pattern of genetic response obtained when an X-ray dose was given in two fractions at various intervals was similar in 101/H and the hybrid mice, suggesting that their kinetics of stem cell repopulation following depletion differ little.
Assuntos
Espermatogônias/efeitos da radiação , Espermatozoides/efeitos da radiação , Translocação Genética , Animais , Sobrevivência Celular , Relação Dose-Resposta à Radiação , Hibridização Genética , Masculino , Camundongos , Camundongos Endogâmicos , Especificidade da EspécieRESUMO
The specific-locus mutation yield from 101/H mice following single, acute 6 Gy spermatogonial X-irradiation was significantly higher than both a concurrent C3H/HeH x 101/H F1 hybrid control and historical data obtained with mice of equivalent genotype. There is therefore discord with the reduced translocation response to single X-ray doses previously identified in this strain. By contrast, the mutation yield following a 24-h interval 3 + 3 Gy fractionated X-ray dose was not significantly different from that of its concurrent hybrid control, nor from results obtained by others with mice of the equivalent or different genotypes. Here there is no discord with the translocation response obtained with a 1 + 5 Gy 24-h interval fractionated regime. Appraisal of comparable specific-locus and translocation data indicates that the differing results obtained with the two genetic end-points are not without precedence. This, together with the observation that the responses for cytologically visible deletions and specific-locus mutations are similar, suggests that the latter two events predominantly derive from one-hit events, with translocations deriving from two-hit events and that the probabilities of induction of each type of lesion vary according to the organisation of the nucleus in different phases of the cell cycle.
Assuntos
Mutação , Lesões Experimentais por Radiação/genética , Animais , Genes Dominantes , Masculino , Camundongos , Camundongos Endogâmicos C3H , Tamanho do Órgão , Testículo/patologia , Testículo/efeitos da radiação , Raios XRESUMO
Earlier studies have shown that the spermatogonial stem cells of the mouse testis recovering from previous radiation or chemical mutagen exposure give subnormal yields of genetic damage with subsequent X-irradiation. This response has been investigated further: (a) with a high, 9-Gy X-ray dose given 4, 12 or 21 days after a 1-Gy conditioning dose (Expt. 1), and (b) with a 1 + 7-Gy, 24-h fractionation regime given 4 or 14 days after a 1-Gy conditioning dose (Expt. 2). In Expt. 1 the 1 + 9-Gy, 4-day interval regime gave a very low response, lower than obtained previously with an equivalent 1 + 5-Gy treatment. This suggests that a heterogeneity in radiosensitivity, such as exists in unirradiated stem cell populations and absent 24-48 h after radiation depletion, is quickly re-established among the stem cells repopulating the testis. By contrast, the 1 + 7-Gy, 24-h fractionation when given 4 days after the 1-Gy conditioning dose (Expt. 2) gave a very high yield of genetic damage, almost as high as that given by the fractionated (1 + 7 Gy) dose applied to previously unirradiated stem cells. This suggests that the newly established heterogeneity is removed by the second 1-Gy conditioning dose. With longer intervals between treatments, genetic yields consistent with additivity were obtained in Expt. 1; less clear results were obtained Expt. 2. Comparison with earlier data generally suggested that the duration of the repopulating period is dose-dependent. In a third experiment evidence was obtained that genetic damage induced by X-irradiation can be reduced by a subsequent treatment with triethylenemelamine (TEM) during the repopulating phase. This confirmed an earlier finding. Such an interaction could not be demonstrated with two X-ray treatments. An explanation for the X-ray/TEM interaction is offered.
Assuntos
Espermatogênese/efeitos da radiação , Espermatogônias/efeitos da radiação , Espermatozoides/efeitos da radiação , Testículo/efeitos da radiação , Animais , Relação Dose-Resposta à Radiação , Fertilidade/efeitos da radiação , Masculino , Camundongos , Tamanho do Órgão/efeitos da radiação , Testículo/citologia , Translocação Genética/efeitos da radiação , Raios XRESUMO
The specific-locus mutation frequency obtained from mouse spermatogonial stem cells following unequal, 1 + 9 Gy X-ray fractionation with a 24-h fractionation interval is low, and consistent with the two fractions acting additively. The response is therefore markedly different from the augmented mutation frequencies obtained with 500 + 500 R and 100 + 500 R, 24-h fractionations. The lower yield compared with the 100 + 500 R response also indicates a clear difference from the translocation data which demonstrate increases in yield with increasing second dose over the same dose range. The decline in specific locus mutation yield with the increase in the second dose from 500 R to 9 Gy suggests that the stem cells surviving the first fraction are heterogeneous in their sensitivities to this class of genetic damage. A similar, additive specific locus mutation frequency is obtained with unequal, 1 + 9 Gy X-irradiation when the interval between fractions is 4 days. This is consistent with 500 + 500 R, 4-day and 7-day interval responses obtained previously but again differs from the sub-additive translocation responses obtained with such X-ray fractionation. Taken together with the data from previous studies the present results suggest that (1) 24 h after the first fraction, (a) the surviving stem cell have two components; survivors of the formerly radiosensitive, cycling component of the normal stem cell population and the formerly radioresistant, G0 or arrested G1 cells, which are being 'triggered' into a rapid cell cycle to achieve repopulation of the testis; (b) these two components are of near-equal sensitivity to translocation induction and cell killing, hence the additive translocation yields with equal X-ray fractionations and yields consistent with those extrapolated from lower doses with higher, unequal fractionations, e.g. 1 + 7 Gy, 1 + 9 Gy; but (c) the formerly radioresistant, triggered component is much more sensitive than the surviving cycling component to specific locus mutation and cell killing, hence the augmented mutation response with 500 + 500 R fractionation and the drop in yield with 1 + 9 Gy compared with 100 + 500 R X-irradiation.(ABSTRACT TRUNCATED AT 400 WORDS)
Assuntos
Mutação , Espermatogônias/efeitos da radiação , Espermatozoides/efeitos da radiação , Animais , Biometria , Ciclo Celular , Relação Dose-Resposta à Radiação , Masculino , Camundongos , Tolerância a Radiação , Células-Tronco/efeitos da radiação , Fatores de TempoRESUMO
Data that demonstrate how the biology of spermatogenesis plays an important role in determining the yield of genetic damage from ionizing radiation are briefly reviewed. It is suggested that for valid extrapolations of data from mouse mutation experiments to man detailed knowledge of the spermatogonial stem cell systems in the two species is required. Two new sets of mouse specific mutation data are presented. (1) When a 2 mg/kg dose of triethylenemelamine (TEM) was used as a conditioning dose and followed 24 h later by 6 Gy X-rays, the mutation yield from spermatogonial stem cells was over twice as high (30.20 X 10(-5)/locus/gamete) as that when the X-ray dose was given alone (13.75 X 10(-5)/locus/gamete). No such effect was found when the TEM was given only 3 h prior to the X-irradiation. Since TEM at the dose used is inefficient at inducing specific-locus mutations, an augmentation of the X-ray response is indicated. It has therefore been concluded that the augmented mutation responses obtained with equal 24 h X-ray fractionations at high doses are attributable to mutation induction by the second dose. The responsive cells would be the formerly resistant component of the stem cell population that had survived the TEM treatment and that had been 'triggered' into a radiosensitive phase by the population depletion. (2) When 2 doses of 500 mg/kg hydroxyurea (HU) were given 3 h apart 3 h prior to 6 Gy X-rays to reduce the numbers of stem cells in the S and G2 phases of the cell cycle exposed to the radiation, the mutation responses was greatly enhanced to a level that is the highest yet recorded per unit X-ray dose (7.10 X 10(-5)/locus/gamete/Gy). No such effect was obtained when the intervals between the HU and X-ray treatments were either shorter (less than 0.5 h) or longer (24 h). It was concluded that X-ray-induced specific-locus mutations derive principally from stem cells in the G1 phase of the cell cycle. The reasons why the X-ray-induced mutation-yields from repopulating stem cells (with a short cell cycle and, hence, short G1 phase) are similar to those from undamaged stem cell populations, in contrast to translocation yields, therefore remains unresolved.
Assuntos
Mutação , Espermatogônias/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Animais , Hemoglobinas/genética , Humanos , Infertilidade/genética , Tamanho da Ninhada de Vivíparos , Masculino , Camundongos , Camundongos Endogâmicos C3H , Mutagênicos/farmacologia , Doses de Radiação , Espermatogônias/efeitos da radiação , Translocação Genética , Raios XRESUMO
Two new genetic methods for detecting autosomal non-disjunction and chromosome loss in mice are described. Both methods involve the use of marker genes and Robertsonian translocations, the latter present only in tester parents, to detect events in chromosomally normal mice. With the Rb method, the tester parent carries one or more Robertsonian translocations heterozygously; with the MBH method the tester parent carries two Robertsonian translocations showing monobrachial homology. The high rates of meiotic non-disjunction in the tester mice provide gametes with specific extra or missing chromosomes which, at fertilization, can allow the survival of a proportion of the zygotes lacking or carrying an extra specific chromosome from tested chromosomally normal parents. The Rb method has been assessed for X-ray-induced chromosome 1 loss and non-disjunction in mature oocytes and also for such chromosome 1 loss from the maternal pronuclei of 1-cell zygotes. The MBH method has been assessed for X-ray-induced chromosome 1 loss in male postmeiotic cells and for non-disjunction in spermatocytes. Both methods proved effective in detecting chromosome 1 loss. A single case of the much rarer non-disjunctional event was also found. As applied, both methods compared favourably with the numerical sex chromosome anomaly (NSA) method and have considerable potential for further development.