RESUMO
Gut microbiome development affects infant health and postnatal physiology. The gut microbe assemblages of preterm infants have been reported to be different from that of healthy term infants. However, the patterns of ecosystem development and inter-individual differences remain poorly understood. We investigated hospitalised preterm infant gut microbiota development using 16S rRNA gene amplicons and the metabolic profiles of 268 stool samples obtained from 17 intensive care and 42 term infants to elucidate the dynamics and equilibria of the developing microbiota. Infant gut microbiota were predominated by Gram-positive cocci, Enterobacteriaceae or Bifidobacteriaceae, which showed sequential transitions to Bifidobacteriaceae-dominated microbiota. In neonatal intensive care unit preterm infants (NICU preterm infants), Staphylococcaceae abundance was higher immediately after birth than in healthy term infants, and Bifidobacteriaceae colonisation tended to be delayed. No specific NICU-cared infant enterotype-like cluster was observed, suggesting that the constrained environment only affected the pace of transition, but not infant gut microbiota equilibrium. Moreover, infants with Bifidobacteriaceae-dominated microbiota showed higher acetate concentrations and lower pH, which have been associated with host health. Our data provides an in-depth understanding of gut microbiota development in NICU preterm infants and complements earlier studies. Understanding the patterns and inter-individual differences of the preterm infant gut ecosystem is the first step towards controlling the risk of diseases in premature infants by targeting intestinal microbiota.
Assuntos
Fezes/microbiologia , Microbioma Gastrointestinal , Cocos Gram-Positivos/classificação , Unidades de Terapia Intensiva , Acetatos/análise , Bifidobacterium/classificação , Bifidobacterium/isolamento & purificação , Enterobacteriaceae/classificação , Enterobacteriaceae/isolamento & purificação , Feminino , Cocos Gram-Positivos/isolamento & purificação , Hospitalização , Humanos , Concentração de Íons de Hidrogênio , Recém-Nascido , Recém-Nascido Prematuro , Masculino , Metaboloma , RNA Ribossômico 16S/genética , Staphylococcaceae/classificação , Staphylococcaceae/isolamento & purificaçãoRESUMO
Primary graft failure (pGF) is associated with considerable morbidity and mortality. Salvage hematopoietic SCT (HSCT) can rescue pGF patients; however, the optimal preconditioning regimen and stem cell source are yet to be determined, particularly in children. In this study, we retrospectively analyzed 102 pediatric patients who received salvage allogeneic HSCT for pGF. Salvage HSCT from matched or one-Ag-mismatched related donors (rMM01) provided superior OS compared with that from two- or three-Ags-mismatched related donors (rMM23) or cord blood transplantation (CBT). CBT showed a trend toward a slightly lower engraftment rate and late engraftment achievement compared with rMM23; however, the OS rate was similar between the two groups (47.6±7.7% for rMM23 and 45.7±8.6% for CBT, at 1 year after salvage HSCT). Multivariate analysis showed that preconditioning regimens with fludarabine or irradiation were associated with a higher engraftment rate and those with alkylating agents were associated with better OS. In conclusion, our results showed that rMM01 was the most suitable donor for salvage HSCT for pediatric pGF, and that CBT was an equally important option compared with rMM23 for patients without rMM01.
Assuntos
Rejeição de Enxerto/cirurgia , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Transplante de Células-Tronco Hematopoéticas/métodos , Terapia de Salvação/métodos , Condicionamento Pré-Transplante/métodos , Adolescente , Criança , Pré-Escolar , Sobrevivência de Enxerto , Humanos , Lactente , Prognóstico , Fatores de Risco , Taxa de Sobrevida , Transplante Homólogo , Resultado do TratamentoRESUMO
MicroRNAs (miRNAs) regulate cell proliferation and differentiation by controlling the expression of proteins involved in many signaling pathways. Recent studies have shown that dysregulation of miRNA expression is associated with increased tumorigenicity and a poor prognosis in several types of cancers. The miRNA let-7b is one of the severely downregulated miRNAs in mixed-lineage leukemia (MLL)-rearranged acute lymphoblastic leukemia (ALL) patients. In vitro transfection of leukemogenic MLL fusion genes into human embryonic kidney-293 cells suppressed let-7b expression. In leukemic cells with an MLL fusion gene, the regulatory region for let-7b expression was hypermethylated, and its expression was partially recovered after culturing the cells with the demethylating agent 5-azacitidine. These results suggest that loss of let-7b expression may be one of the consequences of oncogenic MLL fusion proteins, and contributes to leukemogenesis possibly through the upregulation of let-7b-regulated target genes with leukemogenic potential in hematopoietic cells. The enforced expression of let-7b in ALL cell lines with an MLL fusion gene inhibited their growth, indicating the possible use of let-7b as a new therapeutic tool for refractory infant ALL with an MLL fusion gene.
Assuntos
Metilação de DNA , Regulação Leucêmica da Expressão Gênica , Rearranjo Gênico , MicroRNAs/genética , Proteína de Leucina Linfoide-Mieloide/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/genética , Antimetabólitos Antineoplásicos/farmacologia , Azacitidina/farmacologia , Sequência de Bases , Proliferação de Células , Histona-Lisina N-Metiltransferase , Humanos , Lactente , Dados de Sequência Molecular , Proteínas de Fusão Oncogênica/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Regiões Promotoras Genéticas/genética , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais CultivadasRESUMO
PURPOSE: 5-chloro-2,4-dihydroxypyridine (gimeracil) is a component of the oral fluoropyrimidine derivative S-1. Gimeracil was originally added to S-1 to yield prolonged 5-fluorouracil (5-FU) concentrations in serum and tumor tissues by inhibiting dihydropyrimidine dehydrogenase, which degrades 5-FU. We previously demonstrated that gimeracil enhances the efficacy of radiotherapy through the suppression of homologous recombination (HR) in DNA double strand repair. The goal of this paper was to examine the effects of gimeracil on the sensitivity of anticancer drugs and hyperthermia in order to obtain effective radiosensitization. MATERIALS AND METHODS: Various cell lines, including DLD 1 (human colon carcinoma cells) and cells deficient in HR or nonhomologous end-joining (NHEJ), were used in clonogenic assays. The survival of these cells after various treatments (e.g., drug treatment, heat treatment, and radiation) was determined based on their colony-forming ability. RESULTS: Gimeracil enhanced cell-killing effects of camptothecin (CPT), 5-FU, and hydroxyurea. Gimeracil sensitized effects of CPT or 5-FU to cells deficient in HR or NHEJ to a similar extent as in other cells (DLD1 and a parent cell), indicating that its sensitizing mechanisms may be different from inhibition of HR or NHEJ. Combination of gimeracil and CPT or 5-FU sensitized radiation more effectively than each modality alone. Gimeracil also enhanced heat sensitivity at 42°C or more. The degree of heat sensitization with gimeracil increased as the temperature increased, and the combination of gimeracil and heat-sensitized radiation was more effective than each modality alone. CONCLUSION: Gimeracil enhanced sensitivity of CPT, 5-FU, and hyperthermia. Combination of these modalities sensitized radiation more efficiently than each modality alone.
Assuntos
Antineoplásicos/farmacologia , Temperatura Alta , Piridinas/farmacologia , Tolerância a Radiação/efeitos dos fármacos , Radiossensibilizantes/farmacologia , Raios X , Animais , Células CHO , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Cricetinae , Cricetulus , Hipertermia InduzidaRESUMO
Transient leukemia (TL) has been observed in approximately 10% of newborn infants with Down syndrome (DS). Although treatment with cytarabine is effective in high-risk TL cases, approximately 20% of severe patients still suffer early death. In this study, we demonstrate abundant KIT expression in all 13 patients with GATA1 mutations, although no significant difference in expression levels was observed between TL and acute myeloid leukemia. Stem cell factor (SCF) stimulated the proliferation of the TL cells from five patients and treatment with the tyrosine kinase inhibitor imatinib suppressed the proliferation effectively in vitro. To investigate the signal cascade, we established the first SCF-dependent, DS-related acute megakaryoblastic leukemia cell line, KPAM1. Withdrawal of SCF or treatment with imatinib induced apoptosis of KPAM1 cells. SCF activated the RAS/MAPK and PI3K/AKT pathways, followed by downregulation of the pro-apoptotic factor BIM and upregulation of the anti-apoptotic factor MCL1. Although we found novel missense mutations of KIT in 2 of 14 TL patients, neither mutation led to KIT activation and neither reduced the cytotoxic effects of imatinib. These results suggest the essential role of SCF/KIT signaling in the proliferation of DS-related leukemia and the possibility of therapeutic benefits of imatinib for TL patients.
Assuntos
Proliferação de Células , Síndrome de Down/complicações , Leucemia/patologia , Transdução de Sinais/fisiologia , Fator de Células-Tronco/fisiologia , Apoptose/efeitos dos fármacos , Benzamidas , Linhagem Celular Tumoral , Feminino , Fator de Transcrição GATA1/genética , Humanos , Mesilato de Imatinib , Lactente , Recém-Nascido , Leucemia/etiologia , Masculino , Mutação , Piperazinas/farmacologia , Piperazinas/uso terapêutico , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Fator de Células-Tronco/análise , Fator de Células-Tronco/genéticaRESUMO
Nijmegen breakage syndrome shares several common cellular features with ataxia telangiectasia, including chromosomal instability and aberrant S- and G2-phase checkpoint regulation. We show here that after irradiation, NBS1 interacts physically with both BRCA1 and SMC1, a component of the cohesin complex, and that their interactions are completely abolished in AT cells. It is noted that BRCA1 is required for the interaction of NBS1 with SMC1, whereas the reverse is not the case, since BRCA1 is able to bind to NBS1 in the absence of an NBS1/SMC1 interaction as observed in MRE11- or RAD50-deficient cells. This indicates that ATM and BRCA1 are upstream of the NBS1/SMC1 interaction. Furthermore, the interaction of NBS1 with SMC1 requires both conserved domains of NBS in the N-terminus and the C-terminus, since they are indispensable for binding of NBS1 to BRCA1 and to MRE11/ATM, respectively. The interaction of NBS1 with SMC1 and the resulting phosphorylation are compromised in the clones lacking either the N- or C-terminus of NBS1, and as a consequence, chromatid-type aberrations are enhanced after irradiation. Our results reveal that ATM plays a fundamental role in promoting the radiation-induced interaction of NBS1 with SMC1 in the presence of BRCA1, leading to the maintenance of chromosomal integrity.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Cromátides/genética , Cromátides/efeitos da radiação , Proteínas Cromossômicas não Histona/metabolismo , Aberrações Cromossômicas , Proteínas de Ligação a DNA/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Lesões por Radiação/genética , Transdução de Sinais/fisiologia , Proteínas Supressoras de Tumor/metabolismo , Proteínas Mutadas de Ataxia Telangiectasia , Células Cultivadas , Relação Dose-Resposta à Radiação , Humanos , Doses de Radiação , Transdução de Sinais/efeitos da radiaçãoRESUMO
The proteins responsible for radiation sensitive disorders, NBS1, kinase ataxia-telangiectasia-(A-T)-mutated (ATM) and MRE11, interact through the C-terminus of NBS1 in response to the generation of DNA double-strand breaks (DSBs) and are all implicated in checkpoint regulation and DSB repair, such as homologous recombination (HR). We measured the ability of several NBS1 mutant clones and A-T cells to regulate HR repair using the DR-GFP or SCneo systems. ATM deficiency did not reduce the HR repair frequency of an induced DSB, and it was confirmed by findings that HR frequencies are only slightly affected by deletion of ATM-binding site at the extreme C-terminus of NBS1. In contrast, The HR-regulating ability is dramatically reduced by deletion of the MRE11-binding domain at the C-terminus of NBS1 and markedly inhibited by mutations in the FHA/BRCT domains at the N-terminus. This impaired capability in HR is consistent with a failure to observe MRE11 foci formation. Furthermore, normal HR using sister chromatid was completely inhibited by the absence of FHA/BRCT domains. These results suggested that the N- and C-terminal domains of NBS1 are the major regulatory domains for HR pathways, very likely through the recruitment and retention of the MRE11 nuclease to DSB sites in an ATM-independent fashion.
Assuntos
Proteínas de Ciclo Celular/genética , Reparo do DNA/genética , Proteínas de Ligação a DNA/genética , Proteínas Nucleares/genética , Proteínas Serina-Treonina Quinases/genética , Recombinação Genética , Proteínas Supressoras de Tumor/genética , Adenina , Proteínas Mutadas de Ataxia Telangiectasia , Quebras de DNA de Cadeia Dupla , Fibroblastos/fisiologia , Frequência do Gene , Humanos , TiminaRESUMO
Nijmegen breakage syndrome (NBS) is a rare recessive genetic disorder, characterized by bird-like facial appearance, early growth retardation, congenital microcephaly, immunodeficiency and high frequency of malignancies. NBS belongs to the so-called chromosome instability syndromes; in fact, NBS cells display spontaneous chromosomal aberrations and are hypersensitive to DNA double-strand break-inducing agents, such as ionizing radiations. NBS1, the gene underlying the disease, is located on human chromosome 8q21. The disease appears to be prevalent in the Eastern and Central European population where more than 90% of patients are homozygous for the founder mutation 657del5 leading to a truncated variant of the protein. NBS1 forms a multimeric complex with MRE11/RAD50 nuclease at the C-terminus and retains or recruits them at the vicinity of sites of DNA damage by direct binding to histone H2AX, which is phosphorylated by PI3-kinase family, such as ATM, in response to DNA damage. Thereafter, the NBS1-complex proceeds to rejoin double-strand breaks predominantly by homologous recombination repair in vertebrates. NBS cells also show to be defective in the activation of intra-S phase checkpoint. We review here some cellular and molecular aspects of NBS, which might contribute to the clinical symptoms of the disease.
Assuntos
Proteínas de Ciclo Celular/genética , Síndrome de Quebra de Nijmegen/genética , Proteínas Nucleares/genética , Ciclo Celular , Instabilidade Cromossômica , Dano ao DNA , Reparo do DNA , Efeito Fundador , Histonas/metabolismo , Homozigoto , Humanos , Modelos Biológicos , Modelos Genéticos , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , População BrancaRESUMO
It is well established that cells synchronised at the G1-S phase are highly radiosensitive. In this study, p16-null human glioma cell lines were induced into G1 cell cycle arrest by adenovirus-mediated p16 gene transfer, and examined for radiation-induced cell killing. Clonogenic analysis and trypan blue extraction test showed that the p16 gene transfer enhanced radiation-induced cell killing in p16-null glioma cell lines. TUNEL assays and pulse-field gel electrophoresis confirmed that the radiation-induced cell killing of p16-transfected cells could be caused by a nonapoptotic mechanism. Gimsa staining demonstrated that irradiation alone or Ax-mock infection plus irradiation results in a slight increase in the frequency of cells with abnormal nucleus, compared to unirradiated uninfected or Ax-mock infected cells. However, Ax-hp16 or Ax-hp21 infection alone modestly increased the frequency of cells with abnormal nucleus (especially bi- and multinucleation), and 4-Gy irradiation of Ax-hp16 or Ax-hp21 infected cells substantially enhanced this frequency. These results suggest that there exists some unknown interaction between radiation and p16 in cytoplasm/membranes, which decreases cytokinesis and promotes abnormal nucleation. Thus, p16 expression prevented radiation-induced apoptosis by promoting abnormal nucleation, thereby leading to another mode of cell death.
Assuntos
Núcleo Celular/efeitos da radiação , Técnicas de Transferência de Genes , Genes p16/efeitos da radiação , Glioma/genética , Glioma/patologia , Adenoviridae/genética , Apoptose/efeitos da radiação , Linhagem Celular Tumoral , Núcleo Celular/patologia , Eletroforese em Gel de Campo Pulsado , Vetores Genéticos , Humanos , Marcação In Situ das Extremidades Cortadas , Radiação Ionizante , TransfecçãoRESUMO
The checkpoint kinase Chk2 has a key role in delaying cell cycle progression in response to DNA damage. Upon activation by low-dose ionizing radiation (IR), which occurs in an ataxia telangiectasia mutated (ATM)-dependent manner, Chk2 can phosphorylate the mitosis-inducing phosphatase Cdc25C on an inhibitory site, blocking entry into mitosis, and p53 on a regulatory site, causing G(1) arrest. Here we show that the ATM-dependent activation of Chk2 by gamma- radiation requires Nbs1, the gene product involved in the Nijmegen breakage syndrome (NBS), a disorder that shares with AT a variety of phenotypic defects including chromosome fragility, radiosensitivity, and radioresistant DNA synthesis. Thus, whereas in normal cells Chk2 undergoes a time-dependent increased phosphorylation and induction of catalytic activity against Cdc25C, in NBS cells null for Nbs1 protein, Chk2 phosphorylation and activation are both defective. Importantly, these defects in NBS cells can be complemented by reintroduction of wild-type Nbs1, but neither by a carboxy-terminal deletion mutant of Nbs1 at amino acid 590, unable to form a complex with and to transport Mre11 and Rad50 in the nucleus, nor by an Nbs1 mutated at Ser343 (S343A), the ATM phosphorylation site. Chk2 nuclear expression is unaffected in NBS cells, hence excluding a mislocalization as the cause of failed Chk2 activation in Nbs1-null cells. Interestingly, the impaired Chk2 function in NBS cells correlates with the inability, unlike normal cells, to stop entry into mitosis immediately after irradiation, a checkpoint abnormality that can be corrected by introduction of the wild-type but not the S343A mutant form of Nbs1. Altogether, these findings underscore the crucial role of a functional Nbs1 complex in Chk2 activation and suggest that checkpoint defects in NBS cells may result from the inability to activate Chk2.
Assuntos
Dano ao DNA , Ativação Enzimática , Proteínas Nucleares/metabolismo , Proteínas Quinases/metabolismo , Proteínas Serina-Treonina Quinases , Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular , Núcleo Celular/metabolismo , Quinase do Ponto de Checagem 2 , Ativação Enzimática/efeitos da radiação , Fibroblastos/metabolismo , Raios gama , Deleção de Genes , Humanos , Immunoblotting , Microscopia de Fluorescência , Mitose , Mutação , Fosforilação , Fosfotransferases/metabolismo , Testes de Precipitina , Radiação Ionizante , Fatores de Tempo , Transfecção , Fosfatases cdc25/metabolismoRESUMO
Proteoglycans were localized immunohistochemically in the dermis of Donryu rats, using monoclonal antibodies raised against large proteoglycan (PG-M/versican) and small proteoglycan (decorin). The localizations of these proteoglycans in the dermis were compared between young rats (22-day old) and old ones (24 or 30 months of age), and distinct age differences were observed. In the young dermis, PG-M/versican was observed to be abundant in almost all fibroblastic cells (both cytoplasm and cell processes) whose cellularity was very rich compared with the dermis of old rats. Decorin was only faintly visible in the interstitial fibrous elements of young dermis. In the old dermis, however, decorin was distinctly detected on the fibrous elements, which were diffusely distributed as a fibrous network, and likewise PG-M/versican was visible in only a few fibrous elements which seemed to be the fine processes of fibroblastic cells. In the border layer between epidermis and dermis as well as the basal layer surrounding hair follicles, both large and small proteoglycans could be observed in old dermis. Since decorin, which was abundant in old dermis, has been found to have a growth inhibitory effect, it is conceivable that decorin may be one of the Cell Growth Inhibitory Factors in aging as proposed by Tauchi et al. [17, 18].
Assuntos
Envelhecimento , Proteoglicanas de Sulfatos de Condroitina/análise , Derme/química , Imuno-Histoquímica , Proteoglicanas/análise , Animais , Anticorpos Monoclonais , Decorina , Epiderme/química , Proteínas da Matriz Extracelular , Feminino , Fibroblastos/química , Lectinas Tipo C , Masculino , Ratos , VersicanasRESUMO
BACKGROUND: DNA double-strand breaks (DSB) are the major lethal lesions induced by ionizing radiation. The capability for DNA DSB repair is crucial for inherent radiosensitivity of tumor and normal cells. DNA-PKcs, Ku 70, Ku 85, Xrcc4, and Nbs1 play a critical role in DNA DSB repair. METHODS: We immunohistochemically investigated the expression of DNA-PKcs, Ku 70, Ku85, Xrcc4, and Nbs1 in 134 specimens from various normal and tumor tissues with different radiosensitivity. RESULTS AND CONCLUSION: Immunopositivity to Ku70, Ku85, DNA-PKcs, Xrcc4, and Nbs1 was found in all tumor tissues examined. The staining for Ku70, Ku85, and DNA-PKcs was nuclear; but, for Xrcc4 and Nbs1, it was nuclear and cytoplasmic. There were no apparent differences in the expression of these five proteins among cancerous tissues and the corresponding normal tissues. No apparent differences in nuclear staining intensity were detected in the expression of these five proteins among tumor tissues with different radiosensitivity, although non-Hodgkins' lymphoma (B or T cell) tended to show a lower expression than the others. The stromal cells generally expressed these five proteins at much lower frequency than either tumor or epithelial cells in both tumor and normal tissues.
Assuntos
Antígenos Nucleares , DNA Helicases , Reparo do DNA/genética , DNA de Neoplasias/genética , Proteínas de Ligação a DNA/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Expressão Gênica , Humanos , Autoantígeno Ku , Linfoma não Hodgkin/metabolismo , Linfoma não Hodgkin/radioterapia , Neoplasias/radioterapia , Proteínas Nucleares/metabolismo , Oligopeptídeos/metabolismo , Tolerância a Radiação/genéticaRESUMO
NBS1 (p95), the protein responsible for Nijmegen breakage syndrome, shows a weak homology to the yeast Xrs2 protein at the N terminus region, known as the forkhead-associated (FHA) domain and the BRCA1 C terminus domain. The protein interacts with hMRE11 to form a complex with a nuclease activity for initiation of both nonhomologous end joining and homologous recombination. Here, we show in vivo direct evidence that NBS1 recruits the hMRE11 nuclease complex into the cell nucleus and leads to the formation of foci by utilizing different functions from several domains. The amino acid sequence at 665-693 on the C terminus of NBS1, where a novel identical sequence with yeast Xrs2 protein was found, is essential for hMRE11 binding. The hMRE11-binding region is necessary for both nuclear localization of the complex and for cellular radiation resistance. On the other hand, the FHA domain regulates nuclear foci formation of the multiprotein complex in response to DNA damage but is not essential for nuclear transportation of the complex and radiation resistance. Because the FHA/BRCA1 C terminus domain is widely conserved in eukaryotic nuclear proteins related to the cell cycle, gene regulation, and DNA repair, the foci formation could be associated with many phenotypes of Nijmegen breakage syndrome other than radiation sensitivity.
Assuntos
Núcleo Celular/metabolismo , Reparo do DNA , Proteínas de Ligação a DNA , Endodesoxirribonucleases , Exodesoxirribonucleases , Proteínas Fúngicas/metabolismo , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Proteínas de Saccharomyces cerevisiae , Sequência de Aminoácidos , Animais , Linhagem Celular , Núcleo Celular/efeitos da radiação , Galinhas , Dano ao DNA/efeitos da radiação , Imunofluorescência , Raios gama , Humanos , Substâncias Macromoleculares , Dados de Sequência Molecular , Complexos Multiproteicos , Mutação , Proteínas Nucleares/genética , Estrutura Terciária de Proteína , Alinhamento de Sequência , Transfecção , Técnicas do Sistema de Duplo-HíbridoRESUMO
We analyzed the chromosomal gains and losses that occur in 30 non-small cell lung carcinomas by comparative genomic hybridization. Their chromosomal imbalances showed histological type-specific patterns in adenocarcinomas and in squamous cell carcinomas. The genetic changes in non-small cell lung carcinoma were also strongly dependent on metastasis to lymph node. The average numbers of chromosomal alterations were increased from 6.2 to 9.1 along with the presence of metastasis, and it gave rise to the increased copy number in specific chromosomes. In particular, a novel imbalance at 7p12-21 was recognized in a half of carcinoma with metastasis, although no genetic alteration was observed in 15 non-metastasizing lung carcinoma tested here.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Cromossomos Humanos Par 7/ultraestrutura , DNA de Neoplasias/genética , Neoplasias Pulmonares/genética , Metástase Linfática/genética , Adenocarcinoma/genética , Idoso , Carcinoma de Células Escamosas/genética , Aberrações Cromossômicas , Cromossomos Humanos Par 7/genética , Progressão da Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Hibridização de Ácido NucleicoRESUMO
Life span, which is mainly influenced by pathologic lesions, was compared between specific pathogen-free male Donryu rats fed ad libitum (AL group) and those with dietary restriction (DR group, restricted to 60% of the ad libitum intake). The major age-related lesions observed were pituitary tumor, chronic nephropathy, cardiomyopathy, and myopathy (anterior tibial and masseter muscle). Dietary restriction was effective in slowing the progression of pituitary tumor, chronic nephropathy and myopathy in anterior tibial muscle. Although cardiomyopathy worsened with age, no difference was seen between the AL and DR group. In conclusion, 1) dietary restriction acts to suppress or delay the development of pathologic lesions that occur with age, 2) the onset phase of a pathologic lesion differs with the lesion and organ involved, 3) inhibiting the development of pituitary adenoma and chronic nephropathy can help prolong the life span of rats, 4) for muscle lesions, depending on their anatomical location, the physiological condition of exercise, as well as the relationship with other organs can be involved.
Assuntos
Cardiomiopatias , Dieta Redutora , Falência Renal Crônica , Doenças Musculares , Neoplasias Hipofisárias , Envelhecimento/fisiologia , Animais , Progressão da Doença , Masculino , RatosRESUMO
BACKGROUND: Nijmegen breakage syndrome (NBS), also known as ataxia-telangiectasia (AT) variant, is an autosomal recessive disorder characterized by microcephaly, growth retardation, severe combined immunodeficiency and a high incidence of lymphoid carcinoma, the majority of which are B-cell lymphomas. To determine whether the NBS1 gene is a tumor suppressor gene in B-cell lymphoma, we screened B-cell malignant lymphoma (ML) for any evidence of NBS1 mutation. MATERIALS AND METHODS: Sequence analysis of the NBS1 gene was performed from PCR products amplified from the DNA of 12 extracranial ML or RT-PCR products amplified from cDNA of 8 primary central nervous system lymphoma. RESULTS: Direct sequence analysis revealed that no NBS1 mutations were present in any of these patients. CONCLUSION: The present results suggested that the contribution of NBS1 mutations to B-cell ML was minimal, despite the fact that the NBS1 gene was causative factor in these cases.
Assuntos
DNA de Neoplasias/genética , Linfoma de Células B/genética , Proteínas de Neoplasias/genética , Síndromes Neoplásicas Hereditárias/genética , Proteínas Nucleares/genética , Adolescente , Adulto , Neoplasias Encefálicas/genética , Criança , Pré-Escolar , Códon/genética , Análise Mutacional de DNA , Genes Recessivos , Genes Supressores de Tumor , Humanos , Lactente , Polimorfismo Genético , Polimorfismo Conformacional de Fita Simples , Tolerância a Radiação/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Skin fibroblast cells from two unrelated male infants with a chromosome-instability disorder were analyzed for their response to colcemid-induced mitotic-spindle checkpoint. The infants both had severe growth and developmental retardation, microcephaly, and Dandy-Walker anomaly; developed Wilms tumor; and one died at age 5 mo, the other at age 3 years. Their metaphases had total premature chromatid separation (total PCS) and mosaic variegated aneuploidy. Mitotic-index analysis of their cells showed the absence of mitotic block after the treatment with colcemid, a mitotic-spindle inhibitor. Bromodeoxyuridine-incorporation measurement and microscopic analysis indicated that cells treated with colcemid entered G1 and S phases without sister-chromatid segregation and cytokinesis. Preparations of short-term colcemid-treated cells contained those cells with chromosomes in total PCS and all or clusters of them encapsulated by nuclear membranes. Cell-cycle studies demonstrated the accumulation of cells with a DNA content of 8C. These findings indicate that the infants' cells were insensitive to the colcemid-induced mitotic-spindle checkpoint.
Assuntos
Anormalidades Múltiplas/genética , Aneuploidia , Cromátides/patologia , Fragilidade Cromossômica/genética , Segregação de Cromossomos/genética , Mosaicismo/genética , Fuso Acromático/patologia , Anormalidades Múltiplas/patologia , Células Cultivadas , Pré-Escolar , Cromátides/efeitos dos fármacos , Cromátides/genética , Segregação de Cromossomos/efeitos dos fármacos , DNA/biossíntese , Demecolcina/farmacologia , Fibroblastos , Citometria de Fluxo , Transtornos do Crescimento/genética , Transtornos do Crescimento/patologia , Humanos , Lactente , Masculino , Metáfase/efeitos dos fármacos , Índice Mitótico/efeitos dos fármacos , Mosaicismo/patologia , Membrana Nuclear/efeitos dos fármacos , Pele/patologia , Fuso Acromático/efeitos dos fármacos , Síndrome , Tumor de Wilms/genética , Tumor de Wilms/patologiaRESUMO
Nijmegen breakage syndrome (NBS) is a rare autosomal recessive disorder characterized by microcephaly, combined immunodeficiency, and a high incidence of lymphoid tumor. Cells from NBS patients show chromosomal instability, hypersensitivity to ionizing radiation and abnormal p53-mediated cell cycle regulation. We cloned the underlying gene for NBS, designated NBS1, by complementation-assisted positional cloning from the candidate region 8q21. Large genomic sequencing, as well as a search using computer programs, provides a powerful approach for identifying the underlying gene for a disease. The NBS1 gene encodes a protein of 754 amino acids that has FHA and BRCT domains which often are conserved in cell-cycle checkpoint proteins. The gene has weak homology to the yeast (Saccharomyces cerevisiae) Xrs2 protein in the N-terminus region. Like yeast Xrs2, the NBS1 protein forms a complex with hRAD50/hMRE11, and the complex is condensed as foci in the nucleus after irradiation, indicative that this triple-complex is a crucial factor in DNA repair. Functional analysis of the NBS1 protein is in progress and it should provide further clues to understanding the repair mechanism of radiation-induced DNA double-strand breaks.
Assuntos
Ataxia Telangiectasia/genética , Cromossomos Humanos Par 8/genética , Clonagem Molecular , Reparo do DNA/genética , Genes Recessivos , Microcefalia/genética , Síndromes Neoplásicas Hereditárias/genética , Proteínas Nucleares/genética , Tolerância a Radiação/genética , Proteínas de Saccharomyces cerevisiae , Imunodeficiência Combinada Severa/genética , Sequência de Aminoácidos , Ataxia Telangiectasia/classificação , Proteínas de Ciclo Celular/química , Linhagem Celular Transformada , Cromossomos Humanos/efeitos da radiação , Cromossomos Humanos/ultraestrutura , Etnicidade/genética , Proteínas Fúngicas/genética , Teste de Complementação Genética , Humanos , Dados de Sequência Molecular , Proteínas Nucleares/fisiologia , Polônia/epidemiologia , Estrutura Terciária de Proteína , Saccharomyces cerevisiae/genética , Homologia de Sequência de Aminoácidos , SíndromeRESUMO
PURPOSE: To investigate how DNA-dependent protein kinase (DNA-PK) contributes to p53-dependent signal transduction after heat shock, thermosensitivity and accumulation of p53 and hsp72 after heat shock in lung fibroblasts derived from the SCID mouse were analysed. MATERIALS AND METHODS: Thermosensitivity at 44 degrees C in colony-forming units and Western blot analysis of p53 and hsp72 were analysed. RESULTS: The results indicated that (1) the thermosensitivity at 44 degrees C of SCID cells was higher than that of parental cells and (2) heat-induced accumulation of p53 and hsp72 was abolished and suppressed in SCID cells as compared with that in parental cells respectively. CONCLUSIONS: The findings suggest that the catalytic subunit of DNA-PK may play an important role upstream of p53 and hsp72, which are possible determinants of cellular thermosensitivity.