RESUMO
Cellular DNA damage responses (DDRs) are induced by unrepaired DNA lesions and constitute a protective back-up system that prevents the expansion of damaged cells. These cellular signaling pathways trigger either growth arrest or cell death and are believed to be major components of an early anti-cancer barrier. Cultures of C57BL/6J keratinocytes with various defects in NER sub-pathways allowed us to follow the kinetics of DDRs in an isogenic background and in the proper (physiologically relevant) target cells, supplementing earlier studies in heterogenic human fibroblasts. In a series of well-controlled parallel experiments we have shown that, depending on the NER deficiency, murine keratinocytes elicited highly selective DDRs. After a dose of UV-B that did not affect wild-type keratinocytes, Xpa(-/-) keratinocytes (complete NER deficiency) showed a rapid depletion of DNA replicating S-phase cells, a transient increase in quiescent S-phase cells (not replicating DNA), followed by massive apoptosis. Csb(-/-) keratinocytes (TC-NER deficient) responded by a more sustained increase in QS-phase cells and appeared more resistant to UV-B induced apoptosis than Xpa(-/-). In irradiated Xpc(-/-) keratinocytes (GG-NER deficient) the loss of replicating S-phase cells was associated with a gradual build-up of both QS-phase cells and cells arrested in late-S phase, in complete absence of apoptosis. Our analysis complements and extends previous in vivo investigations and highlights both similarities and differences with earlier fibroblast studies. In vitro cultures of murine keratinocytes provide a new tool to unravel the molecular mechanisms of UV-induced cellular stress responses in great detail and in a physiologically relevant background. This will be essential to fully appreciate the implications of DDRs in tumor suppression and cancer prevention.
Assuntos
Dano ao DNA/genética , Enzimas Reparadoras do DNA/genética , Reparo do DNA/genética , Proteínas de Ligação a DNA/genética , Queratinócitos/patologia , Neoplasias Cutâneas/genética , Raios Ultravioleta , Proteína de Xeroderma Pigmentoso Grupo A/genética , Animais , Apoptose/genética , Apoptose/efeitos da radiação , Morte Celular/efeitos da radiação , Divisão Celular/efeitos da radiação , Células Cultivadas , Dano ao DNA/fisiologia , Reparo do DNA/fisiologia , Enzimas Reparadoras do DNA/fisiologia , Enzimas Reparadoras do DNA/efeitos da radiação , Replicação do DNA/efeitos da radiação , Proteínas de Ligação a DNA/fisiologia , Proteínas de Ligação a DNA/efeitos da radiação , Predisposição Genética para Doença , Queratinócitos/metabolismo , Queratinócitos/efeitos da radiação , Camundongos , Proteínas de Ligação a Poli-ADP-Ribose , Tolerância a Radiação , Fase S/genética , Fase S/efeitos da radiação , Transdução de Sinais/genética , Transdução de Sinais/efeitos da radiação , Neoplasias Cutâneas/etiologia , Neoplasias Cutâneas/patologia , Proteína de Xeroderma Pigmentoso Grupo A/fisiologia , Proteína de Xeroderma Pigmentoso Grupo A/efeitos da radiaçãoRESUMO
The association of variants of genes encoding interleukin (IL)-4 and the IL-4 receptor alpha chain (IL-4Ralpha) with respiratory syncytial virus (RSV) bronchiolitis was examined in hospitalized infants. Polymorphisms in IL-4 (C-590T) and IL-4Ralpha (I50V and Q551R) were genotyped by restriction fragment-length polymorphism analysis. Control subjects included parents of the hospitalized children (for the transmission/disequilibrium test), and a random population sample (for the case-control study). Results were also analyzed in a combination of these 2 tests, using Fisher's method. The IL-4 590T allele was found more frequently among children hospitalized with RSV than expected in the case-control (odds ratio [OR], 1.43; P=.04) and combination (OR, 1.41; P=.02) tests. Among children who were >6 months old when they were hospitalized, compared with the control group or with the <6 months old who were hospitalized for RSV infection, higher frequencies of both the IL-4 590T allele and the IL-4Ralpha R551 allele were found. These results indicate that gain-of-function variants of T helper type 2 cytokine genes may play a role in increasing the severity of RSV disease, which appears more pronounced after the first half-year of life.