Premature senescence is induced in human diploid fibroblasts between day 2 and 3 after treatment with hydrogen peroxide at sublethal concentration. This phenotype is namely characterized by a spread out morphology, irreversible growth arrest and senescence associated ß-galactosidase activity. Previous studies suggested that protein neosynthesis is important for establishment of the flattened, widened and irregular morphology of the senescent cells induced by H2O2. Moreover, it was shown that the neutralization of TGF-ß1 by specific antibodies alters this morphology induced by treatment with H2O2. Since the mechanisms involved in senescent morphogenesis are not well understood, we studied the TGF-ß1-dependent and -independent neosynthesis of cytoplasmic proteins, between the days 2 and 3 after treatment of fibroblasts with H2O2 and identified proteins involved in the organization of cytoskeleton: ezrin, caldesmon and HSP27. Rac1, Cdc42 and caveolin 1 were defined in the literature as proteins involved in the regulation of replicative senescent morphology and in the senescent morphology induced by treatment of fibroblasts with H2O2 (in the case of caveolin 1). We studied activation of Rac1 and Cdc42, the phosphorylation of caveolin 1 after treatment with H2O2 et effect of invalidation of these 3 proteins and p38áMAPK on fibroblasts morphology. Moreover, we showed that Cdc42 could activate p38áMAPK and vice-versa, during the H2O2 treatment. In parallel of this study, we tried to analyze the differential abundance of membrane proteins at day 3 after the treatment of fibroblasts with H2O2. This allowed to study further a protein associate with the membranes and actin cytoskeleton: annexin 2. This work allows a better understanding of mechanisms whereby fibroblasts enter premature senescence, and more particularly senescent morphogenesis.
Sénescence prématurée induite par un stress au peroxyde d’hydrogène chez des fibroblastes diploïdes de poumon humain: Etude de la néosynthèse protéique, de marqueurs associés aux membranes et du rôle de Cdc42, Rac1 et p38MAPK
Chrétien, A. (Author). 21 Apr 2008
Student thesis: Doc types › Doctor of Sciences