Altered response of progeria fibroblasts to epidermal growth factor A. COLIGE, B. NUSGENS and CH. M. LAPIERE* Laboratory of Experimental Dermatology, Tour de Pathologie, B23, CHU Sart-Tilman, University of Liege, B-4000 Sart-Tilman, Liege 1, Belgium * Author for correspondence Summary The Hutchinson-Gilford syndrome (progeria) is a rare disorder in childhood characterized by prema- ture and accelerated aging. This study reports the effect of a potent growth factor, EGF, on the proliferative capacities and extracellular matrix macromolecules and collagenase expression of two strains of progeria skin-derived cells. At low popu- lation doubling levels (PDL<10), confluent cultures of progeria fibroblasts made quiescent by lowering the concentration of serum in the medium did not respond to EGF while the mitotic activity of normal PDL-matched fibroblasts was almost maximally re- stored upon addition of EGF. No obvious difference between normal and low PDL progeria fibroblasts was observed in the number and in the affinity of the receptors measured by [ 125 I]EGF binding. The syn- thesis of collagen and non-collagen proteins was similar in normal and affected cells at low and high serum concentration and both types of cells re- sponded to EGF by a specific inhibition of collagen synthesis. Besides a normal level of mRNA coding for type I and type III collagens, collagenase and laminin, progeria fibroblasts expressed a high level of elastin and type IV collagen mRNA. Like normal fibroblasts, progeria cells responded to EGF by a decrease in the level of mRNA for fibrillar collagens and elastin. In contrast, a complete lack of response to EGF was observed for collagenase mRNA whereas the expression of this enzyme was strikingly induced by EGF in normal PDL-matched cells. The abnormal expression of type IV collagen was not significantly modified by EGF. At PDL>10, progeria cells exhibited features of senescence. A significant re- duction of collagen synthesis was observed and no further inhibition by EGF was recorded. Key words: aging, growth factor, collagens, collagenase, elastin, premature aging syndrome. Introduction The Hutchinson-Gilford syndrome (progeria) is a rare disorder of childhood characterized by selected aspects of premature and accelerated aging (Debusk, 1972). The exact mode of inheritance is still a matter of discussion, since some cases most likely occur as the result of a sporadic dominant mutation (Brown et al. 1985; McKusick, 1982) and others as an autosomal recessive heritable event (Mostafa and Gabr, 1954; Goldstein and Moerman, 1975). Affected patients display striking clini- cal similarities such as short stature, scleroderma-like skin, beaked nose, skeletal alterations and cardiovascular abnormalities (Rautenstrauch et al. 1977; Baker et al. 1981). Progeria patients usually die in the second or third decade of respiratory failure or myocardial infarction secondary to acute coronary thrombosis (Debusk, 1972). In vitro studies of skin fibroblasts from affected individuals have demonstrated a growth-potential deficiency (Gold- stein, 1969, 1979; Danes, 1971), low HLA expression (Goldstein et al. 1975), decreased DNA repair capacity (Epstein et al. 1973), increased elastin production (Sephel et al. 1988) and elevated c-myc gene expression (Naka- mura and Hart, 1988). We recently described a new case of progeria expressing increased levels of elastin and mostly type IV collagen mRNAs (Colige et al. 1991). The etiology of progeria is not yet clear. However, as senescent cells are characterized by alterations in the Journal of Cell Science 100, 649-655 (1991) Printed in Great Britain © The Company of Biologists Limited 1991 responsiveness to growth factors in vitro (Carlin et al. 1983; Phillips et al. 1984; Cristofalo et al. 1989), a better knowledge of the influence of growth factors on progeria fibroblasts should help in understanding the mechanisms underlying this disease. A number of growth factors are involved in the regulation of cellular growth and differen- tiation. Among these, epidermal growth factor (EGF) plays an important role at different levels of cellular functions, modulating, for example, cellular proliferation and migration (Watanabe et al. 1987), the biosynthetic capacity of fibroblasts (Hata etal. 1988; Colige etal. 1990), the expression of collagenase through c-fos- and c-jun- dependent activation (Angel et al. 1987), and the wound healing process ire vivo (Nanney, 1990; Franklin et al. 1986). As we had recently detected a new case of progeria (Colige et al. 1991), it was of interest to investigate the effect of EGF in vitro on the phenotype and the growth capacities of the skin-derived cells of this patient. Materials and methods FCS was from Gibco, [5- 3 H]proline, [ 3 H]thymidine, [ 32 P]dCTP and [ 3 H]Enhancer were from New England Nuclear, bis- benzimide H33258 from Hoechst, acrylamide, n, rc'-methylene- bis-acrylamide and AWW.AT-tetramethylethylenediamine from Biorad Laboratories, X-ray films Royal X-S-0 Mat from Kodak and [ 125 I]EGF from Amersham. The random priming DNA labeling kit was from Boehringer. 649