Differences in oxidative damage, as measured by an increase in the carbonylation of macromolecules, were determined in situ with skin biopsies from psoriatic patients and controls. High levels of carbonyl residues were consistently detected in the dermis and never in the epidermis of sections of these skin biopsy samples. The dermis of psoriatic skin without lesions had a higher level of carbonylation than the dermis of normal skin. In this study, we found that there was more oxidative damage in cultured fibroblasts prepared from skin with and without lesions from psoriasis patients than in normal fibroblasts from the skin of age-matched controls. The extent of protein carbonylation in cell extracts was determined by immunoblotting, using an antidinitrophenylhydrazone antibody, and in intact cells was determined by immunocytochemical analysis with the same antibody. The higher level of carbonylation detected was used here as a measure of oxidative stress, and showed that some oxidative damage occurred before the appearance of typical psoriatic plaques. These results suggest that fibroblasts are affected before the onset of psoriasis and that this damage is independent of any inflammatory infiltrate.
Oxygen free radicals may act as second messengers in signal transduction pathways and contribute to inflammatory diseases. We studied the action in vitro of radiolytically generated hydroxyl radicals (
Several studies indicate that dermal fibroblasts have a specific role in the pathophysiology of psoriasis. We have previously found that cultured fibroblasts from psoriatic patients are hyperproliferative and have low cyclic AMP-dependent protein kinase activity. In this study, we observed that these cells are also larger than normal. Given the key role of mitogen-activated protein kinases (MAPK) in the regulation of cell proliferation and cytoskeleton function, we characterized MAPK in psoriatic fibroblasts and in normal fibroblasts. Serum and platelet-derived growth factor treatment of serum-deprived fibroblasts led to a larger increase in MAPK activity in psoriatic cells than in normal cells. We then purified MAPK by ion-exchange chromatography. MAPK activity was again found to be significantly higher in psoriatic fibroblasts than in normal cells, both when deprived of serum (p < 0.01) and when stimulated with serum (p < 0.05). Interestingly, 8-bromo-cAMP treatment inhibited serum-stimulated MAPK phosphorylation in normal fibroblasts but had no effect in psoriatic fibroblasts. We observed a temporal variation in nuclear localization of phosphorylated MAPK in cultured fibroblasts stimulated by either serum or platelet-derived growth factor. No difference in the localization of phosphorylated MAPK in normal and psoriatic skins was found. Psoriatic fibroblasts are the first example of a MAPK pathway abnormality in large human benign hyperproliferative cells.
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