To gain insight into the molecular mechanisms underlying cutaneous wound repair, we performed a large scale screen to identify novel injury-regulated genes. Here we show a strong up-regulation of the RNA and protein levels of the two Ca 2؉ -binding proteins S100A8 and S100A9 in the hyperthickened epidermis of acute murine and human wounds and of human ulcers. Furthermore, both genes were expressed by inflammatory cells in the wound. The increased expression of S100A8 and S100A9 in wound keratinocytes is most likely related to the activated state of the keratinocytes and not secondary to the inflammation of the skin, since we also found up-regulation of S100A8 and S100A9 in the epidermis of activin-overexpressing mice, which develop a hyperproliferative and abnormally differentiated epidermis in the absence of inflammation. Furthermore, S100A8 and S100A9 expression was found to be associated with partially differentiated keratinocytes in vitro. Using confocal microscopy, both proteins were shown to be at least partially associated with the keratin cytoskeleton. In addition, cultured keratinocytes efficiently secreted the S100A8/A9 dimer. These results together with previously published data suggest that S100A8 and S100A9 are novel players in wound repair, where they might be involved in the reorganization of the keratin cytoskeleton in the wounded epidermis, in the chemoattraction of inflammatory cells, and/or in the defense against microorganisms.After cutaneous injury, a series of biological events takes place that aims at the reconstruction of the damaged skin. Among them are the migration, proliferation, and differentiation of inflammatory, epithelial, and mesenchymal cells. These cells exert specific functions in a temporally and spatially coordinated manner such as the removal of irreversibly destructed tissue, the deposition of new extracellular matrix, and the reestablishment of the cutaneous barrier (1, 2). These processes are well described at the histological level, but little is known about their molecular basis.To gain insight into the molecular mechanisms that underlie the repair process, we performed a large scale subtractive hybridization screen to systematically identify genes that are differentially expressed in injured compared with normal skin. To minimize the risk of detecting differences in gene expression levels due to changes in cellular composition rather than to transcriptional regulation, we compared normal skin with early (24 h) wounds, because only minor changes in cell type composition occur during the initial wound healing period.One of the cDNA clones that we obtained encodes the murine S100A8 protein (also known as calgranulin A, MRP8, leukocyte protein L1, or cytokine CP-10). S100 proteins are intracellular Ca 2ϩ -binding and Ca 2ϩ -modulated proteins that form antiparallel noncovalently linked dimers in solution and play a role in various Ca 2ϩ -mediated cellular functions including cell growth and differentiation, energy metabolism, cytoskeletalmembrane interactions; some of th...

Dexamethasone (DEX)-mediated inhibition of druginduced, but not CD95 ligand-induced, apoptosis in malignant glioma cells correlates with wild-type p53 status. Here, we examined mechanisms underlying DEXmediated protection from apoptosis. DEX did not induce p53 expression in two p53 wild-type cell lines (U87MG, LN-229) and did not alter drug-induced p53 accumulation. human colon carcinoma cells. Paradoxically, while only p21 +/+ and p21 +/7 mouse embryonic ®broblasts showed enhance p21 WAF1/CIP1 levels after exposure to DEX, only p21 7/7 ®broblasts were protected from drug toxicity by DEX. The present study links DEX-mediated protection from cancer chemotherapy to a p53-independent pathway of regulating p21 WAF1/CIP1 expression in glioma cells but this eect appears to cell type-speci®c.

Exogenous glucocorticoids are known to inhibit wound repair, but the roles and mechanisms of action of endogenous glucocorticoids during the healing process are as yet unknown. Therefore, we wounded mice expressing a DNA-binding-defective mutant version of the glucocorticoid receptor (GR dim mice) and also analysed fibroblasts from these animals in vitro. We found a remarkably enlarged granulation tissue with a high fibroblast density in GR dim mice. This difference is likely to result from an increased migratory and proliferative capacity of GR dim fibroblasts and from elevated expression levels of soluble factors involved in granulation tissue formation in wounds of GR dim mice. In spite of the larger granulation tissue seen in early wounds, late wounds appeared normal, most likely due to an enhanced ability of GR dim fibroblasts to contract collagen. These results demonstrate an as yet unidentified role of endogenous glucocorticoids in the regulation of wound repair.

BACKGROUND:More than 900 hemoglobin (Hb) variants are currently known. Common techniques used in Hb analysis are electrophoretic and chromatographic assays. In our laboratory, we routinely apply chromatographic methods. To ascertain whether Hb variants are missed with our procedures, we additionally analyzed all samples with mass spectrometry (MS).

In this study, we present a versatile new procedure for the analysis of transferrin and its isoforms isolated from human body fluids such as serum, plasma, and cerebrospinal fluid. This method is based on a three-step procedure: (i) isolation of transferrins using anion-exchange chromatography with UV detection; (ii) concentration of the transferrin fraction; (iii) detection of the transferrins with liquid chromatography-electrospray mass spectrometry. Pre-analytical sample procedures can be omitted and no immunoaffinity columns or transferrin-specific immunoassays were used. Anticoagulants such as heparin, EDTA, citrate, and oxalate do not interfere with our analysis. According to their respective molecular masses, up to ten different isoforms of transferrin could be identified in a serum sample from a patient with a congenital disorder of glycosylation type Ia (CDG-Ia). The method was successfully applied to different pathological samples from patients with CDG-Ia, CDG-Ib, CDG-Ic, CDG-Ie, CDG-If, and CDG-IIa. Additionally, samples from alcohol consumers that were found with turbidimetric immunoassay to contain increased levels of carbohydrate-deficient transferrin were analyzed.