Title : Delayed production of biologically active O-glycosylated forms of human
eotaxin by
tumor-necrosis-factor-alpha-stimulated dermal fibroblasts
Abstract :
- Since a number of inflammatory skin diseases are characterized by selective eosinophil infiltration preferentially in the dermis, we speculated that dermal fibroblasts might represent a potential cellular source of eosinophil-selective attractants
- Cultivated dermal fibroblasts treated with tumor necrosis factor alpha secreted, not before day 3 of stimulation, eosinophil-specific chemotactic activity
- Purification of this activity revealed a heparin-binding protein with an apparent molecular mass of 13 kDa in SDS /polyacrylamide gel electrophoresis
- Peptide mapping with subsequent amino acid sequence analyses revealed it to be human eotaxin
- Natural eotaxin preparations contain 50% N-terminally truncated forms missing two or three amino acids
- It is O-glycosylated at Thr71 , resulting in at least two sialylated O-glycosylated variants
- Electrospray ionization mass spectrometric analyses revealed the natural eotaxin preparation to be heterogeneous with principal masses of 9033 Da and 9317 Da
- Natural eotaxin stimulated eosinophil chemotaxis with identical potency and efficacy as recombinant human eotaxin
- Neither neutrophils, monocytes or lymphocytes responded towards natural eotaxin preparations indicating that N-terminal truncation and O-glycosylation did not affect the cell-specificity of chemotactic activity
- Treatment of eosinophils with natural eotaxin desensitizes chemotactic responses towards eotaxin , regulated an normal T-lymphocyte expressed and secreted ( RANTES ) and monocyte chemotactic protein 3 ( MCP-3 ), whereas RANTES and MCP-3 were unable to desensitize natural eotaxin-dependent responses