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Cyclosporine A(CsA) is a widely used immunosuppressant, with clinical applications ranging from argan transplants to chronic inflammatory disease. One of the side effects associated with CsA treatment is the development of gingival overgrowth. Although CsA has been shown to stimulate IL-6 production in human gingiva fibroblast cells in culture, the signaling mechanism that regulate IL-6 production is not fully understood. The aim of this study was to examine the role of the p38 and extracellular signal-regulated kinase(ERK) mitogen-activated protein kinase (MAPK) pathway in regulating CsA mediated IL-6 production from human gingiva fibroblast. Human gingiva fibroblast cultures were established from systemically healthy gingival tissue donors. Calls were treated fot various time with CsA in the presence and absence of the specific p38 and ERK1,2 MAPK inhibitors SB203580 and PD98059 respectively, either untreated or treated. IL-6 levels in the cell culture media were measured by enzyme-linked immunosorbent assay. MAPK activation was demonstrated by immunoblot for the active kinase (tyrosine/threonine phosphorylated) in whole cell extracts using phospho-specific antibodies. p38 MARK activity in human gingival fibroblast cells was measuared using an in vitro immunokinase assay using ATP as the substrate. CsA (500ng/ml) stimulated a time-dependent increase in IL-6 production in human gingival fibroblast. The activation of the p38 and the ERK1,2MAPKs occured following CsA stimulation. Pretreatment with various inhibitors had effect on CsA stimulated p38 MAPK activity. PD98059 decreased CsA stimulated phosphorylation of ERK1,2. Additionally, Wortmannin and LY290042, a potent inhibitor of the PI3K-kinase were able to inhibit IL-6 activation, byt they had no effect on IL-6 activation when stimulated by CsA. This study provides evidence that both the p38 and ERK MAPK pathways are important for the regulation of the production of IL-6 from the human gingival fibroblast in response to CsA.
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