ABSTRACT
Obsjective To investigate the effect of signal transduction pathway of p38 mitogen-activated protein kinase(p38MAPK) in epithelial cells activated by eosinophils on the expression and release of inflammatory mediators during co-culturing of two types of cells.Methods Eosinophils were isolated from human peripheral blood with magnetic micro beads conjugated CD16 antibody,LS separation column and magnetic cell sorting system(MCSS);The proteins of phospho-p38 MAPK and phospho-ATF-2 in BEAS-2B cells were analyzed by Western blot and enzyme-linked immuno sorbent assay(ELISA);The activity of phospho-p38MAPK was assayed by micro-bead Immunoprecipitatioin assay and Western blot.Results The interaction of eosinophils and BEAS-2B cells in co-culturing was found to elevate the protein expression and the activity of phospho-p38 MAPK in BEAS-2B cells;Phosphorylation of ATF-2 protein,a transcription factor that could regulate genes to express in cells and a substrate of phospho-p38 MAPK in the cascades of p38 MAPK signal transduction pathway was increased and it could be effectively suppressed by SB 203580,a selective inhibitor of p38 MAPK;eosinophils fixed with paraformaldehyde could also activate BEAS-2B cells to increase the phosphorylation of p38 MAPK protein.Conclusion To summarize the above results,it is showed that BEAS-2B cells activated by co-culturing with eosinophils to express and release inflammatory mediators are,at least partly,regulated by p38 MAPK signal transduction pathway.
ABSTRACT
Objective To investigate p38 MAPK signal transduction pathway of cytokine release in co-culture of BEAS-2B cells and eosinophils.Methods Eosinophils in human peripheral blood were isolated using anti-CD16 magnetic micro beads.Co-culture of BEAS-2B cells and eosinophils was taken as experimental model to investigate the inhibitive effect of SB 203580,a selective inhibitor of p38 MAPK,on release of IL-6,IL-8,IP-10 and MIG in culture supernatant.IL-6 was determined by ELISA kit and IL-8,IP-10 and MIG were measured using cytometric bead array(CBA)kit.Results SB 203580 could inhibit IL-6 and IL-8 release from BEAS-2B cells(P