摘要
AIM To define signaling events initiating healing after intestinal epithelial injury, activation of mitogen activated protein kinase (MAPK) pathways was assessed after wounding using an in vitro model. METHODS Proteins isolated from wounded monolayers of nontransformed intestinal epithelial cells (IEC 6) were analyzed for tyrosine phosphorylation and MAPK expression by Western blot. Extracellular signal regulated kinase (ERK) 1, ERK2, and Raf 1 activities were assessed by immune complex kinase assays. RESULTS Tyrosine phosphorylation of several proteins including ERK1 was substantially increased 5 minutes after injury. Another MAPK, c Jun N terminal protein kinase (JNK), was also activated after wounding. Conditioned medium from wounded but not intact IEC 6 monolayers resulted in increased activity of ERK1, ERK2, and Raf 1 kinase. Wound conditioned medium stimulated proliferation of subconfluent IEC 6 cells compared with conditioned medium from intact IEC 6 cultures and contained higher amounts of transforming growth factor (TGF) α than supernatants of confluent IEC 6 cultures. Activation of ERK1 and ERK2 was partially inhibited by neutralizing anti TGF α. CONCLUSION Wounding of intestinal epithelial cells results in activation of Raf 1, ERK1, ERK2, and JNK1 MAPKs and subsequent cell proliferation in vitro. Activation of ERK1 and ERK2 is mediated in part by TGF α.
