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Rapid mitogen-activated protein kinase activation by transforming growth factor alpha in wounded rat intestinal
M Göke1, M Kanai, K Lynch-Devaney
1Gastrointestinal Unit, Department of Medicine, and Center for the Study of Inflammatory Bowel Disease, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA.
Background & Aims:
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)-alpha than supernatants of confluent IEC-6 cultures. Activation of ERK1 and ERK2 was partially inhibited by neutralizing anti-TGF-alpha.
Conclusions:
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-alpha.
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