Submitted on April 30, 2006
Accepted on July 16, 2006
Morphogenic protein epimorphin protects intestinal epithelial cells from oxidative stress by the activation of EGF receptor and MEK/ERK, PI3 kinase/Akt signals
Masahiro Iizuka1*, Kenji Sasaki1, Yohei Hirai2, Kenichi Shindo1, Shiho Konno1, Hiroaki Itou1, Shigetoshi Ohshima1, Yasuo Horie1, and Sumio Watanabe1
1 Internal Medicine, Akita University School of Medicine, Akita, Japan
2 Morphoregulation, Institute for Frontier Medical Science, Kyoto University, Kyoto, Japan
* To whom correspondence should be addressed. E-mail: maiizuka{at}doc.med.akita-u.ac.jp.
Epimorphin is a mesenchymal protein that regulates morphogenesis of epithelial cells. Our preliminary study suggested a novel function of epimorphin in enhancing survival of intestinal epithelial cells (IEC). Oxidative stress leads to cell injury and death and is suggested to be a key contributor to pathogenesis of inflammatory bowel disease. This study was conducted to determine whether epimorphin protects IEC from oxidative stress. Rat intestinal epithelial cell line IEC-6 was cultured with epimorphin (10, 20 µg/ml), and the life span of IEC was assessed. The mean life span of IEC-6 cells was prolonged 1.9 fold (p<0.0006) by treatment with epimorphin. We then examined the signaling pathways of epimorphin. Epimorphin phosphorylated EGF receptor, activated the MEK/ERK1/2 MAPK and PI3 kinase/Akt pathways, phosphorylated Bad, and induced Bcl-XL and survivin. Hydrogen peroxide (1mM) induced cell death in 92% of IEC-6 cells, but epimorphin dramatically diminished (88.7%) cell death induced by hydrogen peroxide (p<0.0001). This protective effect of epimorphin was significantly attenuated by inhibitors of MEK and PI3 kinase (p<0.0001), or EGF receptor-neutralizing antibody (p=0.0007). In the wound assays, the number of migrated cells in the wound area decreased 72.5% by treatment with 30 µM of hydrogen peroxide, but epimorphin increased the number of migrated cells 3.18 fold (p<0.0001). These results support a novel function of epimorphin in protecting IEC from oxidative stress. This anti-oxidative function of epimorphin is dramatic and is likely mediated by the activation of EGF receptor and the MEK/ERK, PI3 kinase/AKT signaling pathways and through the induction of anti-apoptotic factors.
