Research on inhibitors for autocrine and cross-induction mechanism in human keratinocytes

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It has been reported that several EGF family are autocrine gowth factors in normal human keratinocytes (NHEK) and can induce the other EGF family known as cross-induction mechanism. To elucidate autocrine and cross-induction mechanism, we investigated the signal transduction pathways of auto- and cross-induction by HB-EGF. HB-EGF phosphorylated EGFR after 5 min, and induction of HB-EGF was inhibited by the addition of AG1483, an EGFR inhibitor. ERK, p38, and JNK were phosphorylated by the addition of HB-EGF after 10 min, suggesting the activation of all three pathways by HB-EGF. NHEK were treated with various signal transduction inhibitors ; then, the induction of EGF family mRNA was examined by RPA. The auto- and cross-induction of HB-EGF was completely blocked by treatment with curcumin, a natural JNK inhibitor, but not by p38 or MEK inhibitors. These results demonstrate that the auto- and cross-induction mechanisms of HB-EGF are mediated by the EGFR and JNK pathways. For cutaneous w … More ound healing, the most important growth factor is EGF family. HB-EGF, an autocrine growth factor in NHEK, is thought to play an important role in skin wound healing, although there have been no study about an involvement of HB-EGF in in vivo skin wound healing using mouse model. Since germline targeting of the HB-EGF gene resulted in embryonic lethality, we generated keratinocyte specific HB-EGF deficient mice (KS-HB-EGF-/-mice) using Cre/loxP technology in combination with keratin 5 promoter. A cutaneous wound was made on the mouse back skin using 6-mm punch biopsy and wound closure was measured by migration of epidermal edge stained by anti-keratin antibody. Wound closure was markedly retarded in KS-HB-EGF-/-mice compared to wild type mice on day 9. To further clarify the role of HB-EGF in cutaneous wound healing, we performed scraping wound assay using NHEK. HB-EGF mRNA was rapidly induced at 2 h after scraping, although the slight increase of TGF-alpha, amphiregulin and epiregulin m RNA was observed. Taking together, HB-EGF plays an important role in skin wound healing by accelerating keratinocyte migration. Less