title: Heparin-binding epidermal growth factor-like growth factor is an autocrine growth factor for human keratinocytes.
abstract: Since heparin-binding epidermal growth factor (HB-EGF) is a member of the EGF family and binds to EGF receptor, we tested recombinant HB-EGF for its ability to stimulate human keratinocyte proliferation. The effect of HB-EGF on human keratinocytes was dependent on the cell density. HB-EGF optimally increased the cell number by 1.8-fold at 1.0 ng/ml for a 4-day incubation period under subconfluent culture. In contrast, under confluent culture, 10 ng/ml HB-EGF optimally increased the DNA synthesis 2.1-fold. To examine the production of HB-EGF by human keratinocytes, the analysis of human keratinocyte-conditioned medium was undertaken by a combination of heparin affinity column chromatography, EGF receptor-stimulating assay, immunoblotting, and neutralization. Heparin column chromatography fractionated three activities, peaks 1, 2, and 3, which contained immunoreactive 30- and 27-, 19-, and 14.5-kDa bands, respectively. The anti-HB-EGF-blocking antibody neutralized the activities of peaks 2 and 3 by 38 and 22%, respectively, but did not neutralize the activity of peak 1 at all. The antibody reduced the cell growth by 37% for a 4-day incubation period. Northern blot analysis detected a 2.5-kilobase transcript of HB-EGF. The addition of 1 ng/ml HB-EGF optimally increased the levels of HB-EGF mRNA 5.4-fold at 1 h and TGF-alpha mRNA 3.1-fold at 3 h. Interestingly, the addition of TGF-alpha at 1 ng/ml to keratinocyte cultures enhanced the level of HB-EGF mRNA 10.2-fold at 6 h. 1 ng/ml EGF also increased HB-EGF mRNA levels 10.9-fold at 1 h. These results suggest that HB-EGF is an autocrine growth factor for human keratinocytes, and HB-EGF and TGF-alpha act not only by an autoinductive mechanism but also by mutual amplification.
authors.affiliation: Department of Dermatology, Osaka University Medical School, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Hashimoto
authors.forename: S
authors.initials: S
authors.lastname: Higashiyama
authors.forename: H
authors.initials: H
authors.lastname: Asada
authors.forename: E
authors.initials: E
authors.lastname: Hashimura
authors.forename: T
authors.initials: T
authors.lastname: Kobayashi
authors.forename: K
authors.initials: K
authors.lastname: Sudo
authors.forename: T
authors.initials: T
authors.lastname: Nakagawa
authors.forename: D
authors.initials: D
authors.lastname: Damm
authors.forename: K
authors.initials: K
authors.lastname: Yoshikawa
authors.forename: N
authors.initials: N
authors.lastname: Taniguchi
mesh_terms: D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004815:Epidermal Growth Factor / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D006493:Heparin / Q000378:metabolism*; D066257:Heparin-binding EGF-like Growth Factor; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D015603:Keratinocytes / Q000166:cytology* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000494:pharmacology
chemical_list: C587883:HBEGF protein, human; D066257:Heparin-binding EGF-like Growth Factor; D036341:Intercellular Signaling Peptides and Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D004815:Epidermal Growth Factor; D006493:Heparin