Angiotensin II enhances EGF receptor expression levels via ROS formation in HaCaT cells.
BACKGROUND
Recent work has shown a novel function of angiotensin II (Ang II) in skin wound healing in which reactive oxygen species might be involved. As Ang II is known to increase superoxide production by activating NADPH oxidase in some non-phagocytic cells, we hypothesized that the produced superoxide by NADPH activation could contribute to the regulation of epidermal growth factor receptor (EGFR) in keratinocytes.


OBJECTIVE
We examined whether Ang II could generate superoxide and enhance EGFR expression levels in HaCaT cells.


METHODS
Superoxide formation was assessed by using hydroethidine. EGFR expression levels were examined by Western blotting.


RESULTS
Ang II (1-100 microM) increased the superoxide formation. Ang II (1-100 microM) resulted in a dose-dependent increase in cell proliferation in HaCaT cells. Heparin-binding epidermal growth factor activated the EGFR at 5-10 min. Although Ang II did not activate the EGFR, the expression levels of EGFR protein were increased in HaCaT cells treated with Ang II (1 microM) at 6h. Apocynin, a NADPH oxidase inhibitor, decreased the expression levels of EGFR. Xanthine/xanthine oxidase system, an exogenous superoxide generating system, enhanced the EGFR protein expression. Although Ang II did not affect the nitric oxide (NO) production, a NO synthase inhibitor N(omega)-nitro-l-arginine methyl ester suppressed the Ang II-induced EGFR expression levels in HaCaT cells. Thus, constitutive NO is required for the Ang II-induced EGFR expression in HaCaT cells.


CONCLUSIONS
These results suggest that Ang II enhances the cell proliferation and EGFR expression via superoxide production under the regulation of NO in HaCaT cells, implying that Ang II may regulate the proliferation, differentiation and tumorigenesis of the epidermis by harmonizing the superoxide and NO production.
Nakai
Kozo
K
Department of Dermatology, 1750-1 Ikenobe, Miki-cho, Kita-gun, Kagawa, Japan. kozo@kms.ac.jp
Yoneda
Kozo
K
Igarashi
Junsuke
J
Moriue
Tetsuya
T
Kosaka
Hiroaki
H
Kubota
Yasuo
Y
D000804:Angiotensin II / Q000378:metabolism / Q000494:pharmacology*; D001483:Base Sequence; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D017931:DNA Primers / Q000235:genetics; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D019820:Xanthine / Q000494:pharmacology; D014969:Xanthine Oxidase / Q000378:metabolism
D017931:DNA Primers; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D000804:Angiotensin II; D019820:Xanthine; D009569:Nitric Oxide; D014969:Xanthine Oxidase; D066246:ErbB Receptors; D019331:NG-Nitroarginine Methyl Ester