Modification of anoxic neuronal injury by human recombinant epidermal growth factor and its possible mechanism.
A primary culture of cortical neurons was established from a 17-day rat embryo. Twenty-four hours after plating, the culture medium was changed to a chemically defined one, and human recombinant EGF (rEGF) was added to the medium at concentrations of 1, 10, and 100 ng/ml. Anoxic stress was exerted on the cultured neurons for 4 hr. Without anoxia, rEGF at a concentration of 10 ng/ml supported neuronal survival. When rEGF was not present in the culture medium, anoxic stress reduced neuronal density to one-tenth that of the nonanoxic group. The rEGF improved neuronal recovery from anoxia significantly, with survival dependent on rEGF dosage. Monoclonal antibody for EGF receptors canceled the effect of rEGF on survival of neurons in a dose-dependent manner. Therefore, rEGF functioned to support neuronal survival from anoxia through EGF receptors. Ontogeny of the EGF receptors was detected immunocytochemically with anti-EGF receptor antibody. About 70% of the cultured neurons were stained by the anti-EGF receptor antibody 3 days after change of medium, and immunoreactivity for EGF receptors was found to be located on the soma of neurons. Binding activity of EGF receptors was detected 5 days after change of medium with receptor binding assay and Scatchard analysis. High-affinity binding sites were detected only in neurons cocultured with rEGF (10 ng/ml) for 5 days. The data indicate that rEGF in sufficient concentration induces high-affinity sites of the EGF receptor and that EGF may have an antianoxic effect through the EGF receptors.
Kinoshita
A
A
Department of Neurosurgery, Osaka University Medical School, Japan.
Yamada
K
K
Hayakawa
T
T
Kataoka
K
K
Mushiroi
T
T
Kohmura
E
E
Mogami
H
H
D000818:Animals; D015687:Cell Hypoxia / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D066246:ErbB Receptors / Q000378:metabolism; D005333:Fetus; D051381:Rats; D011994:Recombinant Proteins / Q000494:pharmacology*
D011994:Recombinant Proteins; D004815:Epidermal Growth Factor; D066246:ErbB Receptors