Induction of adrenomedullin by hypoxia in cultured retinal pigment epithelial cells.
OBJECTIVE
To explore the effects of hypoxia on the production and secretion of adrenomedullin (ADM) and endothelin (ET)-1 in human retinal pigment epithelial (RPE) cells.


METHODS
RPE cells were cultured under normoxic or hypoxic (1% O2) conditions. Expression of ADM and ET-1 was examined by Northern blot analysis and radioimmunoassay. Effects of ADM and ET-1 on the number of RPE cells were examined by modified 3-(4,5-dimetylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay.


RESULTS
ADM mRNA expression levels and immunoreactive ADM levels in the medium were increased by hypoxia in all three human RPE cell lines (ARPE-19, D407, and F-0202). Immunoreactive ET was detected in the cultured media of D407 cells and ARPE-19 cells and identified as ET-1 by reversed-phase high performance liquid chromatography. Hypoxia treatment for 48 hours increased immunoreactive ET levels approximately 1.3-fold in the cultured media of D407, but not ARPE-19 cells. Hypoxia decreased the number of ARPE-19 cells and F-0202 cells, and the treatment with ADM ameliorated the hypoxia-induced decrease in the cell number. In contrast, exogenously added ET-1 had no significant effects on the number of ARPE-19 cells under normoxia and hypoxia.


CONCLUSIONS
Hypoxia increased the expression of ADM in all three human RPE cell lines, whereas the induction of ET-1 by hypoxia was found only in D407 cells. ADM induced by hypoxia may have protective roles against hypoxic cell damage in RPE cells.
Udono
T
T
Department of Molecular Biology and Applied Physiology, Tohoku University School of Medicine, Miyagi, Japan.
Takahashi
K
K
Nakayama
M
M
Yoshinoya
A
A
Totsune
K
K
Murakami
O
O
Durlu
Y K
YK
Tamai
M
M
Shibahara
S
S
D053607:Adrenomedullin; D015152:Blotting, Northern; D002452:Cell Count; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D019332:Endothelin-1 / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology; D006801:Humans; D000860:Hypoxia / Q000378:metabolism*; D010455:Peptides / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D011863:Radioimmunoassay; D014665:Vasodilator Agents / Q000378:metabolism* / Q000494:pharmacology
D019332:Endothelin-1; D010455:Peptides; D012333:RNA, Messenger; D014665:Vasodilator Agents; D053607:Adrenomedullin