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(Investigative Ophthalmology and Visual Science. 2001;42:1080-1086.)
© 2001 by The Association for Research in Vision and Ophthalmology, Inc.

Induction of Adrenomedullin by Hypoxia in Cultured Retinal Pigment Epithelial Cells

Tetsuo Udono1,2, Kazuhiro Takahashi1, Masaharu Nakayama1, Ayako Yoshinoya1, Kazuhito Totsune3, Osamu Murakami3, Yusuf K. Durlu2, Makoto Tamai2 and Shigeki Shibahara1

1 From the Department of Molecular Biology and Applied Physiology, the 2 Department of Ophthalmology, and the 3 Second Department of Internal Medicine, Tohoku University School of Medicine, Miyagi, Japan.

PURPOSE. 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.




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