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Functional Estrogen Receptors in Osteoblastic Cells Demonstrated by Transfection with a Reporter Gene Containing an Estrogen Response Element

Matthias Ernst^*, Malcolm G. Parker and Gideon A. Rodan

Department of Bone Biology, Merck, Sharp, and Dohme Research Laboratories West Point, Pennsylvania 19486

Address requests for reprints to: Dr. Gideon A. Rodan, Department of Bone Biology, Merck, Sharp, and Dohme Research Laboratories, West Point, Pennsylvania 19486.

Abstract

Although a small number of estrogen receptors (ER) were visualized in osteoblastic cells, and estradiol (E₂) has some effects on osteoblasts in vitro, the direct action of E₂ on osteoblasts has not been fully established. To determine the presence of functional ER in osteoblasts, we transfected cells with a plasmid containing the chloramphenicol acetyl transferase (CAT) reporter gene and the estrogen-responsive element (ERE) from the vitellogenin A2 gene. E₂-dependent induction of CAT activity was determined 48 h after transient transfection and subsequent treatment with 10–100 nM 17β-E₂. 17β-E₂, but not 17-E₂, dihydrotestosterone, or progesterone, induced CAT activity in a dose-dependent manner (up to 6-fold) in rat calvarial fraction-3, RCT-3, PyMS, and UMR-106 cells as well as in the human osteosarcoma cell line SaOS-2/B-10. In contrast, E₂ had no effect on the induction of CAT activity in the preosteoblastic cell lines RCT-1 and TRAB-11, in the rat osteosarcoma cell line ROS 17/2.8, and in the fibroblastic cell lines BALB-c/3T3 and NRK. Overexpression of ER using a simian virus-40-based expression vector not only conferred or enhanced E₂-dependent induction of CAT in all cell types, but augmented E₂-dependent expression of insulin-like growth factor-I and E₂-stimulated DNA synthesis in primary calvarial and PyMS osteoblastic cells, respectively. These data show the presence of low levels of functional endogenous ER in some, but not all, osteoblastic cells and suggest that the abundance of ER may be rate limiting in the action of E₂ on these cells.

FOOTNOTES

^* Present address: Ludwig Institute for Cancer Research, Royal Melbourne Hospital, Victoria 3050, Australia.

Present address: Molecular Endocrinology Laboratory, Imperial Cancer Research Fund, London, WC2A 3PX United Kingdom.

Received for publication April 23, 1989. Revision received August 19, 1991. Accepted for publication August 20, 1991.

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