FUNCTIONAL ESTROGEN-RECEPTORS IN A HUMAN PREOSTEOCLASTIC CELL-LINE

The primary biological effect of the estrogen estradiol-17 beta (17 be ta E(2)) on bone is to decrease bone resorption. However, whether 17 b eta E(2) affects osteoclast differentiation or function directly or th rough its action on osteoblasts is unclear. To investigate this questi on we examined the human preosteoclastic cell Line FLG 29.1 for eviden ce of functional estrogen receptors (ERs). Southern blotting of revers e transcription-PCR amplification products with a P-32-labeled cDNA pr obe for the human ER mRNA demonstrated that FLG 29.1 cells express ER mRNA. Binding of [H-3]17 beta E(2) to nuclear ERs was steroid specific with approximate to 400 saturable, high affinity (K-d approximate to 1 nM) binding sites per cell nucleus. Nuclear ERs covalently labeled w ith [H-3]tamoxifen aziridine showed an apparent molecular weight of 65 ,000 by SDS/PAGE and Western blotting with the D75 monoclonal antibody to human ER. Pretreatment of cells with 0.1, 1.0, or 10 nM 17 beta E( 2) induced a dose- and time-dependent specific binding of progesterone to FGL 29.1 cells, and stimulation of the cells with 10 nM and 100 nM 17 beta E(2) significantly (P < 0.05) reduced cell proliferation. Tra nscriptional activity of the ER gene was detected by transient transfe ction of cells with the pERE-BLCAT plasmid containing the estrogen res ponse element for the vitellogenin A2 gene and the bacterial chloramph enicol acetyltransferase reporter gene. Treatment of FLG 29.1 cells wi th 10 nM 17 beta E(2) increased chloroamphenicol acetyltransferase exp ression from 5- to 29-fold compared to controls. These observations su ggest a potential role for estrogen in osteoclastogenesis.