Testosterone Prevents Orchidectomy-Induced Bone Loss in Estrogen Receptor-  Knockout Mice Liesbeth Vandenput,* Antwan G. H. Ederveen,† Reinhold G. Erben,‡ Kerstin Stahr,‡ Johannes V. Swinnen,* Erik Van Herck,* Annemieke Verstuyf,* Steven Boonen,§ Roger Bouillon,* and Dirk Vanderschueren* ,1 *Laboratorium voor Experimentele Geneeskunde en Endocrinologie and §Leuven University Centre for Metabolic Bone Diseases, Katholieke Universiteit Leuven, Leuven, Belgium; †Department of Pharmacology, Research and Development, NV Organon, Oss, The Netherlands; and ‡Institute of Physiology, Physiological Chemistry and Animal Nutrition, Ludwig Maximilians University, Munich, Germany Received May 24, 2001 To examine the role of the estrogen receptor- (ER) during male skeletal development, bone density and structure of aged ERKO mice and wild-type (WT) lit- termates were analyzed and skeletal changes in re- sponse to sex steroid deficiency and replacement were also studied. In comparison to WT, ERKO mice had smaller and thinner bones, arguing for a direct role of ER to obtain full skeletal size in male mice. However, male ERKO mice had significantly more trabecular bone as assessed both by pQCT and histomorphom- etry, indicating that ER is not essential to maintain cancellous bone mass. Six weeks following orchidec- tomy (ORX), both WT and ERKO mice showed high- turnover osteoporosis as revealed by increases in se- rum osteocalcin and decreases in trabecular (38% and 58% in WT and ERKO, respectively) and corti- cal bone density (5% and 4% in WT and ERKO, respectively). Administration of testosterone propi- onate (T, 5 mg/kg/day) completely prevented bone loss both in ERKO and in WT mice. As expected, estradiol (E2, 60 g/kg/day) replacement did not prevent cancel- lous bone loss in ORX ERKO mice. However, E2 stim- ulated bone formation at the endocortical surface in ORX ERKO, suggesting that osteoblasts may respond to nonER-mediated estrogen action. In conclusion, although functional ER may play a significant role during male skeletal development, this receptor does not seem essential for androgen-mediated skeletal maintenance in older male mice. © 2001 Academic Press Key Words: androgens; estrogens; estrogen recep- tor-; mice; orchidectomy; bone. Androgens are crucial for both skeletal development and maintenance in men and male rodents. The effects of androgens may be mediated directly by the androgen receptor (AR) or indirectly via aromatization of andro- gens into estrogens and subsequent stimulation of es- trogen receptor- (ER) and/or estrogen receptor- (ER). That estrogens may indeed play an important role in male skeletal growth was suggested by studies using an aromatase inhibitor, showing delayed skele- tal modeling in growing male rats (1). Moreover, both aromatase knockout (ArKO) (2) and estrogen recep- tor- knockout (ERKO) mice (3) show impaired skel- etal development. Also in humans, men with muta- tions in either the ER (4) or the aromatase gene (5, 6) show delayed skeletal maturation. Aromatization of androgens may also be important for skeletal mainte- nance after growth. Indeed, administration of an aro- matase inhibitor induces bone loss in aged male rats (7). Additionally, estrogen (8, 9) and selective estrogen receptor modulators (10, 11) prevent orchidectomy- induced bone loss in rodents. In elderly men, bone density seems to correlate more closely with estrogen than with androgen concentrations (12–14). In these men, estrogen also appears to be the dominant sex steroid regulating bone resorption (15). Finally, im- paired ER expression in osteoblasts and osteocytes in men suffering from idiopathic osteoporosis has been reported (16). ER-mediated androgen action may, however, not ex- plain all aspects of androgen effects on the male skel- eton. This is illustrated by observations that adminis- tration of nonaromatizable androgens prevents castra- tion-induced bone loss in adult rats (8). Furthermore, androgen-resistant rats (17) have decreased longitudi- nal and radial bone growth. The ERKO model is a unique model to study the relevance of ER- versus nonER-mediated androgen 1 To whom correspondence should be addressed at Laboratorium voor Experimentele Geneeskunde en Endocrinologie, Onderwijs en Navorsing, Campus Gasthuisberg, Herestraat 49, B-3000 Leuven, Belgium. Fax: +32-16-34 42 68. E-mail: dirk.vanderschueren@ med.kuleuven.ac.be. Biochemical and Biophysical Research Communications 285, 70 –76 (2001) doi:10.1006/bbrc.2001.5101, available online at http://www.idealibrary.com on 70 0006-291X/01 $35.00 Copyright © 2001 by Academic Press All rights of reproduction in any form reserved.