Although bone resorption is accelerated with menopause, the means by which diminished circulating 17 β-estradiol (E2) promotes osteoclastic activity are unknown. We hypothesized that since the integrin αvβ3 is essential to the resorptive process, reduced E2 levels may increase the integrin's expression by osteoclast precursors. Thus, avian osteoclast precursors (known to contain E2 receptors) were exposed ± E2, surface iodinated, and lysed. The lysate was immunoprecipitated with an antibody recognizing the intact αvβ3 heterodimer. We find E2 alone fails to impact on αvβ3 expression. Most importantly, however, picomolar (i.e. post-menopausal), but not nanomolar (i.e. premenopausal) concentrations of E2, when added in conjunction with 1,25-dihydroxyvitamin D2 [1,25-(OH)2D2], enhance αvβ3 expression on the plasma membrane of avian osteoclast precursors relative to 1,25(OH)2D2 alone. Induction of αvβ3 by picomolar levels of E2 is dose-dependent, maximizing at 10-11-10-12 M, wherein the sex steroid enhances 1,25-(OH)2-D2-stimulated integrin expression approximately 2.5-fold. Northern analysis reveals that β3 mRNA levels parallel those of αvβ3. E2 (10-12 M) increases expression of β3 mRNA induced by a range of 1,25-(OH)2D2 concentrations extending from 10-10 m-10-8 M. The E2 + 1,25-(OH)2D2 additive effect on β3 mRNA appears as early as 1 day of treatment and progresses for at least 3 days. Consistent with evidence that the β3 subunit regulates heterodimer expression, the sex steroid does not impact αv mRNA. Attesting to the specificity of E2 on β3 mRNA expression, the steroid does not impact on β5 mRNA, and its stereoisomer, 17αE2, is inactive in these experiments. Likewise, E2 has no effect on retinoic acid-induced stimulation of β3 mRNA levels. While 1,25(OH)2D3 induction of β3 mRNA reflects transcriptional activation, nuclear run-on studies indicate that, despite its inductive effect on β3 mRNA levels, E2 does not alter β3 gene transcription. Transcriptional arrest experiments demonstrate the t1/2 of β3 mRNA derived from 1,25-(OH)2D3-treated cells cultured in the presence or absence of 10-8 M E2 is approximately 4 h. On the other hand, 10-l2 M E2 in conjunction with 1,25-(OH)2D3, more than triples stability of β3 message. Thus, in conjunction with 1,25(OH)2D3, E2, at levels circulating in postmenopausal (but not premenopausal) females, in whom the rate of bone resorption is accelerated, up-regulates, post-transcriptionally, in osteoclast precursors, αvβ3, an integrin heterodimer pivotal to the resorptive process.
ASJC Scopus subject areas
- Molecular Biology