Osteoblast Deletion of Exon 3 of the Androgen Receptor Gene Results in Trabecular Bone Loss in Adult Male Mice

Abstract

The mechanism of androgen action on bone was studied in male mice with the AR deleted in mature osteoblasts. These mice had decreased trabecular bone volume associated with a decrease in trabecular number, suggesting that androgens may act directly on osteoblasts to maintain trabecular bone.Introduction: Androgens modulate bone cell activity and are important for the maintenance of bone mass. However, the mechanisms by which they exert these actions on bone remain poorly defined. The aim of this study was to investigate the role of androgens acting through the classical androgen receptor (AR) signaling pathways (i.e., DNA-binding dependent pathways) in osteoblasts using male mice in which exon 3 of the AR gene was deleted specifically in mature osteoblasts. Materials and Methods: Mice with a floxed exon 3 of the AR gene were bred with Col 2.3-cre transgenic mice, in which Cre recombinase is expressed in mineralizing osteoblasts. The skeletal phenotype of mutant mice was assessed by histomorphometry and quantitative CT at 6, 12, and 32 weeks of age (n ‫ס‬ 8 per group). Wildtype, hemizygous exon 3 floxed and hemizygous Col 2.3-cre male littermates were used as controls. Data were analyzed by one-way ANOVA and Tukey's posthoc test. Results: CT analysis of the fifth lumbar vertebral body showed that these mice had reduced trabecular bone volume (p < 0.05) at 32 weeks of age compared with controls. This was associated with a decrease in trabecular number (p < 0.01) at 12 and 32 weeks of age, suggesting increased bone resorption. These effects were accompanied by a reduction in connectivity density (p < 0.01) and an increase in trabecular separation (p < 0.01). A similar pattern of trabecular bone loss was observed in the distal femoral metaphysis at 32 weeks of age. Conclusions: These findings show that inactivation of the DNA binding-dependent functions of the AR, specifically in mature osteoblasts in male mice, results in increased bone resorption and decreased structural integrity of the bone, leading to a reduction in trabecular bone volume at 32 weeks of age. These data provide evidence of a role for androgens in the maintenance of trabecular bone volume directly through DNA binding-dependent actions of the AR in mature osteoblasts.

Androgen receptor (AR) in osteocytes is important for the maintenance of male skeletal integrity: Evidence from targeted AR disruption in mouse osteocytes

“…(27) Our findings suggest that this age-related decline in trabecular integrity is enhanced by inactivation of the AR in osteocytes, in line with results from previous mice models. (12,13) Studies using the col2.3Cre AR-knockout mice and osteocalcin Cre ARknockout mice also demonstrated that an inactivation of the DNA binding-dependent functions of the AR, specifically in mature osteoblasts of male mice, increases bone resorption and reduces structural integrity of the bone, leading to a similar acceleration of the age-related reduction in trabecular bone volume. (12,13) Expanding the findings of these studies, our current data support the concept that, for the maintenance of the trabecular network after puberty, a functional AR is not only required at the osteoblast stage but also at the osteocyte stage.…”

“…In line with our findings, previous models inactivating the AR at earlier stages of the osteoblast did not observe a significant difference in cortical structure, but mechanical properties were not documented. (12,13) The absence of a major cortical phenotype across a number of similar mice models contrasts with the significant cortical phenotype characterized by reduction of periosteal expansion observed in the ubiquitous ARKO model. (5) Puberty appears to be a critical period for the development of skeletal sexual dimorphism as characterized by wider and longer bones in male.…”

“…In these two models, both the osteoblast and the osteocyte were targeted, which resulted in a trabecular bone phenotype, confirming that the osteoblast and/or osteocyte is a target cell for AR-mediated maintenance of trabecular bone volume. (12,13) Osteoblast development, however, involves three separate stagesproliferation, matrix development, and maturation-as well as mineralization of the bone matrix, (14) and it remains unknown at which stage(s) androgens act. Interestingly, the highest expression of AR has been observed in osteocytes.…”

Inactivation of the androgen receptor in bone-forming cells leads to trabecular bone loss in adult female mice

“…Some results have indicated a reduction in cortical bone thickness but no change in periosteal circumference when the osteocalcin promoter was used to target the exon 3 of the AR gene, 19 whereas no effect on the cortical bone was observed in another study utilizing the type I collagen 2.3 promoter. 20 In a recent study by Sinnesael et al, 21 osteocytic AR was shown to be crucial for the maintenance of trabecular bone volume in male mice. The role of AR in the regulation of the female skeleton is currently incompletely characterized.…”

“…Inactivation of the AR DNA-binding activity in osteocalcinexpressing cells 19 and in collagen-1-synthesizing osteoblasts 20 has been shown to reduce trabecular bone volume in male mice. In cortical bone various effects have been reported.…”