Introduction: Pubertal sex steroids are indispensable for peak bone mass acquisition as well as skeletal sexual dimorphism. In males, both androgen receptor (AR) and estrogen receptor alpha (ERα) are required for optimal periosteal cortical bone expansion, while estrogens limit bone size in females. In early puberty, estrogens directly stimulate longitudinal growth via ERα in cartilage, but also indirectly via growth hormone (GH) and insulin-like growth factor I (IGF-I). Estrogens in late puberty however, also exert direct inhibitory effects on growth plate cartilage, hereby limiting longitudinal bone growth. Recently, it has become more evident that there is possible neuroendocrine regulation of ERα on bone metabolism, since bone is highly innervated and ERα is enriched throughout the central nervous system. Here, we aimed to study to what extent neuronal ERα contributes to the estrogenic action on skeletal growth during puberty.
Materials and methods: ERα flox mice were crossed with Thy1-Cre mice to generate mice with specific, tamoxifen-inducible inactivation of ERα in extrahypothalamic neurons (N-ERαKO). Tissue-specific ERα deletion in the central nervous system was validated using qPCR. Serum E2/T were measured with gas chromatography/mass spectrometry (GC-MS). At 16 weeks of age, both cortical and trabecular bone morphology was analyzed by microCT. A three-point bending test was conducted to measure the mechanical strength of the femur. To assess potential contribution of GH signaling, serum IGF-I and IGFBP-3 were measured by an in-house RIA and ELISA, respectively.
Results: A tamoxifen-induced knockout at 6 weeks of age reduced ERα mRNA levels in both cerebral cortex and brain stem by 50%, while ERα expression was unaffected in hypothalamus and non-neuronal tissues. Serum T and E2 remained normal in both sexes, indicating that hypothalamus-pituitary-gonadal function was not affected by ERα disruption. Inactivation of neuronal ERα did not alter body weight in males, but female N-ERαKO were 6.3% heavier (P < 0.01) and 2% longer (P < 0.05) compared to control littermates. In female N-ERαKO mice, femoral and L5 vertebral lengths increased by 2.4% (P < 0.01) and 4.8% (P < 0.01) respectively. Radial bone expansion at femoral midshaft also increased in female N-ERαKO concomitantly with higher serum IGF-I as well as IGFBP-3, which is a marker of GH secretion. Furthermore, the three-point bending test revealed increased bone strength in female N-ERαKO compared to control littermates. In contrast, inactivation of neuronal ERα had no major effect on bone growth in males.
Conclusions: Neuronal ERα limits female bone size and strength. The potential indirect effect of estrogens on GH/IGF-1 signaling represents an additional central inhibitory mechanism of action of estrogens on growth.
21 Oct 2019
Belgian Endocrine Society