The role of mechanical strain and estrogen status in regulating ERα levels in bone cells was studied in female rats. OVX is associated with decreased ERα protein expression/osteocyte, whereas habitual strain and artificial loading has only a small but positive effect, except on the ulna's medial surface, where artificial loading stimulates reversal of resorption to formation. Introduction: Osteoporosis is the most widespread failure of bones' ability to match their architectural strength to their habitual load bearing. In men and women, the severity of bone loss is associated with bioavailability of estrogen. This association could result from the estrogen receptor (ER) involvement in bone cells' adaptive response to loading. Materials and Methods: In vivo semiquantitative analysis of the amount of ERα protein per osteocyte was performed in immuno-cytochemically stained sections from control and loaded rat ulna, as well as tibias of ovariectomy (OVX) and sham-operated female rats. In vitro, the effect of exogenous estrogen (10−8 M) and mechanical strain (3400 μϵ, 1 Hz, 600 cycles) on the expression of ERα mRNA levels was assessed in ROS 17/2.8 cells in monolayers using real-time PCR and ER promoter activity. ERα translocation in response to exogenous estrogen and mechanical strain was assessed in both ROS 17/2.8 and MLO-Y4 cells. Results: More than 90 percent of tibial osteocytes express ERα, the level/osteocyte being higher in cortical than cancellous bone. OVX is associated with decreased ERα protein expression/osteocyte, whereas in the ulna habitual strain and that caused by artificial loading had only a small but positive effect, except on the medial surface, where loading stimulates reversal of resorption to formation. In unstimulated osteocytes and osteoblasts in situ, and osteocyte-like and osteoblast-like cells in vitro, ERα is predominantly cytoplasmic. In vitro, both strain and estrogen stimulate transient ERα translocation to the nucleus and transient changes in ERα mRNA. Strain but not estrogen also induces discrete membrane localization of ERα. Conclusions: Bone cells' responses to both strain and estrogen involve ERα, but only estrogen regulates its cellular concentration. This is consistent with the hypothesis that bone loss associated with estrogen deficiency is a consequence of reduction in ERα number/activity associated with lower estrogen concentration reducing the effectiveness of bone cells' anabolic response to strain.

Changes in subchondral bone (SCB) and cross-talk with articular cartilage (AC) have been linked to osteoarthritis (OA). Using micro-computed tomography (micro-CT) this study: (1) examines changes in SCB architecture in a non-invasive loading mouse model in which focal AC lesions are induced selectively in the lateral femur, and (2) determines any modifications in the contralateral knee, linked to changes in gait, which might complicate use of this limb as an internal control. Right knee joints of CBA mice were loaded: once with 2 weeks of habitual use (n = 7), for 2 weeks (n = 8) or for 5 weeks (n = 5). Both left (contralateral) and right (loaded) knees were micro-CT scanned and the SCB and trabecular bone analysed. Gait analysis was also performed. These analyses showed a significant increase in SCB thickness in the lateral compartments in joints loaded for 5 weeks, which was most marked in the lateral femur; the contralateral non-loaded knee also showed transient SCB thickening (loaded once and repetitively). Epiphyseal trabecular bone BV/TV and trabecular thickness were also increased in the lateral compartments after 5 weeks of loading, and in all joint compartments in the contralateral knee. Gait analysis showed that applied loading only affected gait in the contralateral himd-limb in all groups of mice from the second week after the first loading episode. These data indicate a spatial link between SCB thickening and AC lesions following mechanical trauma, and the clear limitations associated with the use of contralateral joints as controls in such OA models, and perhaps in OA diagnosis.

The contribution of the SNS to bone's response to mechanical loading is unclear. Using a noninvasive model of axial loading of the murine tibia, we found that sciatic neurectomy enhances load-induced new cortical bone formation and that pharmacological blockade of the SNS does not affect such responses, indicating that the SNS does not mediate the osteogenic effects of loading in cortical bone. Introduction: There is increasing evidence that the sympathetic nervous system (SNS) contributes to the regulation of bone mass and may influence remodeling by modulating bones' response to mechanical load-bearing. The aim of this study was to examine the effect of sciatic neurectomy (SN) on the changes in cortical bone formation induced in response to mechanical loading and to investigate whether the SNS is directly involved in such load-induced responses. Materials and Methods: Accordingly, load-induced responses were compared in tibias of growing and adult control C57Bl/J6 mice and in mice submitted to unilateral SN; noninvasive axial loading that induced 2000 μstrain on the tibia lateral midshaft cortex was applied cyclically, 5 or 100 days after surgery, for 7 minutes, 3 days/week for 2 weeks, and mice received calcein on the third and last days of loading. Tibias were processed for histomorphometry, and transverse confocal images from diaphyseal sites were analyzed to quantify new cortical bone formation. Chemical SNS inactivation was achieved by prolonged daily treatment with guanethidine sulfate (GS) or by the introduction of propranolol in drinking water. Results: Our results show that new cortical bone formation is enhanced by loading in all tibial sites examined and that load-induced periosteal and endosteal new bone formation was greater in the SN groups compared with sham-operated controls. This SN-related enhancement in load-induced cortical bone formation in tibias was more pronounced 100 days after neurectomy than after 5 days, suggesting that longer periods of immobilization promote a greater sensitivity to loading. In contrast, the increases in new bone formation induced in response to mechanical loading were similar in mice treated with either GS or propranolol compared with controls, indicating that inactivation of the SNS has no effect on load-induced cortical new bone formation. Conclusions: This study shows that SN, or the absence of loading function it entails, enhances loading-related new cortical bone formation in the tibia independently of the SNS.

Mechanoadaptation maintains bone mass and architecture; its failure underlies age-related decline in bone strength. It is unclear whether this is due to failure of osteocytes to sense strain, osteoblasts to form bone or insufficient mechanical stimulus. Mechanoadaptation can be restored to aged bone by surgical neurectomy, suggesting that changes in loading history can rescue mechanoadaptation. We use non-biased, whole-bone tibial analyses, along with characterisation of surface strains and ensuing mechanoadaptive responses in mice at a range of ages, to explore whether sufficient load magnitude can activate mechanoadaptation in aged bone. We find that younger mice adapt when imposed strains are lower than in mature and aged bone. Intriguingly, imposition of short-term, high magnitude loading effectively primes cortical but not trabecular bone of aged mice to respond. This response was regionally-matched to highest strains measured by digital image correlation and to osteocytic mechanoactivation. These data indicate that aged bone’s loading response can be partially recovered, non-invasively by transient, focal high strain regions. Our results indicate that old murine bone does respond to load when the loading is of sufficient magnitude, and bones’ age-related adaptation failure may be due to insufficient mechanical stimulus to trigger mechanoadaptation.