Low-density-lipoprotein receptor-related protein 5 (Lrp5) is a co-receptor in Wnt signaling, which plays a critical role in development and maintenance of bone. Osteoporosis-pseudoglioma syndrome, for instance, arises from loss-of- function mutations in Lrp5, and global deletion of Lrp5 in mice results in significantly lower bone mineral density. Since osteocytes are proposed to act as a mechanosensor in bone, we addressed a question whether a conditional loss-of-function mutation of Lrp5 selective to osteocytes (Dmp1-Cre; Lrp5f/f) would alter responses to ulna loading. Loading was applied to the right ulna for 3 min (360 cycles at 2 Hz) at a peak force of 2.65 N for 3 consecutive days, and the contralateral ulna was used as a non-loaded control. Young’s modulus was determined using a midshaft section of the femur. The results showed that compared to age-matched littermate controls, mice lacking Lrp5 in osteocytes exhibited smaller skeletal size with reduced bone mineral density and content. Compared to controls, Lrp5 deletion in osteocytes also led to a 4.6-fold reduction in Young’s modulus. In response to ulna loading, mineralizing surface, mineral apposition rate, and bone formation rate were diminished in mice lacking Lrp5 in osteocytes by 52%, 85%, and 69%, respectively. Collectively, the results support the notion that the loss-of-function mutation of Lrp5 in osteocytes causes suppression of mechanoresponsiveness and reduces bone mass and Young’s modulus. In summary, Lrp5-mediated Wnt signaling significantly contributes to maintenance of mechanical properties and bone mass.

BackgroundOsteoporosis is a skeletal disease leading to an increased risk of bone fracture. Using a mouse osteoporosis model induced by administration of a receptor activator of nuclear factor kappa-B ligand (RANKL), salubrinal was recently reported as a potential therapeutic agent. To evaluate the role of salubrinal in cellular fates as well as migratory and adhesive functions of osteoclast/osteoblast precursors, we examined the development of primary bone marrow-derived cells in the presence and absence of salubrinal. We addressed a question: are salubrinal’s actions more potent to the cells isolated from the osteoporotic mice than those isolated from the control mice?MethodsUsing the RANKL-injected and control mice, bone marrow-derived cells were harvested. Osteoclastogenesis was induced by macrophage-colony stimulating factor and RANKL, while osteoblastogenesis was driven by dexamethasone, ascorbic acid, and β-glycerophosphate.ResultsThe results revealed that salubrinal suppressed the numbers of colony forming-unit (CFU)-granulocyte/macrophages and CFU-macrophages, as well as formation of mature osteoclasts in a dosage-dependent manner. Salubrinal also suppressed migration and adhesion of pre-osteoclasts and increased the number of CFU-osteoblasts. Salubrinal was more effective in exerting its effects in the cells isolated from the RANKL-injected mice than the control. Consistent with cellular fates and functions, salubrinal reduced the expression of nuclear factor of activated T cells c1 (NFATc1) as well as tartrate-resistant acid phosphatase.ConclusionsThe results support the notion that salubrinal exhibits significant inhibition of osteoclastogenesis as well as stimulation of osteoblastogenesis in bone marrow-derived cells, and its efficacy is enhanced in the cells harvested from the osteoporotic bone samples.

BackgroundModerate loads with knee loading enhance bone formation, but its effects on the maintenance of the knee are not well understood. In this study, we examined the effects of knee loading on the activity of matrix metalloproteinase13 (MMP13) and evaluated the role of p38 MAPK and Rac1 GTPase in the regulation of MMP13.MethodsKnee loading (0.5–3 N for 5 min) was applied to the right knee of surgically-induced osteoarthritis (OA) mice as well as normal (non-OA) mice, and MMP13 activity in the femoral cartilage was examined. The sham-loaded knee was used as a non-loading control. We also employed primary non-OA and OA human chondrocytes as well as C28/I2 chondrocyte cells, and examined MMP13 activity and molecular signaling in response to shear at 2–20 dyn/cm2.ResultsDaily knee loading at 1 N for 2 weeks suppressed cartilage destruction in the knee of OA mice. Induction of OA elevated MMP13 activity and knee loading at 1 N suppressed this elevation. MMP13 activity was also increased in primary OA chondrocytes, and this increase was attenuated by applying shear at 10 dyn/cm2. Load-driven reduction in MMP13 was associated with a decrease in the phosphorylation level of p38 MAPK (p-p38) and NFκB (p-NFκB). Molecular imaging using a fluorescence resonance energy transfer (FRET) technique showed that Rac1 activity was reduced by shear at 10 dyn/cm2 and elevated by it at 20 dyn/cm2. Silencing Rac1 GTPase significantly reduced MMP13 expression and p-p38 but not p-NFκB. Transfection of a constitutively active Rac1 GTPase mutant increased MMP13 activity, while a dominant negative mutant decreased it.ConclusionsKnee loading reduces MMP13 activity at least in part through Rac1-mediated p38 MAPK signaling. This study suggests the possibility of knee loading as a therapy not only for strengthening bone but also preventing tissue degradation of the femoral cartilage.