Osteoblast-specific inactivation of p53 results in locally increased bone formation.

Inactivation of the tumor suppressor p53 (encoded by the Trp53 gene) is relevant for development and growth of different cancers, including osteosarcoma, a primary bone tumor mostly affecting children and young adolescents. We have previously shown that deficiency of the ribosomal S6 kinase 2 (Rsk2) limits osteosarcoma growth in a transgenic mouse model overexpressing the proto-oncogene c-Fos. Our initial aim for the present study was to address the question, if Rsk2 deficiency would also influence osteosarcoma growth in another mouse model. For that purpose, we took advantage of Trp53fl/fl mice, which were crossed with Runx2Cre transgenic mice in order to inactivate p53 specifically in osteoblast lineage cells. However, since we unexpectedly identified Runx2Cre-mediated recombination also in the thymus, the majority of 6-month-old Trp53fl/fl;Runx2-Cre (thereafter termed Trp53Cre) animals displayed thymic lymphomas, similar to what has been described for Trp53-deficient mice. Since we did not detect osteosarcoma formation at that age, we could not follow our initial aim, but we studied the skeletal phenotype of Trp53Cre mice, with or without additional Rsk2 deficiency. Here we unexpectedly observed that Trp53Cre mice display a unique accumulation of trabecular bone in the midshaft region of the femur and the humerus, consistent with its previously established role as a negative regulator of osteoblastogenesis. Since this local bone mass increase in Trp53Cre mice was significantly reduced by Rsk2 deficiency, we isolated bone marrow cells from the different groups of mice and analyzed their behavior ex vivo. Here we observed a remarkable increase of colony formation, osteogenic differentiation and proliferation in Trp53Cre cultures, which was unaffected by Rsk2 deficiency. Our data thereby confirm a critical and tumorigenesis-independent function of p53 as a key regulator of mesenchymal cell differentiation.

Wnt1 is an Lrp5-independent bone-anabolic Wnt ligand.

WNT1 mutations in humans are associated with a new form of osteogenesis imperfecta and with early-onset osteoporosis, suggesting a key role of WNT1 in bone mass regulation. However, the general mode of action and the therapeutic potential of Wnt1 in clinically relevant situations such as aging remain to be established. Here, we report the high prevalence of heterozygous WNT1 mutations in patients with early-onset osteoporosis. We show that inactivation of Wnt1 in osteoblasts causes severe osteoporosis and spontaneous bone fractures in mice. In contrast, conditional Wnt1 expression in osteoblasts promoted rapid bone mass increase in developing young, adult, and aged mice by rapidly increasing osteoblast numbers and function. Contrary to current mechanistic models, loss of Lrp5, the co-receptor thought to transmit extracellular WNT signals during bone mass regulation, did not reduce the bone-anabolic effect of Wnt1, providing direct evidence that Wnt1 function does not require the LRP5 co-receptor. The identification of Wnt1 as a regulator of bone formation and remodeling provides the basis for development of Wnt1-targeting drugs for the treatment of osteoporosis.

[Effect of alendronate on expressions of osteoprotegerin and receptor activator of nuclear factor κB ligand in mouse osteoblasts].

OBJECTIVE: To investigate the effect of alendronate on the expressions of osteoprotegerin (OPG) and receptor activator of nuclear factor-κB ligand (RANKL) in mouse osteoblasts. METHODS: Mouse calvarial osteoblasts cultured in vitro were identified by alkaline phosphatase (ALP) staining and immunofluorescence assay of OPG and RANKL expressions. The second passage of the osteoblasts were treated with different concentrations of alendronate (10(-4) to 10(-7) mol/L) for 48 h, and the changes in OPG and RANKL mRNA and protein expressions were examined using real-time PCR and Western blotting, respectively. RESULTS: The isolated osteoblasts were positive for ALP and expressed OPG and RANKL. Real-time PCR and Western blotting showed that at the concentration of 1×10(-4) mol/L, alendronate caused an obvious down-regulation of OPG and RANKL expressions in the cells, whereas at lower concentrations, alendronate increased the expressions of both genes with the highest expressions occurring after treatment with 1×10(-5) mol/L. CONCLUSION: High concentrations of alendronate (>1×10(-4) mol/L) decrease the expressions of OPG and RANKL, whereas low concentrations (1×10(-5) to 1×10(-7) mol/L) increase their expressions in mouse osteoblasts cultured in vitro.