Cbfa1基因调节骨形成与骨吸收的研究进展

骨的形成与吸收是一个涉及众多的基因和生长因子的复杂过程,凡可使骨形成下降和骨吸收增加的因素都会促进骨质疏松的发生.Cbfa1是成骨细胞分化和成熟的特异性转录因子,研究发现Cbfa1可通过促进骨形成和抑制骨吸收,对骨质疏松的发生、发展起到抑制作用,因此,我们期望随着Cbfa1基因研究的深入将为防治骨质疏松提供新思路.     

淫羊蕾苷抗骨质疏松研究进展

淫羊藿苷是中草药淫羊藿的主要有效成分之一,近年来对其在细胞水平的抗骨质疏松作用研究日益增多.笔者从骨髓间充质干细胞、成骨细胞和破骨细胞三方面入手,综述了淫羊藿苷抗骨质疏松的研究进展,并提出了今后的研究思路.     

Docetaxel inhibits bone resorption through suppression of osteoclast formation and function in different manners

Osteoclasts are formed from the monocyte-macrophage lineage in response to receptor activator of nuclear factor κB ligand (RANKL) expressed by osteoblasts. Bone is the most common site of breast cancer metastasis, and osteoclasts play roles in the metastasis. The taxane-derived compounds paclitaxel and docetaxel are used for the treatment of malignant diseases, including breast cancer. Here we explored the effects of docetaxel on osteoclastic bone resorption in mouse culture systems. Osteoclasts were formed within 6 days in cocultures of osteoblasts and bone marrow cells treated with 1,25-dihydroxyvitamin D₃ plus prostaglandin E₂. Docetaxel at 10⁻⁸ M inhibited osteoclast formation in the coculture when added for the entire culture period or for the first 3 days. Docetaxel, even at 10⁻⁶ M added for the final 3 days, failed to inhibit osteoclast formation. Osteoprotegerin, a decoy receptor of RANKL, completely inhibited osteoclast formation when added for the final 3 days. Docetaxel at 10⁻⁸ M inhibited the proliferation of osteoblasts and bone marrow cells. RANKL mRNA expression induced by 1,25-dihydroxyvitamin D₃ plus prostaglandin E₂ in osteoblasts was not affected by docetaxel even at 10⁻⁶ M. Docetaxel at 10⁻⁶ M, but not at 10⁻⁸ M, inhibited pit-forming activity of osteoclasts cultured on dentine. Actin ring formation and l-glutamate secretion by osteoclasts were also inhibited by docetaxel at 10⁻⁶ M. Thus, docetaxel inhibits bone resorption in two different manners: inhibition of osteoclast formation at 10⁻⁸ M and of osteoclast function at 10⁻⁶ M. These results suggest that taxanes have beneficial effects in the treatment of bone metastatic cancers. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

RNA interference for noggin enhances the biological activity of bone morphogenetic proteins in vivo and in vitro

Noggin is a major extracellular antagonist to bone morphogenetic proteins (BMPs) which binds to BMPs and blocks binding of them to BMP-specific receptors and negatively regulates BMP-induced osteoblastic differentiation. In this study, we investigated the effect of noggin silencing by transfection of small interfering RNA (siRNA) on BMP-induced osteoblastic differentiation in vitro and ectopic bone formation in vivo induced by recombinant human BMP-2 (rhBMP-2). Noggin mRNA expression was up-regulated in response to rhBMP-2 in C2C12 cells, a myoblastic cell line, in dose- and time-dependent fashion as determined by real-time RT-PCR assay. Silencing of noggin expression by transfection of noggin siRNA suppressed BMP-stimulated noggin expression, resulting in acceleration of BMP-induced osteoblastic differentiation. For in vivo noggin silencing, siRNA was injected locally into back muscles and transfected into local cells by electroporation, where rhBMP-2-retaining (5 μg) collagen disks had been surgically placed. The implants were harvested at 2 weeks after surgery from experimental and control group mice and analyzed by radiological and histological methods. As a result, bone mineral content of ossicles ectopically induced by rhBMP-2 was significantly increased by silencing of noggin. Our findings suggest that silencing of noggin enhances the osteoblastic differentiation of BMP-responding cells in vitro and new bone formation induced by rhBMP-2 in vivo by eliminating negative regulation of the effects of BMP. RNA interference might be useful for intensifying the effects of BMP in promoting new bone (callus) formation in repair of damaged bone.     

New anthracene glycosides from <Emphasis Type="Italic">Rhodomyrtus tomentosa</Emphasis> stimulate osteoblastic differentiation of MC3T3-E1 cells

Two new anthracene glycosides (1, 2) were isolated from aerial parts of Rhodomyrtus tomentosa, along with three known compounds (3–5). The structures of two new compounds were established to be 4,8,9,10-tetrahydroxy-2,3,7-trimethoxyanthracene-6-O-β-D-glucopyranoside (1) and 2,4,7,8,9,10-hexahydroxy-3-methoxyanthracene-6-O-α-L-rhamnopyranoside (2) based on spectroscopic and chemical methods. Among them, compound 1, 2, and 5 significantly (P<0.05) increased the alkaline phosphatase activity, collagen synthesis, and mineralization of the nodules of MC3T3-E1 osteoblastic cells compared to those of the control, respectively.     

人健康和炎性牙槽骨成骨细胞COX-2、PGE2、OPG,RANKL的表达

目的：初步探讨COX-2、PGE2、OPG和RANKL在牙周炎发病机制中所起的作用及其相互关系。方法：采用组织块法对人健康和牙周炎性牙槽骨进行体外培养和传代,加入含100μg/LrhOPG的不含胎牛血清的DMEM2mL。严格按照ELISA试剂盒说明进行操作,获得各指标的检测数值。结果：牙周炎组RANKL、PGE2、COX-2的表达较健康组明显增高,OPG的表达较健康组明显降低。加入rhOPG后,牙周炎组RANKL、PGE2、COX-2的表达明显降低;OPG表达明显增加。健康组RANKL、PGE2、COX-2表达明显降低;OPG表达增加。结论：RANKL、PGE2、COX-2可促进牙周炎的发生、发展,而OPG对牙周炎的发生、发展可起抑制作用,同时表明人工重组OPG可能协同其内源性OPG来共同抑制RANKL、PGE2、COX-2的活性。     

Aminobisphosphonates Cause Osteoblast Apoptosis and Inhibit Bone Nodule Formation In Vitro

Bisphosphonates are widely used for the treatment of bone diseases associated with increased osteoclastic bone resorption. Bisphosphonates are known to inhibit biochemical markers of bone formation in vivo, but it is unclear to what extent this is a consequence of osteoclast inhibition or a direct inhibitory effect on cells of the osteoblast lineage. In order to investigate this issue, we studied the effects of various bisphosphonates on osteoblast growth and differentiation in vitro. The aminobisphosphonates pamidronate and alendronate inhibited osteoblast growth, caused osteoblast apoptosis, and inhibited protein prenylation in osteoblasts in a dose-dependent manner over the concentration range 20-100 microM. Further studies showed that alendronate in a dose of 0.1 mg/kg inhibited protein prenylation in calvarial osteoblasts in vivo, indicating that alendronate can be taken up by osteoblasts in sufficient amounts to inhibit protein prenylation at clinically relevant doses. Pamidronate and alendronate inhibited bone nodule formation at concentrations 10-fold lower than those required to inhibit osteoblast growth. These effects were not observed with non-nitrogen-containing bisphosphonates or with other inhibitors of protein prenylation and were only partially reversed by cotreatment with a fourfold molar excess of ss-glycerol phosphate. We conclude that aminobisphosphonates cause osteoblast apoptosis in vitro at micromolar concentrations and inhibit osteoblast differentiation at nanomolar concentrations by mechanisms that are independent of effects on protein prenylation and may be due in part to inhibition of mineralization. While these results need to be interpreted with caution because of uncertainty about the concentrations of bisphosphonates that osteoblasts are exposed to in vivo, our studies clearly demonstrate that bisphosphonates exert strong inhibitory effects on cells of the osteoblast lineage at similar concentrations to those that cause osteoclast inhibition. This raises the possibility that inhibition of bone formation by bisphosphonates may be due in part to a direct inhibitory effect on cells of the osteoblast lineage.     

Bisphosphonate Treatment Increases the Size of the Mandibular Condyle and Normalizes Growth of the Mandibular Ramus in Osteoprotegerin-Deficient Mice

Osteoprotegerin (OPG) is a novel secreted member of the tumor necrosis factor receptor family which plays a crucial role in negative regulation of osteoclastic bone resorption. OPG-deficient (OPG–/–) mice develop severe osteoporosis caused by significant enhancement of bone resorption by osteoclasts. We investigated the effect of administering bisphosphonate on mandibular growth and development in OPG–/– mice. Eight-week-old male OPG–/– mice and wild-type (WT) mice were administered bisphosphonate (1.25 mg/kg body weight) intraperitoneally once every 3 days for 30 days. All bone formation-related parameters and bone resorption-related parameters were significantly lower in OPG–/– mice with bisphosphonate than in those without bisphosphonate. The volume of the whole condyle and the mandibular length in OPG–/– mice without bisphosphonate were significantly smaller than in WT mice without bisphosphonate. Bisphosphonate treatment of the OPG–/– mice resulted in an increase in the volume of the mandibular condyle and mandibular ramus length. In fact, the mandibular ramus length in OPG–/– mice with bisphosphonate was similar to the length in WT mice without bisphosphonate. Histologically, the surface irregularity of the mandibular condyle that was observed in the OPG–/– mice without bisphosphonate tended to be less marked in the OPG–/– mice with bisphosphonate, and the proportion of the area of the cartilage layer relative to the whole condyle was significantly larger in OPG–/– mice with bisphosphonate than in those without bisphosphonate. In conclusion, bisphosphonate treatment results in an increase in mandibular condylar dimensions and normalization of mandibular ramus growth.     

Mechanical force-induced midpalatal suture remodeling in mice

Mechanical stress is an important epigenetic factor for regulating skeletal remodeling, and application of force can lead to remodeling of both bone and cartilage. Chondrocytes, osteoblasts and osteoclasts all participate and interact with each other in this remodeling process. To study cellular responses to mechanical stimuli in a system that can be genetically manipulated, we used mouse midpalatal suture expansion in vivo. Six-week-old male C57BL/6 mice were subjected to palatal suture expansion by opening loops with an initial force of 0.56 N for the periods of 1, 3, 5, 7, 14 or 28 days. Periosteal cells in expanding sutures showed increased proliferation, with Ki67-positive cells representing 1.8 ± 0.1% to 4.5 ± 0.4% of total suture cells in control groups and 12.0 ± 2.6% to 19.9 ± 1.2% in experimental/expansion groups (p < 0.05). Starting at day 1, cells expressing alkaline phosphatase and type I collagen were seen. New cartilage and bone formation was observed at the oral edges of the palatal bones at day 7; at the nasal edges only bone formation without cartilage appeared to occur. An increase in osteoclast numbers suggested increased bone remodeling, ranging from 60 to 160% throughout the experimental period. Decreased Saffranin O staining after day 3 suggested decreased proteoglycan content in the secondary cartilage. Micro-CT showed a significant increase in maxillary width at days 14 and 28 (from 2334 ± 4 μm to 2485 ± 3 μm at day 14 and from 2383 ± 5 μm to 2574 ± 7 μm at day 28, p < 0.001). The suture width was increased at days 14 and 28, except in the oral third region at day 28 (from 48 ± 5 μm to 36 ± 4 μm, p < 0.05). Bone volume/total volume was significantly reduced at days 14 and 28 (50.2 ± 0.7% vs. 68.0 ± 3.7% and 56.5 ± 1.0% vs. 60.9 ± 1.3%, respectively, p < 0.05), indicative of increased bone marrow space. These findings demonstrate that expansion forces across the midpalatal suture promote bone resorption through activation of osteoclasts and bone and cartilage formation via increased proliferation and differentiation of periosteal cells. Mouse midpalatal suture expansion would be useful in further studies of the ability of mineralized tissues to respond to mechanical stimulation.