Bone sialoprotein (BSP) and osteopontin (OPN) are two members of the SIBLING (Small Integrin-Binding LIgand, N-linked Glycoprotein) family of genetically related proteins that are clustered on human chromosome 4. We present evidence that this entire family is the result of duplication and subsequent divergent evolution of a single ancient gene. The solution structures of these two post-translationally modified recombinant proteins were solved by one dimensional proton NMR and transverse relaxation times. The polypeptide backbones of both free BSP and OPN rapidly sample an ensemble of conformations consistent with them both being completely unstructured in solution. This flexibility appears to enable these relatively small glycoproteins to rapidly associate with a number of different binding partners including other proteins as well as the mineral phase of bones and teeth. These proteins often function by bridging two proteins of fixed structures into a biologically active complex.

No abstract

Bone sialoprotein (BSP) is a secreted glycoprotein primarily found in sites of biomineralization. Recently, we demonstrated that BSP is strongly upregulated in osteotropic cancers and particularly those that exhibit microcalcifications. BSP contains an Arg-Gly-Asp (RGD) motif found in other adhesive molecules that interact with cellular integrins. In bone, BSP has been shown to mediate the attachment of osteoblasts and osteoclasts via alpha(v)beta(3) integrin receptors. Ligands for alpha(v)beta(3) integrin are considered to play a central role during angiogenesis. Therefore, we used human umbilical vein endothelial cells (HUVECs) to study the potential role of BSP in angiogenesis. We found that purified eukaryotic recombinant human BSP (rhBSP) is able to promote both adhesion and chemotactic migration of HUVECs in a dose-dependent manner. These interactions involve HUVEC alpha(v)beta(3) integrin receptors and the RGD domain of BSP. Indeed, HUVECs attach to a recombinant BSP fragment containing the RGD domain, whereas this response is not observed with the same fragment in which RGD has been mutated to Lys-Ala-Glu (KAE). A cyclic RGD BSP peptide inhibits both adhesion and migration of HUVECs to rhBSP. Moreover, anti-alpha(v)beta(3) but not anti-alpha(v)beta(5) monoclonal antibodies also prevent BSP-mediated adhesion and migration of HUVECs. We observed that both rhBSP and the RGD BSP recombinant fragment stimulated ongoing angiogenesis on the chorioallantoic chick membrane assay. BSP angiogenic activity was inhibited by anti-alpha(v)beta(3) antibody, and the KAE BSP fragment was inactive. Our findings represent the first report implicating BSP in angiogenesis. BSP could play a critical role in angiogenesis associated with bone formation and with tumor growth and metastatic dissemination.

Previously we have shown that two members of the newly named SIBLING (small integrin-binding ligand, N-linked glycoproteins) family of proteins, bone sialoprotein, and osteopontin, bound first to a cell surface receptor and then to complement Factor H thereby blocking the lytic activity of the alternative pathway of complement. Another member of this family, dentin matrix protein 1, is shown in this paper to be very similar to osteopontin in that it can bind strongly to Factor H (K a ϳ 1 nM) and block the lytic activity through either the vitronectin receptor (␣ V ␤ 3 integrin) or CD44. Binding of Factor H to SIBLING localized to the cells surface was demonstrated by fluorescence-activated cell sorting. Extensive overlapping fragment analyses suggests that both dentin matrix protein 1 and osteopontin interact with cell surface CD44 through their amino termini. Similar fragments of bone sialoprotein, like the intact protein, did not functionally interact with CD44. All three proteins are shown to act in conjunction with Factor I, a serum protease that, when complexed to appropriate cofactors, stops the lytic pathway by digesting the bound C3b in a series of proteolytic steps. These results show that at least three members of this family confer membrane cofactor protein-like activity (MCP or CD46) upon cells expressing RGD-binding integrins or CD44. The required order of the assembly of the complex suggests that this cofactor activity is limited to short diffusional distances.

Many cancers have a strong propensity to spread to bone. The processes involved in cancer dissemination to bone are complex and variable, and the changes in bone metabolism, once bony metastases have occurred, are usually profound. This review surveys the usefulness of bone markers in the diagnosis and follow-up of patients with malignant bone disease. In patients with established bone metastases, most markers of bone remodeling are abnormal compared with healthy controls or cancer patients without bone lesions. Although bone markers may have a potential as diagnostic tools in cancer patients, the available data do not allow final conclusions regarding the accuracy and validity of any of the presently used markers in the diagnosis of bone metastases. As regards monitoring of anticancer therapy, most markers of bone remodeling respond to active treatments. These indices therefore may have the potential to be used in the monitoring of antitumor therapies. However, most if not all of the available evidence on the use of bone markers in monitoring anticancer therapy is observational, and it remains unclear whether they have any beneficial effects on overall outcome. The same is true for their prognostic value, although evidence suggests that suppressed levels of bone formation or high rates of bone resorption are independent predictors of poor survival.

This is the first study assessing severe incidents and complications from a national outcome-tracking database. Annual identification and review of cases, perhaps with standardized database queries in the respective departments, might provide more detailed information about the cascades that lead to unfortunate outcomes.

RG seems not to be a long-term treatment option for patients with persistent central ACTH-evcess. Though, in order to reduce perioperative complications, short term treatment of patients could be an alternative.

Although numerous clinical studies have demonstrated the beneficial effect of preventing postmenopausal bone loss in elder women by long-term estrogen administration, effects of estrogen at the cellular level still remain unclear. Efforts to determine the precise role of bone cells in estrogen-mediated pathways are often hampered by the lack of suitable cell culture models. Presuming that sex steroids have a direct, stimulating effect on bone cells in vitro, we investigated the influence of 17beta-estradiol, testosterone and 1,25(OH)2D, on cell proliferation and differentiation using four established human osteosarcoma (HOS) cell lines of different gender of the donors (male origin: MG 63, HOS 58; female origin: SaOS 2, TE 85). These cell lines are believed to represent different stages of osteogenic maturation. Thus, the aim of this study was to clarify if possible responses to sex steroids are related to gender or osteogenic commitment of the individual cell culture. HOS cells were cultured in six-well plates and underwent hormone treatment (1 nM and 10 nM 17beta-estradiol. 0.1 nM and I nM testosterone and 1 microM 1,25(OH)2D3) for 48 h hours. Cell proliferation was determined by measuring total cell numbers. Cell function was studied by measuring alkaline phosphatase activity and secreted osteocalcin. In this study, estrogen significantly increased proliferation of both one male (MG 63) and one female (SaOS 2) cell line, but decreased proliferation of the female HOS TE 85 cell line significantly. Testosterone treatment had a positive effect on proliferation of only one female cell line (SaOS 2). A significant increase of alkaline phosphatase activity in SaOS 2 and HOS 58 cells and of osteocalcin levels in SaOS 2 cells was detected following estrogen treatment. Administration of 1.25(OH)2D3 was followed by an increased cell proliferation in HOS 58, MG 63 and SaOS 2. Significant gender-related differences could not be demonstrated. In conclusion, response to hormonal treatment with sex steroids is not related to the gender of the osteosarcoma cell line, but rather depends on its osteoblastic commitment.