In vivo analysis of osteopontin function

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1)Production and analysis of anti-mouse osteopontin (OPN) monoclonal antibodies (mAb) : mAb, which show different features of tissue reactivity, were produced. mAb A reacts with kidney and bone cells, whereas mAb B reacts with kidney but not with bone cells. Analysis using these mAb showed that the different reactivity is ascribable to masking of epitope. Functional domains were analyzed using mAb and recombinant OPN,and found that an RGD sequence plays a role in cell adhesion to P388D1 and B16 cells and consequtive hydrophobic amino acids just aminoterminal side of the RGD sequence conforms binding site for hydroxyapatite. Analysis for heparin and fibronactin binding regions is now underway. 2)Roles of OPN in various diseases : Serum and local OPN levels were upregulated after lipopolysaccharide (LPS) stimulation and in Arthus reaction. Effect of LPS was abolished in M14M transgenic mice which show CD14 in hepatic cells under the control of metallothionein promoter and competitively u … More ptake LPS.OPN expression was detected in granuloma in lungs in transgenic mice which express TNF-alpha in lung type II epithelia under the control of surfactant promoter. Expression of OPN was strongly found in macrophages in Propionibacterium acnes induced-hepatic granuloma, whereas that in M14M mice was significantly low. Expression of OPN was not detected in islands in transgenic mice which express TNF-alpha in beta-cells under the control of insulin promoter. OPN was strongly expressed in macrophages in bovine type II collagen-induced arthritis in mice. Maternal or fetal injection of mAb resulted in abortion, suggesting inhibition of organ development. OPN was strlongly expressed in tumor cells and surrowunding macrophages. The level corresponded to metastatic and infiltrating activitis. 3)Analysis of OPN transgenic mice : Transgenic mice expressing OPN under the control of alpha antitrypsin 1 promoter showed high levels of serum OPN.Analysis of antibody production, cellmediated immunity, tumorigenesis, proliferation and metastasis are underway. Less