Osteopontin (OPN) is a highly modified integrin-binding protein present in most tissues and body fluids where it has been implicated in numerous biological processes. A significant regulation of OPN function is mediated through phosphorylation and proteolytic processing. Proteolytic cleavage by thrombin and matrix metalloproteinases close to the integrin-binding Arg-Gly-Asp sequence modulates the function of OPN and its integrin binding properties. Osteopontin (OPN) 2 is a highly acidic phosphorylated glycoprotein containing an integrin-binding Arg-Gly-Asp (RGD) sequence. OPN is implicated in a diversity of physiological processes such as bone mineralization, inhibition of ectopic calcification, wound healing, inflammation, regulation of immune cell functions, and tumor growth (1-3). OPN is synthesized by a variety of cells and is present in most tissues and body fluids, including blood, urine, and milk (1).OPN is present in milk in very high concentrations (ϳ138 mg/liter) (4), but the function is not clear. However, several functions can be hypothesized, for instance OPN can inhibit the formation of renal stones by inhibiting growth and aggregation of calcium crystals (1). Similarly, OPN can be speculated to inhibit unintentional calcium crystallization and precipitation in milk. OPN has also been implicated in mammary gland development and differentiation (5). Furthermore, a significant proportion of the milk OPN is expected to pass through the gut and into the intestines largely intact upon milk consumption, as the protein is relatively resistant to proteolysis by neonatal gastric juice (6). This opens the possibility that OPN or OPN fragments could play a role in the infant immune response, as milk OPN can induce the expression of interleukin-12 from human intestinal lamina propria mononuclear cells (4).Many of the cellular functions propagated by OPN are mediated through interactions with integrin receptors. The ␣ v ␤ 6 ,, and ␣ v ␤ 3 integrins bind OPN via the RGD sequence (1,7,8), whereas the ␣ 4 ␤ 1 and ␣ 9 ␤ 1 integrins bind the cryptic non-RGD motif Ser-Val-Val-Tyr-Gly-Leu-Arg (SVVYGLR) (9, 10), and the monocyte ␣ X ␤ 2 -integrin receptor interacts with the highly acidic parts of OPN (11).OPN is extensively altered through post-translational modifications such as phosphorylation, sulfation, and glycosylation. These modifications have significant implications on the interaction with integrins (8). Another modification that can alter the functionality of OPN is proteolytic processing. Recently, intracellular cleavage of OPN by caspase-8 at Asp 119 -Phe 120 and Asp 141 -Gly 142 has been shown to be a regulatory switch in determining cell death of cancer cells (12). OPN is also a substrate for thrombin and matrix metalloproteinase (MMP)-2, -3, -7, and -9 (13-17). Thrombin hydrolyzes human OPN at Arg 152 -Ser 153 , whereas MMPs cleave nearby at Gly 150 -Leu 151 , which in all cases results in the generation of N-terminal OPN fragments containing an exposed RGD 145 sequence. Thrombin and MMP cleavage of OPN ...

Osteopontin (OPN) is a multifunctional bioactive protein that is implicated in numerous biological processes such as bone remodeling, inhibition of ectopic calcification, and cellular adhesion and migration, as well as several immune functions. Osteopontin has cytokine-like properties and is a key factor in the initiation of T helper 1 immune responses. Osteopontin is present in most tissues and body fluids, with the highest concentrations being found in milk. In the present study, ELISA for human and bovine milk OPN were developed and OPN concentration in human breast milk, bovine milk, and infant formulas was measured and compared. The OPN concentration in human milk was measured to approximately 138 mg/L, which corresponds to 2.1% (wt/wt) of the total protein in human breast milk. This is considerably higher than the corresponding OPN concentrations in bovine milk (approximately 18 mg/L) and infant formulas (approximately 9 mg/L). Moreover, bovine milk OPN is shown to induce the expression of the T helper 1 cytokine IL-12 in cultured human lamina propria mononuclear cells isolated from intestinal biopsies. Finally, the OPN concentration in plasma samples from umbilical cords, 3-mo-old infants, and pregnant and nonpregnant adults was measured. The OPN level in plasma from 3-mo-old infants and umbilical cords was found to be 7 to 10 times higher than in adults. Thus, the high levels of OPN in milk and infant plasma suggest that OPN is important to infants and that ingested milk OPN is likely to induce cytokine production in neonate intestinal immune cells.

Osteopontin (OPN) is a cytokine with multiple functions, including immune defense mechanisms against invading microorganisms. OPN-deficient mice are impaired in clearing intracellular pathogens, suggesting an important role of OPN during phagocytosis, but it remains to be defined how OPN may enhance this innate immune process. Here, we demonstrate that OPN binds to monocytes, but not resting T cells, NK cells, or B cells, and mediates chemoattraction of IL-1-activated human monocytes. Moreover, OPN binds in a specific manner to all known serotypes of the two bacterial species Streptococcus agalactiae and Staphylococcus aureus and opsonizes these bacteria for phagocytosis. We identify the integrin αXβ2 (CD11c/CD18), which is highly expressed on the cell surface of monocytes, as a novel OPN receptor. To eliminate the contribution from other molecular interactions between the bacteria and the phagocyte, we show that OPN-coated synthetic beads are phagocytosed in an αXβ2 integrin-dependent manner. The ligand recognition does not involve the RGD motif previously reported to support binding of OPN to integrins. Taken together, these data identify the αXβ2 integrin as a novel OPN receptor that is required for OPN-mediated phagocytosis, thereby elucidating an important mechanism of an innate immune function of OPN.

In Crohn's disease (CD) mucosal T‐cells produce increased interferon‐γ (IFN‐γ) and tumour necrosis factor‐α (TNF‐α) levels and TNF‐α antibody treatment [Infliximab (Ifx)] is effective. Osteopontin (OPN), a glycoprotein stimulating activated T‐lymphocytes, may be involved in the disturbed immune‐regulation but also in normal immune‐homeostasis and mucosal repair, since it is expressed in many tissues and present in human milk. This study investigates plasma‐OPN levels in CD patients during Ifx treatment and the in vitro effect of OPN on intestinal T cells. Thirty‐seven CD patients received three Ifx doses at week 0, 2 and 6. Blood samples, colonic biopsies and clinical scores were obtained before treatment and at week 8, 26 and 52. In‐vivo activated T‐cell cultures were established from colonic biopsies in the presence of interleukin (IL)‐2 and IL‐4. The in vitro effect of OPN stimulation on T‐cell IFN‐γ, TNF‐α, and IL‐10 production was measured. Plasma‐OPN was increased in active CD (increased CRP‐level) compared with quiescent disease (P = 0.02) and declined after three Ifx doses (P = 0.04). It was inversely correlated with in vitro T‐cell IL‐10 production. OPN increased CD69 and CD25 expression and enhanced T‐cell IFN‐γ and TNF‐α production in a dose‐dependent fashion with higher levels in CD than in healthy controls (HC), but induced a concomitant higher IL‐10 production in HC than CD. In conclusion, plasma‐OPN levels are related to CD inflammation. In vitro, OPN‐stimulated IL‐10 production increases less in T‐cell cultures from CD patients than from HC, indicating that IL‐10 deficiency may be involved in the defect immune‐regulation in CD, even after OPN stimulation.

The oxidation resistance proteins (OXR) help to protect eukaryotes from reactive oxygen species. The sole C-terminal domain of the OXR, named TLDc is sufficient to perform this function. However, the mechanism by which oxidation resistance occurs is poorly understood. We present here the crystal structure of the TLDc domain of the oxidation resistance protein 2 from zebrafish. The structure was determined by X-ray crystallography to atomic resolution (0.97Å) and adopts an overall globular shape. Two antiparallel β-sheets form a central β-sandwich, surrounded by two helices and two one-turn helices. The fold shares low structural similarity to known structures.