Detection of Various Epitopes of Murine Osteopontin by Monoclonal Antibodies

Hara, Hiroshi; Kawa, Shigeyuki; Katagiri, Yohko U.; Tosa, Noriko; Tsukamoto, Taiji; Chambers, Ann F.; Uede, Toshimitsu

doi:10.1006/bbrc.1999.0412

Cited by 12 publications

(10 citation statements)

References 19 publications

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“…Immunohistochemistry was performed on paraffin sections according to the avidin–biotin method, as previously described (8, 20). Briefly, 3‐μm deparaffinized sections were pretreated with 10 m M citrate buffer, pH 6.0, in a microwave oven for 20 minutes for antigen retrieval.…”

Section: Methodsmentioning

confidence: 99%

“…Briefly, 3‐μm deparaffinized sections were pretreated with 10 m M citrate buffer, pH 6.0, in a microwave oven for 20 minutes for antigen retrieval. Rat monoclonal IgG anti‐mouse OPN antibody (OPN2.2) (20), hamster monoclonal IgG anti‐mouse CD51 (αv integrin) antibody (H9.2B8; PharMingen, San Diego, CA), and hamster monoclonal IgG anti‐mouse CD61 (β3 integrin) monoclonal antibody (HMb3; PharMingen) were employed as primary antibodies. An equal concentration of either purified rat IgG or hamster IgG (both from PharMingen) was used as a negative control.…”

Section: Methodsmentioning

confidence: 99%

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Expression of osteopontin at sites of bone erosion in a murine experimental arthritis model of collagen‐induced arthritis: Possible involvement of osteopontin in bone destruction in arthritis

Ohshima¹,

Kobayashi²,

Yamaguchi³

et al. 2002

Arthritis & Rheumatism

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Objective. To investigate the involvement of osteopontin (OPN) in bone destruction in a murine experimental arthritis model of collagen-induced arthritis (CIA).Methods. The expression of OPN was examined at both the messenger RNA (mRNA) and protein levels in various arthritic lesions in mice with CIA by in situ hybridization and immunohistochemistry, respectively. In addition, the expression of ␣v␤3 integrin, a receptor for OPN, the ligation of which is thought to be essential for bone resorption by osteoclasts, was examined by immunohistochemistry. Plasma concentrations of OPN were measured at different time points in the course of CIA by enzyme-linked immunosorbent assay.Results. OPN mRNA was detected mainly at sites of bone erosion in arthritic lesions, where activated osteoclasts were present; OPN protein was also detected at sites of bone erosion. In the arthritic synovium, OPN was predominantly expressed in the synovial lining layer, but not in lymphoid aggregates. In addition, ␣v␤3 integrin was detected coincident with OPN at sites of bone erosion (bone-pannus junction). Plasma OPN levels were markedly elevated at the time points that corresponded to arthritis flares, and higher levels were maintained during the progression of arthritis.Conclusion. OPN may mediate bone resorption by osteoclasts in arthritis through ligation with its receptor, ␣v␤3 integrin. OPN may be a useful therapeutic target molecule in the prevention of bone destruction in arthritis.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Expression of osteopontin at sites of bone erosion in a murine experimental arthritis model of collagen‐induced arthritis: Possible involvement of osteopontin in bone destruction in arthritis

Ohshima¹,

Kobayashi²,

Yamaguchi³

et al. 2002

Arthritis & Rheumatism

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“…10 A monoclonal antibody, reacting against ␣-smooth muscle actin, clone 1A4, was purchased from Nichirei, Tokyo, Japan. A monoclonal antibody, reacting against macrophage, MOMA-2, was purchased from Serotec, Tokyo, Japan.…”

Section: Immunohistochemistrymentioning

confidence: 99%

Osteopontin Deficiency Attenuates Atherosclerosis in Female Apolipoprotein E–Deficient Mice

Matsui

Rittling

Okamoto

et al. 2003

ATVB

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Objective-Osteopontin (OPN), a noncollagenous adhesive protein, is implicated in atherosclerosis, in which macrophages within atherosclerotic plaques express OPN. However, it is not known whether the elevated OPN expression is a cause or result of atherosclerosis. Methods and Results-We generated mice that lacked OPN and crossed them with apolipoprotein (apo) E-deficient mice and analyzed these mice with a mixed C57BL/6ϫ129 background after 36 weeks on a normal chow diet. In female mice, OP ϩ/Ϫ E Ϫ/Ϫ and OP Ϫ/Ϫ E Ϫ/Ϫ mice had significantly smaller atherosclerotic and inflammatory lesions compared with OP ϩ/ϩ E Ϫ/Ϫ mice, and that was reflected by smaller area of MOMA-2-positive staining. In male mice, however, there was no significant difference in the atherosclerosis lesion areas among 3 genotypes. In both OP Ϫ/Ϫ E Ϫ/Ϫ and OP ϩ/ϩ E Ϫ/Ϫ mice, typical atherosclerotic lesions were detected, which include necrotic core, foamy cell collections, and cholesterol clefts. However, we found that vascular mineral-deposited areas in 60-week-old male OP Ϫ/Ϫ E Ϫ/Ϫ mice were significantly increased compared with those in OP ϩ/ϩ E Ϫ/Ϫ male mice. Key Words: osteopontin Ⅲ atherosclerosis Ⅲ macrophage Ⅲ calcification Ⅲ lipid metabolism A therosclerosis is characterized as a chronic inflammatory process of the vessel wall. Atherosclerosis is initiated by the infiltration of monocytes and T-lymphocytes into activated endothelium, followed by their migration into the intima and subsequent lipid accumulation within macrophages. In late stages of atherosclerosis, calcification is a common advanced complication. Osteopontin (OPN), a noncollagenous adhesive protein, was first found at sites of dystrophic calcification and is synthesized at high levels by macrophages in calcified aortic valves and atherosclerotic plaques. 1 The expression of OPN protein was detected in not only macrophages but also vascular smooth muscle cells within atherosclerotic lesion. [1][2][3][4][5] In addition, it was shown that vascular smooth muscle cells during the proliferative and migratory phase, but not the quiescent and contractile phase, expressed OPN in a model of balloon catheter injury of rat carotid artery. 2 More importantly, Liaw et al 6 reported that neutralizing antibodies directed against OPN inhibited rat carotid neointimal thickening after endothelial denudation. Taken together, these results suggested that OPN can play a pivotal role in the early stage of atherosclerosis, including proliferation and migration of smooth muscle cells as well as at the late stage of atherosclerosis, characterized by calcified atheromatous plaque formation. Conclusions-TheseTo more directly address the question of whether OPN initiates the development of atherosclerotic lesion and plays a role in calcification, we took advantage of OPN-deficient mice we had recently generated. 7 We crossed them with apolipoprotein (apo) E-deficient mice and made OPN and apoE double-deficient mice. In mice deficient for OPN gene expression, OPN mRNA was not detected in any organs. ...

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“…Previous studies have demonstrated that the detectability of the native OPN molecule by immunoblot depends on epitopes which antibodies can recognize (28). Therefore, we tested two types of antibodies which recognize different domains on an OPN protein, and utilized anti-C-OPN antibody that recognizes the C-terminal end of the OPN molecule.…”

Section: Figmentioning

confidence: 99%

Pure Atmospheric Pressure Promotes an Expression of Osteopontin in Human Aortic Smooth Muscle Cells

Iizuka

Murakami

Kawaguchi

2001

Biochemical and Biophysical Research Communications

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Detection of Various Epitopes of Murine Osteopontin by Monoclonal Antibodies

Cited by 12 publications

References 19 publications

Expression of osteopontin at sites of bone erosion in a murine experimental arthritis model of collagen‐induced arthritis: Possible involvement of osteopontin in bone destruction in arthritis

Expression of osteopontin at sites of bone erosion in a murine experimental arthritis model of collagen‐induced arthritis: Possible involvement of osteopontin in bone destruction in arthritis

Osteopontin Deficiency Attenuates Atherosclerosis in Female Apolipoprotein E–Deficient Mice

Pure Atmospheric Pressure Promotes an Expression of Osteopontin in Human Aortic Smooth Muscle Cells

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