Background: PDI regulates cytoskeleton reorganization by the thiol-disulfide exchange in ␤-actin. Results: PDI directly binds to Cys 374 of ␤-actin during cell adhesion and spreading. Conclusion: Interaction of PDI with ␤-actin is induced by integrin-mediated cell adhesion and promotes cytoskeleton reorganization. Significance: PDI is a new regulator of the intramolecular disulfide-thiol rearrangement of ␤-actin in response to ␣IIb␤3 integrin engagement.

Recent evidence supports a role of protein-disulfide isomerase (PDI) in redox-controlled remodeling of the exofacial domains of ␣ IIb ␤ 3 in blood platelets. The aim of this study was to explain whether Ero1␣ can be responsible for extracellular reoxidation of the PDI active site. We showed that Ero1␣ can be found on platelets and is rapidly recruited to the cell surface in response to platelet agonists. It is physically associated with PDI and ␣ IIb ␤ 3 , as suggested by colocalization analysis in confocal microscopy and confirmed by immunoprecipitation experiments. Apart from monomeric oxidized Ero1␣, anti-␣ IIb ␤ 3 immunoprecipitates showed the presence of several Ero1␣-positive bands that corresponded to the complexes ␣ IIb ␤ 3 -PDIEro1␣, PDI-Ero1␣, and Ero1␣-Ero1␣ dimers. It binds more efficiently to the activated ␣ IIb ␤ 3 conformer, and its interaction is inhibited by RGD peptides. Ero1␣ appears to be involved in the regulation of ␣ IIb ␤ 3 receptor activity because of the following: (a) blocking the cell surface Ero1␣ by antibodies leads to a decrease in platelet aggregation in response to agonists and a decrease in fibrinogen and PAC-1 binding, and (b) transfection of MEG01 with Ero1␣ increases ␣ IIb ␤ 3 receptor activity, as indicated by increased binding of fibrinogen.

The aim of the study is proteomic analysis of the plasma profile in children with recurrent bone fractures. The study involved 16 children: 6 patients with recurrent low-energy fractures and normal bone mass and 10 with osteogenesis imperfecta. In the analysis of the protein profile, the two-dimensional protein electrophoresis was used (Ettan DALT II, Amersham Bioscience). The images of protein gels were compared with controls. The protein spots with changed expression were cut from the gel and the amino acid sequence was analyzed with the mass spectrometry method (Q-Tof Premier(TM) API MASS SPECTROMETR, Waters) for protein identification. The most prevalent protein with changed expression, with respect to controls, was haptoglobin observed in 6 patients with a severe form of osteogenesis imperfecta. Increased haptoglobin concentration in these patients was confirmed by the ELISA method. Peptides corresponding to alpha-1 acid glycoprotein and serum amyloid P-component, apolipoprotein A-I, and transthyretin were detected in one, two and three children, respectively. 1) The results show increased haptoglobin which may be suggestive of an inflammatory component taking part in the course of osteogenesis imperfecta. 2) Further studies to explain the possible relationship of this protein with increased bone fragility are necessary.

A novel affinity purification method for rapid isolation of vitronectin (VN) from human plasma is described. Recently we have used phage display technology to obtain clones expressing peptides with high binding activity for VN. The isolated "strong VN binders" were covalently coupled to CNBr-activated Sepharose. Human plasma was applied to the column and bound VN was eluted using 0.5 M acetic acid, giving purity exceeding 90%. The developed method is a convenient alternative to conventional antibody-antigen affinity chromatography techniques for purification of VN, as it offers low ligand cost, is rapid and ensures good protein recovery from human plasma.