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Comparative proteomic analysis of serum from patients with systemic sclerosis and sclerodermatous GVHD. Evidence of defective function of factor H.

Scambi C, La Verde V, De Franceschi L, Barausse G, Poli F, Benedetti F, Sorio M, Deriu F, Roncada P, Bortolami O, Turrini F, Caramaschi P, Stranieri C, Bambara LM, Biasi D - PLoS ONE (2010)

Bottom Line: In addition, we observed that FH binding to ECs was reduced when we used serum from these patients.The comparative proteomic analysis of serum from SSc and ScGVHD patients highlighted proteins involved in either promoting or maintaining an inflammatory state.We also found a defective function of Factor H, possibly associated with ECs damage.

View Article: PubMed Central - PubMed

Affiliation: Section of Rheumatology, Department of Clinical and Experimental Medicine, University of Verona, Verona, Italy. cinzia.scambi@univr.it

ABSTRACT

Background: Systemic sclerosis (SSc) is an autoimmune disease characterized by immunological and vascular abnormalities. Until now, the cause of SSc remains unclear. Sclerodermatous graft-versus-host disease (ScGVHD) is one of the most severe complications following bone marrow transplantation (BMT) for haematological disorders. Since the first cases, the similarity of ScGVHD to SSc has been reported. However, both diseases could have different etiopathogeneses. The objective of this study was to identify new serum biomarkers involved in SSc and ScGVHD.

Methodology: Serum was obtained from patients with SSc and ScGVHD, patients without ScGVHD who received BMT for haematological disorders and healthy controls. Bi-dimensional electrophoresis (2D) was carried out to generate maps of serum proteins from patients and controls. The 2D maps underwent image analysis and differently expressed proteins were identified. Immuno-blot analysis and ELISA assay were used to validate the proteomic data. Hemolytic assay with sheep erythrocytes was performed to evaluate the capacity of Factor H (FH) to control complement activation on the cellular surface. FH binding to endothelial cells (ECs) was also analysed in order to assess possible dysfunctions of this protein.

Principal findings: Fourteen differentially expressed proteins were identified. We detected pneumococcal antibody cross-reacting with double stranded DNA in serum of all bone marrow transplanted patients with ScGVHD. We documented higher levels of FH in serum of SSc and ScGVHD patients compared healthy controls and increased sheep erythrocytes lysis after incubation with serum of diffuse SSc patients. In addition, we observed that FH binding to ECs was reduced when we used serum from these patients.

Conclusions: The comparative proteomic analysis of serum from SSc and ScGVHD patients highlighted proteins involved in either promoting or maintaining an inflammatory state. We also found a defective function of Factor H, possibly associated with ECs damage.

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Related in: MedlinePlus

2D serum maps of a patient with diffuse SSc and a healthy control.1200 µg of total serum proteins were focused on non-linear pH 3–10 immobiline dry strips and then separated into 9–16% polyacrilamide gels, which were stained with colloidal Coomassie. Marked are the protein spots, which have been identified by mass spectrometry.
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pone-0012162-g002: 2D serum maps of a patient with diffuse SSc and a healthy control.1200 µg of total serum proteins were focused on non-linear pH 3–10 immobiline dry strips and then separated into 9–16% polyacrilamide gels, which were stained with colloidal Coomassie. Marked are the protein spots, which have been identified by mass spectrometry.

Mentions: This approach allowed us to highlight seven protein spots in pH 3–10 2D maps (Fig. 2) and six in pH 4–5.5 2D maps (Fig. 3), which were differently expressed in SSc patients, compared to healthy controls. These protein spots were excised from the gels and analysed by mass spectrometry (Table 3). We identified thirteen proteins, that were divided into five functional clusters: (i) cell protection, (ii) cell proliferation, (iii) acute phase response, (iv) immune response (v) host infection. Concentration differences of each identified protein had been expressed as ratio between the mean volume of the spot in each subgroup of patients and in healthy controls (Table 3).


Comparative proteomic analysis of serum from patients with systemic sclerosis and sclerodermatous GVHD. Evidence of defective function of factor H.

Scambi C, La Verde V, De Franceschi L, Barausse G, Poli F, Benedetti F, Sorio M, Deriu F, Roncada P, Bortolami O, Turrini F, Caramaschi P, Stranieri C, Bambara LM, Biasi D - PLoS ONE (2010)

2D serum maps of a patient with diffuse SSc and a healthy control.1200 µg of total serum proteins were focused on non-linear pH 3–10 immobiline dry strips and then separated into 9–16% polyacrilamide gels, which were stained with colloidal Coomassie. Marked are the protein spots, which have been identified by mass spectrometry.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2921355&req=5

pone-0012162-g002: 2D serum maps of a patient with diffuse SSc and a healthy control.1200 µg of total serum proteins were focused on non-linear pH 3–10 immobiline dry strips and then separated into 9–16% polyacrilamide gels, which were stained with colloidal Coomassie. Marked are the protein spots, which have been identified by mass spectrometry.
Mentions: This approach allowed us to highlight seven protein spots in pH 3–10 2D maps (Fig. 2) and six in pH 4–5.5 2D maps (Fig. 3), which were differently expressed in SSc patients, compared to healthy controls. These protein spots were excised from the gels and analysed by mass spectrometry (Table 3). We identified thirteen proteins, that were divided into five functional clusters: (i) cell protection, (ii) cell proliferation, (iii) acute phase response, (iv) immune response (v) host infection. Concentration differences of each identified protein had been expressed as ratio between the mean volume of the spot in each subgroup of patients and in healthy controls (Table 3).

Bottom Line: In addition, we observed that FH binding to ECs was reduced when we used serum from these patients.The comparative proteomic analysis of serum from SSc and ScGVHD patients highlighted proteins involved in either promoting or maintaining an inflammatory state.We also found a defective function of Factor H, possibly associated with ECs damage.

View Article: PubMed Central - PubMed

Affiliation: Section of Rheumatology, Department of Clinical and Experimental Medicine, University of Verona, Verona, Italy. cinzia.scambi@univr.it

ABSTRACT

Background: Systemic sclerosis (SSc) is an autoimmune disease characterized by immunological and vascular abnormalities. Until now, the cause of SSc remains unclear. Sclerodermatous graft-versus-host disease (ScGVHD) is one of the most severe complications following bone marrow transplantation (BMT) for haematological disorders. Since the first cases, the similarity of ScGVHD to SSc has been reported. However, both diseases could have different etiopathogeneses. The objective of this study was to identify new serum biomarkers involved in SSc and ScGVHD.

Methodology: Serum was obtained from patients with SSc and ScGVHD, patients without ScGVHD who received BMT for haematological disorders and healthy controls. Bi-dimensional electrophoresis (2D) was carried out to generate maps of serum proteins from patients and controls. The 2D maps underwent image analysis and differently expressed proteins were identified. Immuno-blot analysis and ELISA assay were used to validate the proteomic data. Hemolytic assay with sheep erythrocytes was performed to evaluate the capacity of Factor H (FH) to control complement activation on the cellular surface. FH binding to endothelial cells (ECs) was also analysed in order to assess possible dysfunctions of this protein.

Principal findings: Fourteen differentially expressed proteins were identified. We detected pneumococcal antibody cross-reacting with double stranded DNA in serum of all bone marrow transplanted patients with ScGVHD. We documented higher levels of FH in serum of SSc and ScGVHD patients compared healthy controls and increased sheep erythrocytes lysis after incubation with serum of diffuse SSc patients. In addition, we observed that FH binding to ECs was reduced when we used serum from these patients.

Conclusions: The comparative proteomic analysis of serum from SSc and ScGVHD patients highlighted proteins involved in either promoting or maintaining an inflammatory state. We also found a defective function of Factor H, possibly associated with ECs damage.

Show MeSH
Related in: MedlinePlus