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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

Factor H binding to endothelial cells.HUVEC were incubated in patient and control sera (dSSc n = 8; lSSc n = 6; ScGVHD n = 3; healthy n = 5) and bound FH was visualized with a specific antibody by flow cytometry. The results obtained were compared using box and whiskers plot. Figure inset. Two subtypes of dSSc patients are represented: patients with important pulmonary involvement (dSSc lung, n = 4) and patients with exclusive cutaneous involvement (dSSc skin, n = 4). The results obtained were compared using box and whiskers plot.
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pone-0012162-g008: Factor H binding to endothelial cells.HUVEC were incubated in patient and control sera (dSSc n = 8; lSSc n = 6; ScGVHD n = 3; healthy n = 5) and bound FH was visualized with a specific antibody by flow cytometry. The results obtained were compared using box and whiskers plot. Figure inset. Two subtypes of dSSc patients are represented: patients with important pulmonary involvement (dSSc lung, n = 4) and patients with exclusive cutaneous involvement (dSSc skin, n = 4). The results obtained were compared using box and whiskers plot.

Mentions: To validate these results, the in vitro binding of FH to human ECs was investigated with HUVECs, which had been used as model of self-cells. We incubated HUVECs with human sera from patients and healthy subjects (lSSc n = 6; dSSc n = 8; ScGVHD n = 3; healthy n = 5) and the binding of native FH was measured by flow cytometry. We observed reduced surface-bound FH when HUVECs were incubated with the serum from dSSc patients (Fig. 8), in particular in dSSc patients with pulmonary involvement (Figure inset). For the other cases, the large variability in FH cell binding documented for lSSc patients and the limited number of ScGVHD patients that we analysed, did not allow us to formulate a conclusion.


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)

Factor H binding to endothelial cells.HUVEC were incubated in patient and control sera (dSSc n = 8; lSSc n = 6; ScGVHD n = 3; healthy n = 5) and bound FH was visualized with a specific antibody by flow cytometry. The results obtained were compared using box and whiskers plot. Figure inset. Two subtypes of dSSc patients are represented: patients with important pulmonary involvement (dSSc lung, n = 4) and patients with exclusive cutaneous involvement (dSSc skin, n = 4). The results obtained were compared using box and whiskers plot.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0012162-g008: Factor H binding to endothelial cells.HUVEC were incubated in patient and control sera (dSSc n = 8; lSSc n = 6; ScGVHD n = 3; healthy n = 5) and bound FH was visualized with a specific antibody by flow cytometry. The results obtained were compared using box and whiskers plot. Figure inset. Two subtypes of dSSc patients are represented: patients with important pulmonary involvement (dSSc lung, n = 4) and patients with exclusive cutaneous involvement (dSSc skin, n = 4). The results obtained were compared using box and whiskers plot.
Mentions: To validate these results, the in vitro binding of FH to human ECs was investigated with HUVECs, which had been used as model of self-cells. We incubated HUVECs with human sera from patients and healthy subjects (lSSc n = 6; dSSc n = 8; ScGVHD n = 3; healthy n = 5) and the binding of native FH was measured by flow cytometry. We observed reduced surface-bound FH when HUVECs were incubated with the serum from dSSc patients (Fig. 8), in particular in dSSc patients with pulmonary involvement (Figure inset). For the other cases, the large variability in FH cell binding documented for lSSc patients and the limited number of ScGVHD patients that we analysed, did not allow us to formulate a conclusion.

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