Plasma proteome changes associated with refractory cytopenia with multilineage dysplasia

Abstract

Background: Refractory cytopenia with multilineage dysplasia (RCMD) is a subgroup of myelodysplastic syndrome (MDS), which belongs to oncohematological diseases, occurring particularly in elderly patients, and represents a heterogeneous group of bone marrow diseases. The goal of this study was to look for plasma proteins that changed quantitatively or qualitatively in RCMD patients.

Results: A total of 46 plasma samples were depleted, proteins were separated by 2D SDS-PAGE (pI 4-7), and proteomes were compared using Progenesis SameSpots statistical software. Proteins were identified by nanoLC-MS/MS. Sixty-one unique, significantly (p < 0.05, ANOVA) different spots were found; proteins in 59 spots were successfully identified and corresponded to 57 different proteins. Protein fragmentation was observed in several proteins: complement C4-A, complement C4-B, inter-alpha-trypsin inhibitor heavy chain H4, and endorepellin.

Conclusions: This study describes proteins, which change quantitatively or qualitatively in RCMD patients, and represents the first report on significant alterations in C4-A and C4-B complement proteins and ITIH4 fragments in patients with MDS-RCMD.

Figure 1

Positions of significantly differed spots on a 2D gel. Positions of all spots that were found to significantly differ in 2D gels of RCMD patients and healthy controls when mutually compared. The 2D gel of a patient sample was used as an illustrative gel to display spot positions.

Figure 2

ITIH4 western blot analysis. (A) A segment of 2D SDS-PAGE gel containing spots with uncleaved ITIH4 (spots 43, 50, 53, 60, and 61) and (B) 2D western blot analysis (ITIH4) - a segment corresponding to the highlighted area on the 2D gel; (C) illustrations of 1D western blot analysis of ITIH4 and its bands in both the control and RCMD groups corresponding to the molecular weight of 2D western blot (and 2D SDS-PAGE) ITIH4 spots are shown.

Figure 3

Principal Component Analysis. PCA was performed to assess whether grouping of patients and healthy controls based on proteomic methods reflects their stratification using classical clinical diagnosis. Analysis was based on spots that significantly differ according the mentioned statistical criteria (p < 0.05, ANOVA). Principal Component Analysis (PCA) showed the separation of all samples into two aggregates that corresponded to the RCMD (blue dots) and the control group (pink dots).