Purpose: To investigate protein differences in renal allografts to provide insight into the pathophysiology of cellular rejection. The objective is to determine which proteins are most different in abundance so that these may be targeted as urine biomarkers of allograft rejection.
Methods: Retrospectively, five patients with Banff class 1B rejection were identified and case matched with five control patients that had no identified pathology on biopsy. Excess frozen renal allograft biopsy tissue was retrieved and washed in cold PBS. Tissue was homogenized and proteins digested with trypsin. Tryptic peptides were analyzed by LC/MS/MS (liquid chromatography tandem mass spectrometry) and identified by MASCOT. Normalized spectral counts were used to estimate protein abundance. Groups were compared by the Wilcoxon Rank-Sum test and the mean fold-change (MFC), which were used to construct a volcano plot. Candidate biomarkers were selected using the geometric distance from the origin of the volcano plot. Immunohistochemistry was used to localize Serpin A5.
Results: A total of 721 proteins (FDR 0.4%) were identified. 18 proteins were lower in abundance in the rejection group compared to controls. SerpinA5 appeared to be the least abundant protein, based on the combination of its low p-value (<0.01) and high mean fold-change (MFC=8.92). Based on immunostaining, Serpin A5 was localized to the distal tubular cells and brush border of proximal tubules. Only two proteins were elevated in the rejection group compared to the control group: Transglutaminase C and IgM chain C region.
Conclusion: Proteomic analysis of frozen biopsy samples revealed novel differences between rejected and non-rejected tissue, notably a consistently lower level of Serpin A5 and elevated level of Transglutaminase C. Elevated Transglutaminase C has been described in chronic allograft nephropathy, but the role of Serpin A5 in allograft pathology and role of both these proteins specifically in allograft rejection is not known. Further studies in the use of these as urinary markers for rejection are ongoing.
To cite this abstract in AMA style:Talwar M, Ahmed V, Srinivas T, Bhensdadia N, Alison B, Alge J, Self S, Arthur J, Janech M. Tissue Proteomics Reveals Protein Differences Associated with Allograft Cellular Rejection [abstract]. Am J Transplant. 2013; 13 (suppl 5). http://atcmeetingabstracts.com/abstract/tissue-proteomics-reveals-protein-differences-associated-with-allograft-cellular-rejection/. Accessed September 23, 2017.
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