Allo immune response, epithelial mesenchymal transition and chronic allograft fibrosis – Is there a link?

Abstract

Introduction Mechanisms of chronic rejection manifested by irreversible fibrosis remain unclear. Phenotypic conversion of renal tubular epithelial and endothelial cells of the allograft to mesenchymal cells [Epithelial/Endothelial to Mesenchymal Transitioning (EMT/EndOEMT)] in the onset of fibrosis in the transplanted organ is proposed hypothesis. Objectives To detect EMT by way of gene expression profiling related to EMT using qualitative CR and quantitative real-time PCR covering 84 genes related to EMT. Materials: Anti-HLA Class I chimeric monoclonal antibody to the constant region of HLA Class I antigen (Invivogen, San Diego CA), EMR8-5: (Abcam ® ,Cambrige MA), a monoclonal HLA-Class I antibody that reacts with the heavy chains of Human HLA Class I A, B, and C and Pooled Positive Serum (PPS) containing antibodies to all Class I and Class II HLA allo-antigen and some possible non-HLA antigens. Renal Epithelial Cell (REC) and Human Umbilical Endothelial Cell (HUEC) lines (Life Line Cell Technologies, Frederick MD) were used. A total of 37 paraffin embedded renal allograft biopsy samples with varying etiologies and varying percentages of fibrosis were retrieved from Rush University Medical Center in Chicago, IL. An EMT RT 2 profiler PCR Array (Qiagen) specific for 84 EMT-related genes was used to assess the expression of these genes in 12 renal allograft biopsies and 2 cell cultures Results and conclusions The EMT Gene array profiling by realtime PCR indicated that in both in vitro cell cultures and biopsies there was significant changes in EMT related gene expressions. The overall gene expression pattern revealed up regulation of several EMT related genes in renal epithelial cells, HUEC, and in biopsy samples. The biopsy sample findings on EMT related gene expression array point in the direction of contributing towards finding a molecular signature of chronic allograft rejection.