Renal Transplantation: Examination of the Regulation of Chemokine Binding During Acute Rejection

Abstract

Chemokines recruit leukocytes during allograft rejection. It is thought that the formation of glycosaminoglycan (GAG)-stabilized chemokine concentration gradients within the allograft plays a crucial role in this process. This raises the possibility that changes in GAG biology might regulate chemokine binding and the development of rejection. Immunocytochemical techniques were used to quantify changes in GAG expression within normal and rejection renal biopsy sections. Changes in GAG expression by cultured endothelial cell lines were also examined after stimulation with tumor necrosis factor-alpha and interferon-gamma. Quantitative reverse-transcriptase polymerase chain reaction was used to examine the basis for increased sulphation of heparan sulphate (HS) observed during inflammation. A binding assay was developed to determine how levels of GAG expression correlate with changes in chemokine (CCL5) sequestration. In normal kidney, HS was largely restricted to the tubular basement membrane; chondroitin-4-sulphate and chondroitin-6-sulphate were expressed within the interstitial tissues. The expression of all three GAGs was increased significantly during acute rejection, and heavily sulphated HS remained predominant within the tubular basement membrane. Treatment of endothelial cells with proinflammatory cytokines increased the expression of mRNA encoding N-deacetylase/N-sulphotransferase-1, an isoform of the enzyme responsible for N-sulphation of HS. Cytokine-treated cells and rejection biopsy specimens showed an enhanced capacity to bind CCL5. Chemokine production is known to be increased during acute renal allograft rejection. In this study we showed that the graft tissues also respond by increasing their potential to bind chemokines, a process that is vital for effective chemokine presentation and leukocyte recruitment.