Expression of extra domain A containing fibronectin in chronic cardiac allograft rejection

Abstract

Cardiac allograft vasculopathy (CAV) and fibrosis are important in chronic cardiac allograft rejection. The aim of our study was to analyze the up-regulation of extra domain A (ED-A) containing fibronectin (ED-A(+) Fn) in cardiac allografts after heterotopic rat heart transplantation using a human recombinant antibody applicable for targeted drug delivery. Cardiac allografts were subjected to immunofluorescence double labelling procedures combining a human recombinant small immunoprotein (SIP) format antibody recognizing ED-A(+) Fn (F8) with antibodies recognizing CD31, ASMA or CD45. Protein expression levels of ED-A(+) Fn were measured by quantitative confocal laser scanning microscopy and messenger RNA expression levels by real-time reverse-transcription polymerase chain reaction. A distinct re-expression of ED-A(+) Fn was detectable with the F8 antibody, especially in vessel structures exhibiting CAV and in fibrotic areas. ED-A(+) Fn protein deposition but not messenger RNA expression levels increased with rising rejection grade (p ≤ 0.001). There were clear co-localizations of ED-A(+) Fn and α-smooth muscle actin in vessels and in fibrotic areas. We could show first that ED-A(+) Fn is expressed in rat cardiac allografts in association with CAV and cardiac fibrosis. The protein is detectable with the human recombinant antibody F8 usable for targeted drug delivery to the side of disease. Second, protein expression levels increase with rising rejection grade. Thus, ED-A(+) Fn might be usable to monitor and target CAV as well as fibrosis after heart transplantation.