Decoy Oligodeoxynucleotides against the Transcription Factor AP-1 Decrease the Formation of Allograft Vasculopathy

Objectives: Allograft vasculopathy (AV) is the main limiting factor for long-term graft survival after solid organ transplantation. It is characterized by an increased MMP expression and activity resulting in migration of vascular smooth muscle cells (VSMCs), thereby enhancing neointimal formation and vascular lumen occlusion. The transcription factor activator protein 1 (AP-1) is critically involved in the pathogenesis of AV. AP-1 decoy oligodeoxynucleotides (AP-1 dODNs) are single stranded DNA molecules which neutralize AP-1 in the cytoplasm and nucleus.

Methods: Donor aortic allografts from DBA/2 mice were transplanted infrarenally into C57BL/6 mice. Allografts of the AP-1 dODN group (n = 7) were incubated for 30 minutes in a solution containing 10 µmol/L AP-1 dODNs and compared with a control group (n = 8). 10mg/kg bodyweight Ciclosporine A was administered intraperitoneally once daily. The grafts were harvested 30 days after transplantation. The intima-to-media (I/M) ratio and the degree of stenosis were quantified. The expression of MMP-2, MMP-9, MMP-12 as well as ACTA2, a marker specific for VSMCs or myofibroblasts, and the macrophage marker protein F4/80 were visualized by fluorescence microscopy followed by semiquantitative analysis of their presence within the neointima. The activity of MMP-2 and MMP-9 was evaluated by gelatin in situ zymography.

Results: The degree of stenosis and the I/M ratio were significantly reduced in the AP-1 dODN treated group by 23% (p = 0.03) and 37% (p = 0.011), respectively. The neointimal presence of VSMCs or fibroblasts was reduced (p = 0.05). No difference could be detected in the expression of MMP-2 (p = 0.874), MMP-9 (p = 0.201) and MMP-12 (p = 0.53) and in the activity of MMP-2 and MMP-9 (p = 0.238). In the AP-1 dODN group the number of macrophages correlates with the I/M ratio (r = 0.78; p = 0.037). This correlation did not exist in the control group (r = −0.24; p = 0.56).

Conclusion: AP-1 dODNs are able to reduce the degree of allograft vasculopathy in aortic transplantation of allogenic mice, demonstrated by a reduced number of VSMCs or myofibroblasts migrating into the neointima. The expression of MMP-2, MMP-9 and MMP-12 and the activity of MMP-2 and MMP-9 were not reduced by AP-1 dODNs in our model. In AP-1 treated mice, the invasion of macrophages contributes to the development of AV. The mechanism of the I/M ratio reduction caused by AP-1 dODNs needs to be further analyzed.