Long-term effect of stents eluting 6-mercaptopurine in porcine coronary arteries

Abstract

Background: Drug-eluting stents (DES) have dramatically reduced restenosis rates compared to bare metal stents and are widely used in coronary artery angioplasty. The anti-proliferative nature of the drugs reduces smooth muscle cell (SMC) proliferation effectively, but unfortunately also negatively affects endothelialization of stent struts, necessitating prolonged dual anti-platelet therapy. Cell-type specific therapy may prevent this complication, giving rise to safer stents that do not require additional medication. 6-Mercaptopurine (6-MP) is a drug with demonstrated cell-type specific effects on vascular cells both in vitro and in vivo, inhibiting proliferation of SMCs while promoting survival of endothelial cells. In rabbits, we demonstrated that DES locally releasing 6-MP during 4 weeks reduced in-stent stenosis by inhibiting SMC proliferation and reducing inflammation, without negatively affecting endothelialization of the stent surface. The aim of the present study was to investigate whether 6-MP-eluting stents are similarly effective in preventing stenosis in porcine coronary arteries after 3 months, in order to assess the eligibility for human application.

Methods: 6-MP-eluting and polymer-only control stents (both n = 7) were implanted in porcine coronary arteries after local balloon injury to assess the effect of 6-MP on vascular lesion formation. Three months after implantation, stented coronary arteries were harvested and analyzed.

Results: Morphometric analyses revealed that stents were implanted reproducibly and with limited injury to the vessel wall. Unexpectedly, both in-stent stenosis (6-MP: 41.1 ± 10.3 %; control: 29.6 ± 5.9 %) and inflammation (6-MP: 2.14 ± 0.51; control: 1.43 ± 0.45) were similar between the groups after 3 months.

Conclusion: In conclusion, although 6-MP was previously found to potently inhibit SMC proliferation, reduce inflammation and promote endothelial cell survival, thereby effectively reducing in-stent restenosis in rabbits, stents containing 300 μg 6-MP did not reduce stenosis and inflammation in porcine coronary arteries.

Fig. 1

Characteristics and implantation of 6-MP DES in porcine coronary arteries. Polymer-only control stents (a) and stents loaded with 300 μg 6-MP (b) showed smooth coating surfaces by scanning electron microscopy. The cumulative release of 6-MP from the stents (c) was determined in vitro for up to 38 days (panels a-c adapted from [18]). After applying endothelial damage by balloon inflation, stents were deployed in the RCA (d), LCX (e) and LAD (f). Stent placement was followed by control angiography to ensure patency of the stented coronary artery. After 3 months, control angiography was repeated to reassess patency. After euthanasia, the heart was rapidly excised and the stents, visible as indicated (g), were harvested and fixed. Arrows indicate location of the stents

Fig. 2

Characterization of stented coronary arteries 3 months after placement. Sections of arteries implanted with polymer-only stents (control) or 6-MP-eluting stents (6-MP) were stained with H&E to assess general histology of the stented vessel wall (a, d). Masson trichrome staining visualized the adventitia (Adv), media (Med) and intima (Int) of the vessel wall as well as strut holes (s) (b, e). Lawson-Van Gieson staining was applied to stain the internal and external elastic lamina and thus quantification of the different layers of the vessel wall (c, f), enabling morphometric analysis. Endothelial cell coverage of the vessel wall was demonstrated by immunohistochemical staining with an antibody against vWF (g). The media largely consists of circumferentially aligned SMCs, whereas SMCs in the intima are oriented longitudinally in the vessel wall, as shown by staining with an antibody directed against αSMA (h). Most cells positive for the cell-cycle inhibitor p27^kip1 are localized in media and adventitia, whereas hardly any positive cells were found in the intima, indicating that especially in the intima cells are proliferating (i)

Fig. 3

Morphometric analyses after 3 months of the stented coronary arteries. The outer diameter of the stented arteries (a) was similar within and between the groups, indicating high reproducibility of stent implantation and expansion. The injury score was low in all stents and similar between groups (b). Lumen stenosis (c) showed large variation within the groups and was similar between 6-MP and control stents. Inflammation score (d) was also similar between control and 6-MP. Bars depict mean values, error bars represent SE