In vitro biocompatibility assessment of a novel potential drug eluting stent coating in human coronary artery endothelial cells

McGillicuddy F1, O’Toole D1, Hickey J1, Lynch I2, Dawson KA2, Keenan AK1. 1UCD School of Biomolecular and Biomedical Research, Conway Institute; 2UCD School of Chemistry and Chemical Biology, University College Dublin, Ireland.

While N-isopropylacrylamide-derived copolymers have recently been investigated as potential vehicles for local delivery of fluvastatin from coronary stents (McGillicuddy F et al., 2004) , little is known about the pro-inflammatory effects of such coatings on coronary vascular endothelium. This study aimed to determine the effects of fluvastatin on human coronary artery endothelial cell (HCAEC) proliferation and viability and to assess whether the presence of fluvastatin in these coatings could limit pro-inflammatory responses in these cells.

Stainless steel stent wires were coated with fluvastatin-loaded N-isopropylacrylamide/N-tertbutylacrylamide copolymer (NiPAAm/NtBAAm, 65/35 w/w ratio) microgels dispersed in a NiPAAm/NtBAAm (85/15 w/w ratio) copolymer matrix (Lynch & Dawson, 2003) . Effects of increasing fluvastatin concentrations (0.1-1 μM) on proliferation (BrdU ELISA) of HCAEC and human coronary artery smooth muscle cells (HCASMC) were first compared. Subsequently, effects of fluvastatin on HCAEC viability (MTT assay) and cytoskeletal actin arrangement (rhodamine-phalloidin staining) were determined. The effects of bare-metal and copolymer-coated ( ± fluvastatin) stent wires on ICAM-1, VCAM-1, E-selectin and IL-8 mRNA expression in HCAEC were investigated by RT-PCR analysis. Effects of stent wires on VCAM-1 protein expression were examined using immunofluorescence microscopy. IL-8 and TNF-α secretion into overlying medium after incubation with stent wires placed in the vicinity of HCAEC was assessed by ELISA. Significant differences between treatments were identified by one-way ANOVA.

At all concentrations tested, HCASMC were more sensitive than HAEC to the anti-proliferative effects of fluvastatin, with a significant reduction (p<0.001, n=8) in SMC number to 72.06 ± 2.46 % of control observed after treatment with 0.1 μM fluvastatin whilst no effect was seen on HCAEC proliferation ( 102.1 ± 3.9 % of control, NS, n=8) at this concentration. A loss of HCAEC viability was only evident upon 48 h treatment with 10 m M fluvastatin. Incubation with 1 μM fluvastatin resulted in reduced stress fibre expression in the actin cytoskeleton of HCAEC. Incubation with stent wires ( ± coating, ± fluvastatin) failed to induce mRNA expression of any of the inflammatory markers, secretion of IL-8 or TNF-α into overlying medium, or induction of VCAM-1 protein expression. However, a loss of endothelial cell viability was evident due to mechanical impact of stent wires on the cells. This mechanical impact mimics endothelial denudation which is an important contributing factor to in-stent restenosis in vivo (Gershlick, 2002) . In conclusion, no evidence was found that this novel copolymer coating induced an inflammatory response in vitro strengthening its potential as a biocompatible drug-eluting stent coating in vivo.

Gershlick, A.H. (2002). Atherosclerosis, 160, 259-271.
Lynch, I. & Dawson, K. (2003). J Phys Chem B, 107, 9629-9637.
McGillicuddy F, et al., (2004). Am J Cardiol, 95, 224E.

 Supported by the Irish Health Research Board.