Suppression of Cell Proliferation and Macrophage Infiltration with Localized siRNA Delivery in a Murine Model of Intimal Hyperplasia

Abstract

Intimal hyperplasia limits the durability of peripheral vascular interventions and is associated with cell proliferation and macrophage infiltration. We use an in vivo rat carotid angioplasty model to evaluate inhibition of intimal hyperplasia with localized siRNA delivery targeting Thrombospondin-2 (TSP2) and evaluate its impact on cell proliferation (Ki-67) and macrophage infiltration (CD68). Intimal hyperplasia was induced by Fogarty catheter angioplasty vessel injury of the common carotid artery in 32 rats, which were divided into three control arms and one experimental arm (localized delivery of TSP2-suppressing siRNA). At postoperative day 21, bilateral common carotid arteries were harvested for evaluation of intimal hyperplasia via intima-to-media ratio calculation, PCR quantification of TSP2 mRNA expression, and immunohistochemical staining of Ki-67 proliferative marker and CD68 macrophage marker. RT-PCR data demonstrates successful TSP2 mRNA expression knockdown in the experimental arm compared with the control injury arm (0.49 ± 0.32 vs 20.5 ± 5.47; P = .027). The intima-to-media ratio, a measure of intimal hyperplasia, trended towards a significant decrease in the TSP2 siRNA treatment arm (1.34 ± 0.57 vs 1.94 ± 0.55; P = .056). Reduced macrophage infiltration (CD68 per mm2) was detected in the TSP2 siRNA treatment arm (23 ± 14 vs 72 ± 11; P = .029) as compared with the control injury arm. Furthermore, cell proliferation as measured by Ki-67 per mm2 (Figure) trended towards a significant decrease in the TSP2 siRNA treatment arm (27 ± 19 vs 69 ± 9; P = .067). Localized delivery of TSP2 siRNA can successfully limit the development of intimal hyperplasia in a rat carotid angioplasty model and decrease cell proliferative markers and macrophage infiltration. Further studies in larger animal models are warranted.