Liposomal simvastatin attenuates neointimal hyperplasia in rats

Abstract

Monocytes, macrophages, and inflammation play a key role in the process of neointimal proliferation and restenosis. The present study evaluated whether systemic and transient depletion of monocytes could be obtained by a single intravenous (IV) injection of simvastatin liposomes, for the inhibition of neointima formation. Balloon-injured carotid artery rats (n = 30) were randomly assigned to treatment groups of free simvastatin, simvastatin in liposomes (3 mg/kg), and saline (control). Stenosis and neointima to media ratio (N/M) were determined 14 days following single IV injection at the time of injury by morphometric analysis. Depletion of circulating monocytes was determined by flow cytometry analyzes of blood specimens. Inhibition of RAW264.7, J774, and THP-1 proliferation by simvastatin-loaded liposomes and free simvastatin was determined by the 3-(4, 5-dimethylthiazolyl-2)-2, 5- diphenyltetrazolium bromide assay. Simvastatin liposomes were successfully formulated and were found to be 1.5-2 times more potent than the free drug in suppressing the proliferation of monocytes/macrophages in cell cultures of RAW 264.7, J774, and THP-1. IV injection of liposomal simvastatin to carotid-injured rats (3 mg/kg, n = 4) resulted in a transient depletion of circulating monocytes, significantly more prolonged than that observed following treatment with free simvastatin. Administration to balloon-injured rats suppressed neointimal growth. N/M at 14 days was 1.56 +/- 0.16 and 0.90 +/- 0.12, control and simvastatin liposomes, respectively. One single systemic administration of liposomal simvastatin at the time of injury significantly suppresses neointimal formation in the rat model of restenosis, mediated via a partial and transient depletion of circulating monocytes.

Fig. 1

The effect of liposomal simvastatin (IV, 3 mg/kg) on peripheral blood monocytes in the rat restenosis model. Representative flow cytometry analysis demonstrate the gated monocytes population (CD3⁻ CD4⁺), 48, 96, and 168 h after treatment with liposomal simvastatin in comparison to the free drug and saline. The treatment effect on circulating monocytes number (+SEM) over time is expressed as the percentage of total WBC (n = 4 at each group, *p < 0.05)

Fig. 2

Bar graph depicting reduced stenosis (upper) and neointima/media (N/M) ratio (lower) in liposomal simvastatin treated rats (3 mg/kg, IV, n = 9) 14 days after injury, in comparison to treatments with simvastatin solution (3 mg/kg, IV, n = 9, free drug) and saline (IV, n = 9, control). Below the bar graph are photomicrographs of Verhoeff’s tissue elastin staining of full size sections of control, free simvastatin, and liposomal-simvastatin-treated animals (*p < 0.05)