Effects of survival and proliferation of smooth muscle cells (SMC) by supercooling and re-warming

Abstract

Background: SMC plays a critical role in neointimal hyperplasia after angioplasty. Studies showed that cryoplasty produces less restenosis clinically, however, its underlying mechanisms are still not well understood. In this study, we investigated the effects of SMC survival and proliferation after supercooling and re-warming in an in-vitro model simulating cryoplasty. Methods: Bovine aortic smooth cells were cultured in 6 well plates with medium supplemented with 10% fetal bovine serum. The samples were supercooled to around -10oC for 0 and 60 seconds using a conduction cooling stage and then re-warmed in an incubator at 37oC for 0,6, 12 or 24 hours. TUNEL assay and BrdU were used to measure the degree of apoptosis and proliferation respectively. Western blot measured the activation of p70S6 kinase and p44/42MAP kinase and were quantified using densitometry. Results are given as mean + standard error of mean and analysed by ANOVA. Results: Significant increased SMC apoptosis and decreased proliferation were observed with increasing supercooling and re-warming time. Both p70S6 kinase and p44/42 MAP kinase showed no changes with supercooling and re-warming. Conclusions: The decreased proliferation and increased apoptosis of SMC after supercooling and re-warming may explain why cryoplasty can produce less restenosis. No significant changes in p70S6 kinase and p44/42 MAP kinase level may indicate that other pathways may be involved in both transcription and translation control of SMC after supercooling and re-warming.