Prevention of Simvastatin-Induced Inhibition of Tendon Cell Proliferation and Cell Cycle Progression by Geranylgeranyl Pyrophosphate.

Wen-Chung Tsai,Cheng-Lun Chen,Tung-Yang Yu,Mei-Ling Cheng,Li-Ping Lin,Jong-Hwei S Pang

doi:10.1093/toxsci/kfv239

Abstract

Statins have been reported to induce tendinopathy and even tendon rupture. The present study was designed to investigate the potential molecular mechanism underlying the adverse effect of simvastatin on tendon cells. An invitro tendon healing model was performed using tendon cells isolated from rat Achilles tendons. The viability of tendon cells and cell cycle progression were examined by the MTT assay and flow cytometric analysis, respectively. Immunofluorescent staining for Ki-67 was used to assess the proliferation activity of tendon cells. Western blot analysis and coimmunoprecipitation was used to determine the protein expression of cell cycle-related proteins. To investigate the potential mechanism underlying the effect of statins on tendon cells, mevalonate, farnesyl pyrophosphate (FPP), or geranylgeranyl pyrophosphate (GGPP) was added to simvastatin-treated tendon cells. Simvastatin inhibited the invitro tendon healing model and tendon cell proliferation in a dose-dependent manner. Immunofluorescent staining demonstrated reduced ki-67 expression in simvastatin-treated tendon cells. Furthermore, simvastatin induced cell cycle arrest at the G1 phase. The expression levels of cdk1, cdk2, cyclin A, and cyclin E were downregulated by simvastatin in a dose-dependent manner. The inhibitory effect of simvastatin was proved to mediate the reduction of mevalonate, and the addition of exogenous GGPP completely prevented the inhibitory effect of simvastatin on tendon cells. The present study demonstrated, for the first time, the molecular mechanism underlying simvastatin-induced tendinopathy or tendon rupture. GGPP was shown to prevent the adverse effect of simvastatin in tendon cells without interfering with its cholesterol-reducing efficacy.

Highlights

Statins are a class of lipid-lowering drugs clinically used to inhibit 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA), a key enzyme in the mevalonate metabolic pathway for endogenous cholesterol production
Simvastatin Reduced the Number of Viable Tendon Cells Tendon cells were treated with various concentrations of simvastatin for 24 h, and both the MTT assay and Lactate dehydrogenase (LDH) cytotoxicity assay were used to examine differences in the number of viable tendon cells
The relative LDH released from 0, 1, 5, and 10 lM simvastatin-treated cells was 100.0 6 0%, 99.7 6 3.1%, 103.3 6 3.8%, and 109.3 6 6.2%, respectively (Fig. 2B); the differences between groups were not statistically significant. These results indicated that the simvastatin-induced decrease in the number of viable tendon cells was not due to cytotoxicity under these assay conditions

Summary

Introduction

Statins are a class of lipid-lowering drugs clinically used to inhibit 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA), a key enzyme in the mevalonate metabolic pathway for endogenous cholesterol production. Marie et al retrospectively identified cases of statin-associated tendinopathy, and the results suggested that tendinopathies more often occurred within the first year after statin initiation and most were related to atorvastatin and simvastatin use (37% and 31%, respectively). We reported that inhibition of cell proliferation was responsible for ciprofloxacin-associated tendinopathy (Tsai et al, 2008). The exact mechanism varies among cell types (Denoyelle et al, 2001; Ghosh et al, 1999) and the effect of simvastatin on tendon cell proliferation has not yet been investigated. The present study was designed to investigate whether simvastatin, a commonly prescribed statin, exerts any adverse effect on tendon cell proliferation, to explore the underlying molecular mechanisms, and to determine whether mevalonate, GGPP, and FPP can reverse the inhibition of tendon cell proliferation

Methods

Results

Conclusion