Acta Pharmacologica Sinica (2009) 30: 1392–1398; doi: 10.1038/aps.2009.135; published online 21 Sep 2009

Aim: To examine the effect of atorvastatin on homocysteine (Hcy)-induced reactive oxygen species (ROS) production and apoptosis in human umbilical vein endothelial cells (HUVECs).

Methods: HUVECs were cultured with Hcy (0.1−5 mmol/L) in the presence or absence of atorvastatin (1−100 μmol//L) or various stress signaling inhibitors, including the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor diphenylene iodonium (DPI, 10 μmol/L), the p38 mitogen-activated protein kinase (p38 MAPK) inhibitor SB203580 (10 μmol/L) and antioxidants N-acetyl cysteine (NAC, 1 mmol/L).  Cell apoptosis was evaluated by Annexin V/propidium iodide staining and flow cytometry.  ROS were detected by 2′,7′-dichlorodihydrofluorescein diacetate (H₂DCFH-DA).  NADPH oxidases were evaluated with lucigenin-enhanced chemiluminescence.  Hcy-induced expression of p38MAPK protein was measured by Western blotting analysis. 

Results: Atorvastatin inhibited endothelial cell apoptosis induced by 1 mmol/L Hcy in a dose-dependent manner and the maximal inhibitory effect was reached at 100 μmol/L.  Atorvastatin (10 μmol/L) significantly suppressed Hcy (1 mmol/L for 30 min) induced ROS accumulation (3.17±0.33 vs 4.34±0.31, P<0.05).  Atorvastatin (10 μmol/L) also antagonized Hcy (1 mmol/L for 30 min) induced activation of NADPH oxidase (2.57±0.49 vs 3.33±0.6, P<0.05).  Furthermore, atorvastatin inhibited Hcy-induced phosphorylation of p38 MAPK (1.7±0.1 vs 2.22±0.25, P<0.05), similar effects occurred with DPI, NAC and SB203580. 

Conclusion: Atorvastatin may inhibit Hcy-induced ROS accumulation and endothelium cell apoptosis through an NADPH oxidase and/or p38MAPK-dependent mechanisms, all of which may contribute to atorvastatin-induced beneficial effect on endothelial function.

Keywords: atorvastatin; homocysteine; apoptosis; reactive oxygen species; NADPH oxidase; p38MAPK; human umbilical vein endothelial cells