Technical Reference #1770

1770.

Simvastatin Inhibits Angiotensin II-Induced Abdominal Aortic Aneurysm Formation in Apolipoprotein E-Knockout Mice: Possible Role of ERK Yali Zhang; Jack C. Naggar; C. Michael Welzig; Debbie Beasley; Karen S. Moulton; Ho-Jin Park; Jonas B. Galper, Tufts University School of Medicine, Arteriosclerosis; Thrombosis; and Vascular Biology, 29(1770), (2009)
Link To Paper

Abstract:
Objective—Abdominal aortic aneurysm (AAA) is a life-threatening disease affecting almost 10% of the population over age 65. Generation of AAAs by infusion of angiotensin (Ang) II in apolipoprotein E–knockout (ApoE / ) mice is an animal model which supports an imbalance of the renin–angiotensin system in the pathogenesis of AAA. The effect of statins on AngII-mediated AAA formation and the associated neovascularization is not known. Here we determined the effect of simvastatin and the ERK inhibitor CI1040 on AngII-stimulated AAA formation. Methods and Results—ApoE / mice infused for 28 days with AngII using osmotic minipumps were treated with placebo 10 mg/kg/d simvastatin or 100 mg/kg/d CI1040. 95% of AngII-treated mice developed AAA with neovascularization of the lesion increased ERK phosphorylation MCP-1 secretion and MMP activity. These effects were markedly reversed by simvastatin and in part by CI1040. Furthermore simvastatin and the ERK inhibitor U0126 reversed AngII-stimulated angiogenesis and MMP secretion by human umbilical vein endothelial cells. Conclusions—These data support the conclusion that simvastatin interferes with AAA formation induced by AngII in ApoE / mice at least in part via ERK inhibition.

Keywords:
angiotensin II • abdominal aortic aneurysm • statin • ERK inhibition • angiogenesis

Materials & Methods:
In Vitro Angiogenesis Model HUVECs were isolated by the method of Gimbrone28 and cultured as described previously.24 HUVECs grown to near confluence were pretreated either overnight with 1 mol/L simvastatin or for 1 hour with 10 mol/L U0126 an ERK inhibitor followed by a 6-hour incubation with 200 nmol/L AngII. Conditioned medium was collected for analysis of MMPs. For the study of angiogenesis HUVECs were cultured on Matrigel (Becton Dickinson) and the formation of capillary-like structures honeycombs quantitated.29 Specifically HUVECs were cultured to 80% confluence in 6-well plates and pretreated as described. After pretreatment HUVECs were trypsinized plated onto Matrigel-coated 24-well plates (MatTek) at a density of 5 104 cells per well and incubated overnight with AngII and either simvastatin or U0126. After fixing and washing honeycomb formation was determined by photomicrography. Images were analyzed using Scion Image software and the total length of tube-like structures quantified. The total percent difference in tube formation relative to control was determined for each group. The mean values from 24 fields in each group from 3 independent experiments were calculated.

Microscopic Technique
photomicrography

Cell Type(s)
HUVECs