VASCULAR SURGERY II perfusion fixed, the left and right carotids were harvested, and analyzed by computer-assisted morphometric analysis. Student t test was used to compare the vessel perimeters of each group, with statistical significance determined at p⬍0.05.
Nitric oxide inhibits the 26S proteasome in six cell types important in the vasculature Muneera R Kapadia MD, Oliver Aalami MD, Qun Jiang MD, Adam Elhofy MD, Melina Kibbe MD Northwestern University, Chicago, IL INTRODUCTION: We previously showed that nitric oxide (NO) inhibits the 26S proteasome in vascular smooth muscle cells (VSMC), and this may be one mechanism by which NO inhibits neointimal hyperplasia. The aim of this study was: 1) to evaluate 26S proteasome activity in six cell types important in the vasculature; and 2) to determine the effect of NO on the 26S proteasome in these cells. METHODS: Whole cell lysate was collected from VSMC, fibroblasts (NIH3T3), endothelial cells (HUVEC), macrophages (RAW), dendritic cells (JAWSII), and T-cells. Reaction buffer, ATP, the NOdonor S-nitroso-N-acetylpenicillamine (SNAP) and fluorogenic proteasome-specific substrates (Suc-LLVY-AMC, Bz-VGR-AMC, or Z-LLE-AMC) were added. Proteosome activity was measured. The proteasome inhibitor, MG132, was used as negative control. RESULTS: At baseline, chymotrypsin-like activity was greatest in the HUVEC, trypsin-like activity was greatest in fibroblasts and dendritic cells, and caspase-like activity was greatest in HUVEC, (P⬍0.05). NO significantly inhibited all three 26S proteasome catalytic activities in all six cell types in a time- and concentrationdependent manner. However, chymotrypsin-like activity was inhibited greatest (50%; P⬍0.05). Mercuric chloride, a sulfhydryl reacting agent, inhibited 26S proteasome activity in all cells. Dithiothreitol, a reducing agent, reversed NO- and mercuric chloridemediated inhibition of the 26S proteasome, suggesting that NO inhibits the 26S proteasome through S-nitrosylation. CONCLUSIONS: Baseline 26S proteasome activity is differentially regulated in VSMC, fibroblasts, endothelial cells, macrophages, dendritic cells and T cells. Our findings suggest that NO inhibits 26S proteasome activity through S-nitrosylation of a critical cysteine residue. This represents a novel mechanism of regulation of the 26S proteasome.
The role of the MyD88 pathway in arterial remodeling Matthew M McRae BA, Amy Gallo MD, Ahmad Saad MD, Madison Cuffy MD, George Tellides MD, PhD Yale University, New Haven, CT INTRODUCTION: Chronic reduction in carotid blood flow, secondary to external carotid ligation, has been shown to cause inward vascular remodeling and subsequent reduction in vessel lumen diameter. Inflammation is not thought to play a role in flow-mediated arterial remodeling. Our goal is to test the role of innate immunity, specifically MyD88 signaling in vascular remodeling. METHODS: Sixteen C57BL/6 were used as the control group and 15 MyD88 ⫺/⫺ mice were used as the experimental group. The right external carotid artery was ligated in each animal. The left carotid served as an internal control. After 14 days the arteries were
© 2006 by the American College of Surgeons Published by Elsevier Inc.
RESULTS: Ligation of the external carotid arteries in the control group resulted in a significant reduction in lumen perimeter which did not occur in the carotids of MyD88 ⫺/⫺ mice (p⬍0.05). The mean perimeter of the control group right carotid arteries was 1.1 ⫹ 0.065 mm. The mean perimeter of the MyD88 right carotid arteries was 1.41 ⫹ 0.102 mm. CONCLUSIONS: This preliminary data supports a role of innate immunity, through the MyD88 adaptor molecule in vascular remodeling following chronic blood flow changes.
Upregulated expression of mineralocorticoid receptor (MR) in vein grafts: A potential role for aldosterone signaling in vein graft disease Richard Bafford MD, Xin Xin SUI MD, Min Park BS, Jose Romero PhD, Gail Adler MD, PhD, Gordon Williams MD, Raouf Khalil MD, PhD, Michael Conte MD Brigham and Women’s Hospital, Boston, MA INTRODUCTION: Recent evidence suggests that the mineralocorticoid aldosterone (ALDO), via activation of the MR, is an important amplifier of the local renin-angiotensin system (RAS) in cardiovascular tissue. We hypothesize that MR activation contributes to the injury response in vein bypass grafts. METHODS: Greater saphenous vein (GSV;N⫽6) and excised lesions of human peripheral vein grafts (HVG;N⫽15,84-1330 days post-bypass) were obtained under an IRB approved protocol. NZW rabbits (N⫽15) underwent carotid interposition vein grafts (RVG) and were harvested 1-51 days later. Immunostaining was performed using a mAb to MR with IgG controls. RT-PCR was used to examined MR, 11-Beta-hydroxysteroid-dehydrogenase-2 (11B-HSD2), and angiotensin receptor type-1 (AT1R) gene expression in vein samples, as well as in primary cultures of human GSV smooth muscle cells (SMC). SMC were treated with ALDO (0.1uM) ⫹/⫺ an MRantagonist (MRA). RESULTS: MR immunostaining was observed in all HVG lesions and in RVG, with minimal staining in GSV. RT-PCR revealed upregulated expression of MR and 11B-HSD2 in RVG beginning 1 week post-bypass and increasing through 51 days. HVG demonstrated expression of MR and 11B-HSD2, whereas GSV expressed minimal MR and no detectable 11B-HSD2. MR and 11B-HSD2 were expressed in cultured SMC, and their functional significance was revealed by an ALDO-induced upregulation of AT1R expression (3.3 fold/control) that was sensitive to an MRA. CONCLUSIONS: Upregulation of MR and 11B-HSD2 in arterialized veins suggests that the vein graft may constitute a novel target for ALDO-mediated tissue injury. ALDO/MR signaling may potentiate local RAS activation in the graft, and may have relevance to vein graft disease.