COL4A2 Locus on Chromosome 13q34 to Coronary Artery Disease (CAD)

S257

Abstracts

by Oil Red O staining and confirmed by quantifying the expression levels of gene involved in fatty acid biosynthesis. After 18 hours, both GLN and TM significantly increased lipid accumulation in wild type MEFs, however deficiency of GSK3␣ or GSK3␤ attenuated ER stress induced lipid accumulation. ER stress-induced lipid accumulation was then assessed in MEFs lacking key ER stress response genes PERK and IRE1␣. Relative to wild type cells, lipid accumulation in PERK⫺/⫺ MEFs was limited when treated with either GLN or TM. IRE1␣⫺/⫺ MEFs showed an increase in intracellular lipid levels similar to wild type controls. Pharmacological inhibition of GSK3␣/␤ attenuated lipid accumulation in wild type and IRE1␣⫺/⫺ MEFs, however did not affect lipid levels in PERK⫺/⫺ MEFs. The apparent defect in ER stress-induced lipid accumulation could be rescued by overexpressing a constitutively active form of GSK3␤ containing the S9A point mutation using an adenoviral vector. CONCLUSION: Genetic deletion or pharmacological inhibition of GSK3␣/␤ attenuates ER stress-induced lipid accumulation suggesting that both forms of this kinase play an important role in ER stress signaling. PERK, but not IRE1␣, is required for ER stress-induced lipid accumulation. The effect of PERKdeficiency could be rescued by overexpression of a constitutively active form of GSK3␤. Together these data suggest that ER stress signals through PERK and GSK3␣/␤ to promote the expression of enzymes involved in lipid biosynthesis and cellular lipid accumulation.

420 CONTROLLED NITRIC OXIDE DELIVERY BASED ON SNITROSOTHIOL CONJUGATED INTERPOLYMER COMPLEXES DECREASES RESTENOSIS IN RATS J Guo, H Zhao, P Lee, A Giacca Toronto, Ontario

Nitric oxide (NO) is known to play critical roles in vascular physiology and pathophysiology. In particular, NO has the ability to inhibit the development of neointimal hyperplasia by inhibiting platelet aggregation and adhesion, leukocyte accumulation, vascular smooth muscle cell migration and proliferation and by stimulating re-endothelialization. Not surprisingly, NO supplementation at the site of angioplasty has been investigated in numerous studies. However, NO-based therapies have been limited to the short duration of NO release, short half-life of NO, and instability of available NO donors. In this study, we investigated a NO releasing interpolymer complex, previously shown to accelerate wound closure, in the effect to decrease intimal hyperplasia. This NO-releasing system consists of S-nitrosothiols (RSNOs), derived from endogenous glutathione (GSH) conjugated onto poly(vinyl methyl ether-co-maleic anhydride (PVMMA), that is subsequently hydrogen bonded to a block copolymer of ethylene oxide and propylene oxide (Pluronic F-127) forming interpolymer complex. In vitro NO release profiles showed sustained NO release cu-

mulating to 1ìmol for up to 10 days or more. To study the effect of this NO system on intimal hyperplasia, Sprague Dawley rats initially weighing 400-425g were treated with either NO-releasing complex or vehicle control prepared as a 30% Pluronic gel applied around the injured carotid immediately after carotid balloon injury. NO-releasing complex significantly decreased neointimal hyperplasia measured 28 days after injury by 22% (n ⫽ 7-8 per group; P⫽0.01). Thus, controlled NO delivery is effective in a model of restenosis, suggesting that this NO delivery system has potential to be developed into a drug eluting stent. CDA

421 FUNCTIONAL RELATIONSHIP OF THE COL4A1/COL4A2 LOCUS ON CHROMOSOME 13Q34 TO CORONARY ARTERY DISEASE (CAD) A Turner, P Lau, S Soubeyrand, O Jarinova, R McPherson Ottawa, Ontario BACKGROUND: The COL4A1 and COL4A2 genes on chromosome 13 have been identified as new loci associated with CAD (⬎22,000 CAD cases & ⬎64,000 controls) that overall discovered 13 novel regions associated with CAD. The index SNP at the COL4A1/COL4A2 locus from the CARDIoGRAM study (rs4773144) has a minor allele frequency of 0.4 and is associated with an increased risk of CAD (allele specific odds ratio⫽1.21 in the Ottawa Heart Study (OHS)). The goal of the current project is to elucidate how variants in the COL4A1 and COL4A2 genes associated with CAD functionally and mechanistically contribute to the CAD phenotype. Type IV collagen triple helices constitute the major structural component of basement membranes, consisting primarily of 2 COL4A1 chains arranged with 1 COL4A2 chain. COL4A1 and COL4A2 also have important functional roles in angiogenesis, and mutations are associated with diverse vascular abnormalities. METHODS/RESULTS: In a search for functional genetic variants, we resequenced the bidirectional COL4A1/COL4A2 promoter in 500 CAD cases and 500 controls and identified four novel SNPs, in promoter/enhancer regions essential for COL4A1 and/or COL4A2 gene expression and in strong linkage disequilibrium with several OHS risk SNPs. Three of these novel SNPs are in the region necessary for COL4A2 transcription and in promoter luciferase assays with HT-1080 cells result in 1520% decreases in COL4A2 promoter activity (p⬍0.005). Furthermore, a follow-up study to CARDIoGRAM with more cases and controls has identified an intronic SNP in COL4A2 with an even higher CAD association than rs4773144. 2 kb of this intronic sequence containing the CAD-associated SNP was cloned into the pGL3-Promoter vector (Promega), in which insertion of functional enhancers leads to upregulation of luciferase expression in vitro. Luciferase assays in HT-1080 cells reveal this intronic sequence acts as an enhancer due to its insertion upregulating pGL3-Promoter activity over threefold

S258

relative to controls. Future work entails narrowing down what SNP(s) in this enhancer region has actual functional consequences (ie. disrupting transcription factor binding sites). We also plan on conducting chromosome conformation capture experiments to determine whether this enhancer acts locally on COL4A1/COL4A2 or acts long-range on other genomic targets. CONCLUSION: These findings are important because misregulation of COL4A1 and COL4A2 could have important consequences relevant to CAD, including effects on basement membrane integrity and angiogenesis. OGS

422 ACCELERATED ADIPOSE TISSUE DYSFUNCTION AND CARDIOVASCULAR RISK IN INDIVIDUALS OF SOUTH ASIAN ETHNICITY S Rashid, M Melone, M Tarnopolsky, K Schulze, H Gerstein, S Yusuf, S Anand Hamilton, Ontario

Individuals with similar degrees of obesity can have significantly different risks of developing metabolic diseases. We previously identified that South Asians (SA) have more cardio-metabolic risk factors compared with European caucasians (EC) matched for body mass index (BMI). In the present study, we examined whether ethnic-specific differences in adipose tissue morphology and metabolic function can account for differences in cardiovascular disease (CVD) risk between SA and EC. METHODS: Subcutaneous superficial abdominal adipose tissue biopsies were obtained from 108 males and females of SA or EC descent, with a mean age of 36 years overall, and assessed for adipose tissue: (1.) morphology; (2.) functional metabolic processes, determined through gene expression profiles; and (3.) endocrine functions, measured via serum adipokine levels. All values reported are adjusted for BMI, age and sex. RESULTS: Adipocyte diameter was significantly greater (259⫾4 vs. 238⫾5 units, P⬍0.01) among SA compared with EC. SA also expressed greater levels SREBP1, TIMP1 and CD68 mRNA than EC, indicating increased lipid accumulation, decreased extracellular matrix breakdown and increased macrophage accumulation and inflammation in SA, respectively. Finally, serum adiponectin levels, were markedly lower in SA compared with EC (by 37%, P⬍0.001), and was associated with hyperinsulinemia and a marked 4-fold greater hepatic fat accumulation (P⬍0.01). CONCLUSION: SA have evidence of adipose tissue dysfunction at a relatively early age, as demonstrated by adipocyte hypertrophy, increased lipid accumulation and inflammation and reduced adiponectin secretion compared with EC. Adipose tissue dysfunction is related to greater CVD risk factors in SA and may be the primary defect leading to accelerated CVD and associated high morbidity and mortality rates in SA. AstraZeneca BACKGROUND:

Canadian Journal of Cardiology Volume 28 2012

Canadian Cardiovascular Society (CCS) CCS183 Oral CAD INTERVENTIONAL I Monday, October 29, 2012 Featured Research 425 COMPLETE VERSUS CULPRIT-ONLY REVASCULARIZATION IN ST ELEVATION MYOCARDIAL INFARCTION WITH MULTIVESSEL DISEASE UNDERGOING PRIMARY PERCUTANEOUS CORONARY INTERVENTION: A SYSTEMATIC REVIEW AND META-ANALYSIS KR Bainey, SR Mehta, RC Welsh Edmonton, Alberta BACKGROUND: The management of clinically significant nonculprit coronary artery disease (CAD) in patients with STsegment elevation myocardial infarction (STEMI) and multivessel is uncertain. International guidelines recommend culprit only percutaneous coronary intervention (PCI) during the intial primary PCI, but whether a staged approach should routinely be performed is not known. Accordingly, we conducted a meta-analysis comparing the benefits and risks of routine culprit-only PCI versus multi-vessel PCI in STEMI. METHODS: Studies were identified through comprehensive search of MEDLINE, EMBASE, ISI the Web of Science, CENTRAL, hand searching of conferences, and cross references from original articles and reviews. Studies included patients with STEMI and multi-vessel CAD receiving primary PCI. Major outcomes of death and repeat PCI occurring from index hospitalization to end of follow-up were extracted. Data were combined using a fixed-effects model. RESULTS: Of 507 citations identified, 26 studies (46 249 patients; 7886 multi-vessel PCI and 38 438 culprit-only PCI) were included. There was significant heterogeneity between the randomized trials and registry studies for in-hospital survival (P⬍0.001, Figure). There was a trend toward increased inhospital death in patients receiving multi-vessel PCI compared with culprit-only PCI (OR 1.11; 95% CI 0.98-1.25) but this risk was observed only when multivessel PCI was performed during the index catheterization (OR 1.35; 95% CI 1.19-1.54; p⬍0.001). By contrast, when a staged procedure was performed to a non-culprit lesion, there was a reduction in mortality (OR 0.35; 95% CI 0.21-0.59; p⬍0.001) (p interaction ⬍0.001). A reduction in long-term mortality and repeat PCI was also observed with multi-vessel PCI compared with culprit-only PCI (OR 0.74; 95% CI 0.65-0.85; p⬍0.001; OR 0.65; 95% 0.46-0.90; p⫽0.01 respectively). CONCLUSION: In STEMI patients with multi-vessel CAD, staged PCI of non-culprit lesions reduced long-term mortality and need for repeat PCI compared with PCI to the culprit lesion only. By contrast, PCI of non-culprit lesions performed during the primary PCI procedure was associated with a trend towards higher in-hospital mortality. These data suggest that