- Email: [email protected]
W01-P-005 Anti-endothelial cells antibodies in atherosclerosis and acute coronary syndromes
WI
Workshops Apoptosis, plaque instability and acute coronary events
W01-P-004 ] T H E H M G - C O A REDUCTASE INHIBITOR ATORVASTATIN REDUCES FN14 EXPRESSION INDUCED BY P R O I N F L A M M A T O R Y CYTOKINES IN H U M A N VASCULAR S M O O T H M U S C L E C E L L S L.M. Blanco-Colio 1, B. Munoz-Garcia 1, J.L. Maxtin-Venturn 1, E. Martinez 2 , G. Hemandez 2, J. Egido 1. 1Vascular Research Lab,
Fundacion Jimenez Diaz, Autonoma University, Madrid, Spain; 2R &D Departmeng Medical Division, Pfizer, Spain Background: Fnl4, a cell-surface receptor for the TNF suporfamily, is implicated in cell proliferation, inflammation and angiogeuesis. HMG-CoA reductase inhibitors reduce cardiovascular mortality, though the mechanisms of action have not been completely elucidated. Methods: Fnl4 expression was analyzed in human aortic vascular smooth muscle cells (hASMCs) in culture (Western-blot and real-time PCR) and in atherosclerotic plaques of 20 patients with carotid atheroscleroils randomized to atorvastatin 80 mg/day (ATV) (n=ll) or usual care (n=9) during 4-6 weeks before they underwent carotid endarterectomy (immunohistocbemistry). Results: Proinflammatory cytokiues (100 U/mL IL-I[3; 500 U/mL IN'F-y) upregulate Fnl4 expression in hASMCs, which was prevented by atorvastatin treatment (10 .7-10 .5 mol/L). Mevalonate, the mntabolite synthesized by HMG-CoA reductase, reversed the effect induced by atorvastatin, indicating that atorvastatiu through inhibition of cholesterol pathway can regulate Fnl4 expression. Geranylgerauyl pyrophosphate, but not farnesyl pyrophosphate, two isoprenoids synthesized in the cholesterol pathway, prevented the downregulation induced by atorvastatin. The attachment of an isoprenoid residue to the small G protons is necessary for thor full functionality. Inhibition of gerauylgeranyl trausferase showed similar reduction of Fn 14 expression, demonstrating that small G proteins modified by GGPP are necessary for Fn14 expression. Toxin B (Rac and Rho inhibitor), C3 exoenzyme (Rho protons inhibitor) and Y-27632 (Rho kinase inhibitor) also decrease Fn14 expression, implicating Rho/Rho kinase pathway in the regulation of Fn 14 expression. Finally, atorvastatin treatment reduced Fnl4 expression in human carotid atherosclerotic plaques. Conclusion: Atorvastatin reduces Fnl4 expression in cultured hASMCs and atherosclerotic plaques probably due to the inhibition of small G proteins prenylation. These results provide novel information of the beneficial anti-inflammatory propierties of atorvastatin.
W01-P-005 ] A N T I - E N D O T H E L I A L CELLS ANTIBODIES IN ATHEROSCLEROSIS AND ACUTE CORONARY SYNDROMES I. B urazor 1, A. Vojdani2.1 Clinic for Cardiovascular diseases, Clinical center, Nis, Serbia and Montenegro; 2Immunosciences Lab. Inc, Beverly Hills, Los Angeles, USA Anti-endothelial cells antibodies (AECA) are non specific antibodies that are directed against endothelial antigens. Several effects of AECA are emerging such as endothelial cell lyses, activation of the cells, production of cytokines or EC programmed cell death which can been shown to be participant in vascular injury. Is the appearance of these antibodies result of specific autoimmune response driven by infectious agent? Aim of the study was to determine if AECA can be detected in patients with acute coronary syndromes and do they play a role in the process of plaque instability. Patients and methods: Study included 100 patients with acute coronary syndromes on admission to Coronary Care Unite of Clinic for cardiovascular diseases, 50 controls from Serbia and 50 controls from California. Blood samples were taken, serum was frozen at -40°C and kept in a freezer. The packaging was sent on dry ice to Immunosciences Lab Inc. Beverly Hills, USA where the analyses were done by using ELISA method Traditional risk factors for cardiovascular diseases were noted. Results: Out of 45% patients with acute coronary event had positive AECA versus 22% of healthy controls from Serbia and 4% of healthy controls from California. The mean values of AECA titer was 0.684+0.211 o.d. for patients (max value 1.545 o.d.). Subjects from Serbia had mean value 0.598-t-0.193 o.d. and for subjects from California 0.509+0.102 o.d. (max. 0.811 o.d.) The difference between patients and controls from Serbia was p=0.012. Statistically significant difference between AECA titers in patients and controls from California was p<0.001, and there was a difference between two control groups. Healthy subjects from Serbia had higher titer on AECE (p<0.001).
Conclusion: In patients with acute coronary syndromes activity of AECA is high and can represent the autoimmune process which highlights the development and progression of atherosclerosis. Healthy subjects from Serbia can be at high risk to develop acute event compared to healthy subjects from California. 1
I W01-P-006 / HDL C H O L E S T E R O L , S M O K I N G AND I L l 3 R130Q J P O L Y M O R P H I S M A R E ASSOCIATED W I T H M Y O C A R D I A L INFARCTION IN G R E E K C Y P R I O T MALES M.A. Cariolou 1, M. Hadjivassiliou 1, N. Demetriou 1, I. Marcou 4, A. Karagrigoriou 2, G. Miltiadous 3 , M. Elisaf 3 . 1Molecular~Forensic
Genetics, Cyprus Institute of Neurology & Genetics, Nicosia, Cyprus; 2Dept Mathemalics & Statistics, University of Nicosia, Nicosia, Cyprus; 31oannina Medical Schoo~ Ioannina, Greece; 4Nicosia General Hospita~ Nicosia, Cyprus Objective: An epidemiological study was carried out in Greek Cypriot males to identify risk factors that predispose this population to myocardial infarction. Genetic, biochemical and behavioral risk factors were investigated in a case control study. Methods: The ILl3 R 130Q polymorphism was genotypod using a novel TaqMan assay, other genotyping used established protocols, and statistical analyses were performed with statistical package SPSS version 12. Results: Contrary to other studies, mean LDL cholesterol did not differ between cases and controls. HDL-cholesterol on the other hand, although within normal range in cases and controls, was found to be significantly higher in the control population, (40.22-t-9 versus 45.70+11.99 mgdl); P=0.001). Second, in agreement with many other studies, smoking was more prevalent in cases compared to controls, (83% versus 58%; P=0.000). A smoker is 2.901 times more likely to experience an MI than a non-smoker in this male population. The frequency of the ILl3 R130Q heterozygotes and homozygotes (RQ & QQ respectively) was higher in cases compared to controls, although not statistically significant (40.2% versus 28.8% for RQ & 7.6% versus 2.5% for QQ respectively; P=0.058). In contrast, the difference in allele frequency was statistically significant, (28% in cases versus 17% in controls in the frequency of mutant Q allele, P=0.0240). Further, heterozygotes were found to be 0.675 times more likely to develop an MI than individuals with a wild type genotype, and homozygotes for the mutation were 1.534 times more likely than individuals with a wild type genotype. Conclusion: This study has indicated that smoking, a relatively low HDl.,-cholesterol level and possibly ILl3, an inflammatory cytokine play a role in MI in this male population.
WO1-P-O07]LERCANIDIPINE REDUCES F R E E C H O L E S T E R O L - I N D U C E D CYTOTOXICITY E. Favari, M.P. Adorni, E Zimetti, I. Zanotti, F. Bemini. Dept.
Pharmacological and Biological Sciences and AppL Chem., University of Parrtm, Parrtm, Italy Objective: excess intracellular free cholesterol (F,C) is cytotoxic to many cells types and it was hypothesized that accumulation of FC is a mediator of macrophage death in vivo and contributes to the atherosclerotic lesion progression. Excess cellular F,C generated through the hydrolysis of cytoplasmatic cholesteryl ester (EC) is esterified by ACAT enzyme or is transported to the plasma membrane where it induces a variety of responses as increased phospholipid synthesis, cell death and FC crystals formation. Lercanidipine is a new calcium antagonist with high lipophificity and a chiral center. Lercanidipine reduce both cholesterol esterification and free cholesterol accumulation in macrophages. These effects do not involve a Ca 2+ antagonist action, but are related to the lipophilic characteristic of the molecule. These observations indicate that the drug may interfere with cellular cholesterol trafficking. The aim of this study was to examine lercanidipine activity in preventing FC-induced cytotoxicity.Methods: J774 were cholesterol-enriched using acetylated low-density lipoprotein and FC/phospholipid dispersions. After 48h of incubation monolayers were labeled with lp.Ci [3H]-adeniue. To evaluate lercanidipine effect on FC-induced cytotoxicity J774 foam cells were induced to accumulate excess intracellular FC by incubation with an ACAT inhibitor. Cytotoxicity was measured by cellular release of [3H]-adeuine. Results: the ACAT inhibitor, but not lercanidipine induced a 3-fold increase of adenine release. Moreover, J774 foam cells incubated with the ACAT inhibitor in the
75th EAS Congress, 23-26 April 2005, Prague, Czech Republic
0 I 0 Go
Workshops Apoptosis, plaque instability and acute coronary events
W01-P-004 ] T H E H M G - C O A REDUCTASE INHIBITOR ATORVASTATIN REDUCES FN14 EXPRESSION INDUCED BY P R O I N F L A M M A T O R Y CYTOKINES IN H U M A N VASCULAR S M O O T H M U S C L E C E L L S L.M. Blanco-Colio 1, B. Munoz-Garcia 1, J.L. Maxtin-Venturn 1, E. Martinez 2 , G. Hemandez 2, J. Egido 1. 1Vascular Research Lab,
Fundacion Jimenez Diaz, Autonoma University, Madrid, Spain; 2R &D Departmeng Medical Division, Pfizer, Spain Background: Fnl4, a cell-surface receptor for the TNF suporfamily, is implicated in cell proliferation, inflammation and angiogeuesis. HMG-CoA reductase inhibitors reduce cardiovascular mortality, though the mechanisms of action have not been completely elucidated. Methods: Fnl4 expression was analyzed in human aortic vascular smooth muscle cells (hASMCs) in culture (Western-blot and real-time PCR) and in atherosclerotic plaques of 20 patients with carotid atheroscleroils randomized to atorvastatin 80 mg/day (ATV) (n=ll) or usual care (n=9) during 4-6 weeks before they underwent carotid endarterectomy (immunohistocbemistry). Results: Proinflammatory cytokiues (100 U/mL IL-I[3; 500 U/mL IN'F-y) upregulate Fnl4 expression in hASMCs, which was prevented by atorvastatin treatment (10 .7-10 .5 mol/L). Mevalonate, the mntabolite synthesized by HMG-CoA reductase, reversed the effect induced by atorvastatin, indicating that atorvastatiu through inhibition of cholesterol pathway can regulate Fnl4 expression. Geranylgerauyl pyrophosphate, but not farnesyl pyrophosphate, two isoprenoids synthesized in the cholesterol pathway, prevented the downregulation induced by atorvastatin. The attachment of an isoprenoid residue to the small G protons is necessary for thor full functionality. Inhibition of gerauylgeranyl trausferase showed similar reduction of Fn 14 expression, demonstrating that small G proteins modified by GGPP are necessary for Fn14 expression. Toxin B (Rac and Rho inhibitor), C3 exoenzyme (Rho protons inhibitor) and Y-27632 (Rho kinase inhibitor) also decrease Fn14 expression, implicating Rho/Rho kinase pathway in the regulation of Fn 14 expression. Finally, atorvastatin treatment reduced Fnl4 expression in human carotid atherosclerotic plaques. Conclusion: Atorvastatin reduces Fnl4 expression in cultured hASMCs and atherosclerotic plaques probably due to the inhibition of small G proteins prenylation. These results provide novel information of the beneficial anti-inflammatory propierties of atorvastatin.
W01-P-005 ] A N T I - E N D O T H E L I A L CELLS ANTIBODIES IN ATHEROSCLEROSIS AND ACUTE CORONARY SYNDROMES I. B urazor 1, A. Vojdani2.1 Clinic for Cardiovascular diseases, Clinical center, Nis, Serbia and Montenegro; 2Immunosciences Lab. Inc, Beverly Hills, Los Angeles, USA Anti-endothelial cells antibodies (AECA) are non specific antibodies that are directed against endothelial antigens. Several effects of AECA are emerging such as endothelial cell lyses, activation of the cells, production of cytokines or EC programmed cell death which can been shown to be participant in vascular injury. Is the appearance of these antibodies result of specific autoimmune response driven by infectious agent? Aim of the study was to determine if AECA can be detected in patients with acute coronary syndromes and do they play a role in the process of plaque instability. Patients and methods: Study included 100 patients with acute coronary syndromes on admission to Coronary Care Unite of Clinic for cardiovascular diseases, 50 controls from Serbia and 50 controls from California. Blood samples were taken, serum was frozen at -40°C and kept in a freezer. The packaging was sent on dry ice to Immunosciences Lab Inc. Beverly Hills, USA where the analyses were done by using ELISA method Traditional risk factors for cardiovascular diseases were noted. Results: Out of 45% patients with acute coronary event had positive AECA versus 22% of healthy controls from Serbia and 4% of healthy controls from California. The mean values of AECA titer was 0.684+0.211 o.d. for patients (max value 1.545 o.d.). Subjects from Serbia had mean value 0.598-t-0.193 o.d. and for subjects from California 0.509+0.102 o.d. (max. 0.811 o.d.) The difference between patients and controls from Serbia was p=0.012. Statistically significant difference between AECA titers in patients and controls from California was p<0.001, and there was a difference between two control groups. Healthy subjects from Serbia had higher titer on AECE (p<0.001).
Conclusion: In patients with acute coronary syndromes activity of AECA is high and can represent the autoimmune process which highlights the development and progression of atherosclerosis. Healthy subjects from Serbia can be at high risk to develop acute event compared to healthy subjects from California. 1
I W01-P-006 / HDL C H O L E S T E R O L , S M O K I N G AND I L l 3 R130Q J P O L Y M O R P H I S M A R E ASSOCIATED W I T H M Y O C A R D I A L INFARCTION IN G R E E K C Y P R I O T MALES M.A. Cariolou 1, M. Hadjivassiliou 1, N. Demetriou 1, I. Marcou 4, A. Karagrigoriou 2, G. Miltiadous 3 , M. Elisaf 3 . 1Molecular~Forensic
Genetics, Cyprus Institute of Neurology & Genetics, Nicosia, Cyprus; 2Dept Mathemalics & Statistics, University of Nicosia, Nicosia, Cyprus; 31oannina Medical Schoo~ Ioannina, Greece; 4Nicosia General Hospita~ Nicosia, Cyprus Objective: An epidemiological study was carried out in Greek Cypriot males to identify risk factors that predispose this population to myocardial infarction. Genetic, biochemical and behavioral risk factors were investigated in a case control study. Methods: The ILl3 R 130Q polymorphism was genotypod using a novel TaqMan assay, other genotyping used established protocols, and statistical analyses were performed with statistical package SPSS version 12. Results: Contrary to other studies, mean LDL cholesterol did not differ between cases and controls. HDL-cholesterol on the other hand, although within normal range in cases and controls, was found to be significantly higher in the control population, (40.22-t-9 versus 45.70+11.99 mgdl); P=0.001). Second, in agreement with many other studies, smoking was more prevalent in cases compared to controls, (83% versus 58%; P=0.000). A smoker is 2.901 times more likely to experience an MI than a non-smoker in this male population. The frequency of the ILl3 R130Q heterozygotes and homozygotes (RQ & QQ respectively) was higher in cases compared to controls, although not statistically significant (40.2% versus 28.8% for RQ & 7.6% versus 2.5% for QQ respectively; P=0.058). In contrast, the difference in allele frequency was statistically significant, (28% in cases versus 17% in controls in the frequency of mutant Q allele, P=0.0240). Further, heterozygotes were found to be 0.675 times more likely to develop an MI than individuals with a wild type genotype, and homozygotes for the mutation were 1.534 times more likely than individuals with a wild type genotype. Conclusion: This study has indicated that smoking, a relatively low HDl.,-cholesterol level and possibly ILl3, an inflammatory cytokine play a role in MI in this male population.
WO1-P-O07]LERCANIDIPINE REDUCES F R E E C H O L E S T E R O L - I N D U C E D CYTOTOXICITY E. Favari, M.P. Adorni, E Zimetti, I. Zanotti, F. Bemini. Dept.
Pharmacological and Biological Sciences and AppL Chem., University of Parrtm, Parrtm, Italy Objective: excess intracellular free cholesterol (F,C) is cytotoxic to many cells types and it was hypothesized that accumulation of FC is a mediator of macrophage death in vivo and contributes to the atherosclerotic lesion progression. Excess cellular F,C generated through the hydrolysis of cytoplasmatic cholesteryl ester (EC) is esterified by ACAT enzyme or is transported to the plasma membrane where it induces a variety of responses as increased phospholipid synthesis, cell death and FC crystals formation. Lercanidipine is a new calcium antagonist with high lipophificity and a chiral center. Lercanidipine reduce both cholesterol esterification and free cholesterol accumulation in macrophages. These effects do not involve a Ca 2+ antagonist action, but are related to the lipophilic characteristic of the molecule. These observations indicate that the drug may interfere with cellular cholesterol trafficking. The aim of this study was to examine lercanidipine activity in preventing FC-induced cytotoxicity.Methods: J774 were cholesterol-enriched using acetylated low-density lipoprotein and FC/phospholipid dispersions. After 48h of incubation monolayers were labeled with lp.Ci [3H]-adeniue. To evaluate lercanidipine effect on FC-induced cytotoxicity J774 foam cells were induced to accumulate excess intracellular FC by incubation with an ACAT inhibitor. Cytotoxicity was measured by cellular release of [3H]-adeuine. Results: the ACAT inhibitor, but not lercanidipine induced a 3-fold increase of adenine release. Moreover, J774 foam cells incubated with the ACAT inhibitor in the
75th EAS Congress, 23-26 April 2005, Prague, Czech Republic
0 I 0 Go