- Email: [email protected]
W07.175 Oral tolerance as a new therapeutic modality for atherosclerosis
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Rosenblat
Workshops W7 Immune modulation in the treatment of atherosclerosis
PARAOXONASE 1 (PON1) ENHANCES HDLMEDIATED MACROPHAGE CHOLESTEROL EFFLUX VIA THE ABCA1 TRANSPORTER: A POSSIBLE ROLE F O R LYSOPHOSPHATIDYL CHOLINE
W7 I M M U N E M O D U L A T I O N IN THE T R E A T M E N T OF ATHEROSCLEROSIS
IW07.1741
ATHEROPROTECTIVE IMMUNIZATION WITH MDA-MODIFIED APO B-100 PEPTIDE SEQUENCES IS ASSOCIATED WITH ACTIVATION OF TH2 SPECIFIC ANTIBODY EXPRESSION
M. Rosenblat, D. Shih, J. Vaya, M. Avfl'am. Rambam Medical Center,
Technion Faculty of Medicine, The Rappaport Family Institute for Research in the Medical Sciences, Haifa, Migal-Galilee Technological Center, Kiryat Shmona, Israel; UCLA, LOs Angeles, USA The present study questioned the possible role of HDL-associated paraoxonasel (PON1) in HDL-mediated macrophage cholesterol efflux. Cholesterol effiux fi'om mouse peritoneal macrophages or fi'om J774 A.1 cell line was found to be significantly increased by 70% on using HDL fi'om human PONl-transgenic mice (HDL-PON1Tg) compared to HDL fi'om PON1- knockout mice (HDL-PONI°). Similar effect was observed upon HDL enrichment with purified PON1. The PON1 flee sulfhydryl group (cysteine 284), but not the PON1 N-terminal leader sequence, was requfl'ed for PON1 stimulation of HDL-mediated cholesterol efflux. This latter effect of PON1 could be related to increased HDL binding to the cells, as the binding of HDL-PON1Tg or that of HDL-PON1 ° enriched with PON1 to macrophages was increased by up to 50%, compared to the binding rate of HDL-PON1 °. Upon incubation of macrophages with HDL-PON1Tg, but not with HDL-PON1 °, cellular lysophosphatidylcholine (LPC) content was increased by 7.7 fold. LPC enrichment of macrophages resulted in up to 60% increased HDL binding to the cells, and up to 41% increased HDL-mediated cholesterol efflux. On using a cAMP analog to increase the ABCA1 receptor expression, or the rabbit anti-mouse SR-BI specific antibody to block the SR-BI receptor, PON1 stimulation of HDL binding and HDL-mediated cholesterol efflux fi'om macrophages was found to involve the ABCA1 receptor. In conclusion, PON1 stimulation of HDL-mediated cholesterol efflux fi'om macrophages can contribute to the attenuation of atherosclerosis development.
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INFLAMMATION AND ENDOTHELIAL ACTIVATION ASSOCIATE WITH FAMILIAL LOW HDL
A. Soro-Paavonen, J. Westerbacka, C. Ehnholm, M. Taskinen. University of
Helsinki, and National Public Health Institute, Helsinki, Finland Objective Endothelial activation and low-grade inflammation of the arterial wall are characteristics of atherosclerosis. Inhibition of cytokine-induced expression of cellular adhesion molecules (CAMs) is one of the atheroprotective mechanisms of high density lipoprotein (HDL). We studied whether the levels of soluble CAMs are increased in subjects with familial low HDL, and whether endothelial activation associates with the metabolic syndrome in these subjects. Methods: Serum levels of the soluble forms of intercellular adhesion molecule-1 (slCAM-1), vascular cell adhesion molecule-1 (sVCAM-1), sEselectin and C-reactive protein (CRP) were measured in 91 subjects with familial low HDL fi'om Finnish low HDL families, and in 112 age-sex matched controls. The number of the features of the metabolic syndrome was determined. Results: Low HDL subjects had significantly higher sVCAM-1, slCAM1, sE-selectin and CRP than the controls (for sVCAM-1; 560-4-147ng/ml vs. 496-4-95 ng/ml, P<0.001, for slCAM-1; 247-4-60ng/ml vs. 215-4-47ng/ml, P<0.001, for sE-selectin; 52-4-20ng/ml vs 44-4-16ng/ml, P=0.020, for CRP; 1.73-4-2.05mg/1 vs. 0.85-4-1.10mg/1, P<0.001). HDL cholesterol cora'elated significantly with sVCAM-1 and with slCAM-1 when adjusted for age, gender, body mass index (BMI) and smoking. The levels of sCAMs and CRP increased according to the number of the features of the metabolic syndrome. Conclusions: The familial low HDL trait associates with low-grade inflammation and endothelial activation. Elevated concentrations of sCAMs and CRP predispose these subjects to increased cardiovascular risk.
G. Nordin Fredrikson, L. Andersson, I. S derberg, P. Dimayuga, K. Chyu, P. Shah, J. Nilsson. Exp Cardiovascular Research, Maim , Sweden;
Atherosclerosis Research Center, Los Angeles, USA Background: Oxidized LDL is present in atherosclerotic plaques where it causes cell injury and inflammation. However, immunization with oxidized LDL, as well as apolipoprotein B-100 (apo B-100) fi'agments, induces atheroprotective immune responses in apo E deficient mice. Methods and Results: Apo E deficient mice were immunized with either of three MDA-modified human apo B-100 fi'agments (P45; amino acids 688-707, P74; amino acids 1123-1142 or P240; amino acids 3613-3632) at 6, 9 and 11 weeks of age and compared to controls given carrier alone. The sequence of P240 is not homologous with the cora'esponding mouse sequence and served as control for peptide specificity. Immunization with fi'agments P45 and P74 reduced atherosclerosis in the aorta of 25-week old mice by 48% (p=0.02) and 31% (p=0.06) and macrophage content in atherosclerotic plaques by 33% (p=0.02) and 39% (p=0.02), respectively. The levels of Th2-specific IgG1 against each peptide increased more than fifty-fold in response to immunization, whereas the levels of peptide-specific IgM and Thl-associated IgG2a were only marginally affected. However, there was an increase in the plaque expression of both Thl and Th2 specific cytokines as assessed by real time PCR. Immunization with the non-homologous peptide fi'agment P240 induced a ten-fold increase of specific IgG1 antibodies, but did not influence atherosclerosis or plaque content. Condusions: These findings suggest that immunization with MDA apo B-100 fi'agments induce a shift fi'om Thl to a Th2 specific oxidized LDL antibody expression, but without a concomitant downregulation of plaque IFN-y expression. ORAL TOLERANCE AS A NEW THERAPEUTIC IW07.1751 MODALITY F O R ATHEROSCLEROSIS I i
N. Yacov, J. George, G. Halperin, E. Ishai Kovalevski, E. Breitbart, L. Bangio, A. Shish, H. Levkovitz, Y. Liberrnan, D. Harats. Vascular
Biogenics LTD, Or Yehuda and Institute of Lipid and Atherosclerosis Research, Sheba Medical Center, Tel Hashomer, Israel Inflammation is a major component in atherogenesis and among is the autoimmunity to oxidized LDL that is involved in the progression of atherosclerosis. Oral administration of autoantigens in autoimmune diseases decreases the inflammatory process and the disease outcome. We explored the effect of oral administration of a mimetic epitope of oxidized LDL on aortic sinus lesion formation and on cytokine expression in the aorta in old ApoE KO mice with advanced lesion, on chow diet. Animals were stratified by age, weight and lipid profile to the different groups. Oral administrations of the mimetic epitope of oxidized LDL, was conducted for 10 days (total of 5 doses), at three sets given at 4 weeks intervals. After 12 weeks of treatment the control fed mice exhibit a 50% increase in aortic sinus lesion, while the mimetic epitope of oxidized LDL treatment group completely blocked any fmther progression in aortic sinus lesion as compared to base line group which was sacrificed at study start. We also found an increased expression level of IL-10 and decreased expression level of IFN-y in aortas of mice treated with the mimetic epitope of oxidized LDL as compared to control mice. Our results demonstrates that oral administration of the mimetic epitope of oxidized LDL totally prevents the progression of atherosclerosis which is associated with the induction of immune-modulation fi'om inflammatory response to an anti-inflammatory response and provides a novel approach for the treatment of atherosclerosis.
74th EAS Congress, 17-20 April 2004, Seville, Spain
•
Rosenblat
Workshops W7 Immune modulation in the treatment of atherosclerosis
PARAOXONASE 1 (PON1) ENHANCES HDLMEDIATED MACROPHAGE CHOLESTEROL EFFLUX VIA THE ABCA1 TRANSPORTER: A POSSIBLE ROLE F O R LYSOPHOSPHATIDYL CHOLINE
W7 I M M U N E M O D U L A T I O N IN THE T R E A T M E N T OF ATHEROSCLEROSIS
IW07.1741
ATHEROPROTECTIVE IMMUNIZATION WITH MDA-MODIFIED APO B-100 PEPTIDE SEQUENCES IS ASSOCIATED WITH ACTIVATION OF TH2 SPECIFIC ANTIBODY EXPRESSION
M. Rosenblat, D. Shih, J. Vaya, M. Avfl'am. Rambam Medical Center,
Technion Faculty of Medicine, The Rappaport Family Institute for Research in the Medical Sciences, Haifa, Migal-Galilee Technological Center, Kiryat Shmona, Israel; UCLA, LOs Angeles, USA The present study questioned the possible role of HDL-associated paraoxonasel (PON1) in HDL-mediated macrophage cholesterol efflux. Cholesterol effiux fi'om mouse peritoneal macrophages or fi'om J774 A.1 cell line was found to be significantly increased by 70% on using HDL fi'om human PONl-transgenic mice (HDL-PON1Tg) compared to HDL fi'om PON1- knockout mice (HDL-PONI°). Similar effect was observed upon HDL enrichment with purified PON1. The PON1 flee sulfhydryl group (cysteine 284), but not the PON1 N-terminal leader sequence, was requfl'ed for PON1 stimulation of HDL-mediated cholesterol efflux. This latter effect of PON1 could be related to increased HDL binding to the cells, as the binding of HDL-PON1Tg or that of HDL-PON1 ° enriched with PON1 to macrophages was increased by up to 50%, compared to the binding rate of HDL-PON1 °. Upon incubation of macrophages with HDL-PON1Tg, but not with HDL-PON1 °, cellular lysophosphatidylcholine (LPC) content was increased by 7.7 fold. LPC enrichment of macrophages resulted in up to 60% increased HDL binding to the cells, and up to 41% increased HDL-mediated cholesterol efflux. On using a cAMP analog to increase the ABCA1 receptor expression, or the rabbit anti-mouse SR-BI specific antibody to block the SR-BI receptor, PON1 stimulation of HDL binding and HDL-mediated cholesterol efflux fi'om macrophages was found to involve the ABCA1 receptor. In conclusion, PON1 stimulation of HDL-mediated cholesterol efflux fi'om macrophages can contribute to the attenuation of atherosclerosis development.
~
INFLAMMATION AND ENDOTHELIAL ACTIVATION ASSOCIATE WITH FAMILIAL LOW HDL
A. Soro-Paavonen, J. Westerbacka, C. Ehnholm, M. Taskinen. University of
Helsinki, and National Public Health Institute, Helsinki, Finland Objective Endothelial activation and low-grade inflammation of the arterial wall are characteristics of atherosclerosis. Inhibition of cytokine-induced expression of cellular adhesion molecules (CAMs) is one of the atheroprotective mechanisms of high density lipoprotein (HDL). We studied whether the levels of soluble CAMs are increased in subjects with familial low HDL, and whether endothelial activation associates with the metabolic syndrome in these subjects. Methods: Serum levels of the soluble forms of intercellular adhesion molecule-1 (slCAM-1), vascular cell adhesion molecule-1 (sVCAM-1), sEselectin and C-reactive protein (CRP) were measured in 91 subjects with familial low HDL fi'om Finnish low HDL families, and in 112 age-sex matched controls. The number of the features of the metabolic syndrome was determined. Results: Low HDL subjects had significantly higher sVCAM-1, slCAM1, sE-selectin and CRP than the controls (for sVCAM-1; 560-4-147ng/ml vs. 496-4-95 ng/ml, P<0.001, for slCAM-1; 247-4-60ng/ml vs. 215-4-47ng/ml, P<0.001, for sE-selectin; 52-4-20ng/ml vs 44-4-16ng/ml, P=0.020, for CRP; 1.73-4-2.05mg/1 vs. 0.85-4-1.10mg/1, P<0.001). HDL cholesterol cora'elated significantly with sVCAM-1 and with slCAM-1 when adjusted for age, gender, body mass index (BMI) and smoking. The levels of sCAMs and CRP increased according to the number of the features of the metabolic syndrome. Conclusions: The familial low HDL trait associates with low-grade inflammation and endothelial activation. Elevated concentrations of sCAMs and CRP predispose these subjects to increased cardiovascular risk.
G. Nordin Fredrikson, L. Andersson, I. S derberg, P. Dimayuga, K. Chyu, P. Shah, J. Nilsson. Exp Cardiovascular Research, Maim , Sweden;
Atherosclerosis Research Center, Los Angeles, USA Background: Oxidized LDL is present in atherosclerotic plaques where it causes cell injury and inflammation. However, immunization with oxidized LDL, as well as apolipoprotein B-100 (apo B-100) fi'agments, induces atheroprotective immune responses in apo E deficient mice. Methods and Results: Apo E deficient mice were immunized with either of three MDA-modified human apo B-100 fi'agments (P45; amino acids 688-707, P74; amino acids 1123-1142 or P240; amino acids 3613-3632) at 6, 9 and 11 weeks of age and compared to controls given carrier alone. The sequence of P240 is not homologous with the cora'esponding mouse sequence and served as control for peptide specificity. Immunization with fi'agments P45 and P74 reduced atherosclerosis in the aorta of 25-week old mice by 48% (p=0.02) and 31% (p=0.06) and macrophage content in atherosclerotic plaques by 33% (p=0.02) and 39% (p=0.02), respectively. The levels of Th2-specific IgG1 against each peptide increased more than fifty-fold in response to immunization, whereas the levels of peptide-specific IgM and Thl-associated IgG2a were only marginally affected. However, there was an increase in the plaque expression of both Thl and Th2 specific cytokines as assessed by real time PCR. Immunization with the non-homologous peptide fi'agment P240 induced a ten-fold increase of specific IgG1 antibodies, but did not influence atherosclerosis or plaque content. Condusions: These findings suggest that immunization with MDA apo B-100 fi'agments induce a shift fi'om Thl to a Th2 specific oxidized LDL antibody expression, but without a concomitant downregulation of plaque IFN-y expression. ORAL TOLERANCE AS A NEW THERAPEUTIC IW07.1751 MODALITY F O R ATHEROSCLEROSIS I i
N. Yacov, J. George, G. Halperin, E. Ishai Kovalevski, E. Breitbart, L. Bangio, A. Shish, H. Levkovitz, Y. Liberrnan, D. Harats. Vascular
Biogenics LTD, Or Yehuda and Institute of Lipid and Atherosclerosis Research, Sheba Medical Center, Tel Hashomer, Israel Inflammation is a major component in atherogenesis and among is the autoimmunity to oxidized LDL that is involved in the progression of atherosclerosis. Oral administration of autoantigens in autoimmune diseases decreases the inflammatory process and the disease outcome. We explored the effect of oral administration of a mimetic epitope of oxidized LDL on aortic sinus lesion formation and on cytokine expression in the aorta in old ApoE KO mice with advanced lesion, on chow diet. Animals were stratified by age, weight and lipid profile to the different groups. Oral administrations of the mimetic epitope of oxidized LDL, was conducted for 10 days (total of 5 doses), at three sets given at 4 weeks intervals. After 12 weeks of treatment the control fed mice exhibit a 50% increase in aortic sinus lesion, while the mimetic epitope of oxidized LDL treatment group completely blocked any fmther progression in aortic sinus lesion as compared to base line group which was sacrificed at study start. We also found an increased expression level of IL-10 and decreased expression level of IFN-y in aortas of mice treated with the mimetic epitope of oxidized LDL as compared to control mice. Our results demonstrates that oral administration of the mimetic epitope of oxidized LDL totally prevents the progression of atherosclerosis which is associated with the induction of immune-modulation fi'om inflammatory response to an anti-inflammatory response and provides a novel approach for the treatment of atherosclerosis.
74th EAS Congress, 17-20 April 2004, Seville, Spain