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4P-1074 Postprandial activation of neutrophilic granulocytes and monocytes in humans
310
Thursday October 2, 2003: Poster Session Immune response and inflammation
is known regarding the dynamics of host-derived cells in the graft media, we investigated this question and in this context the build up of the neointima after mouse carotid artery transplantation. Methods: C57BL/6 carotid arteries were orthotopically transplanted into BALB/c mice ubiquitously expressing enhanced green fluorescent protein (GFP). Grafts were harvested at 1, 2, 4 and 8 weeks after transplantation for histology. No immunosuppression was used. Results: Immunostaining and semiquantitative analysis of cross sections showed that donor medial smooth muscle cells (SMCs) decreased time dependently, whereas GFP-positive/α-actin positive cells, i.e. cells of host origin, increased over time. Interestingly, host cells were located only in the inner media and the neointima at 2 weeks, thereafter also in the outer media, indicating that the host cells entered the media from the luminal side of the vessel rather than from the adventitia. Conclusion: The medial cell replacement and the neointima formation by host cells expressing α-actin, probably derived from circulating precursor cells, might be a healing process to restore vascular function following transplantation. Our observations differ from the traditional view that the donor medial SMCs dedifferentiate and migrate to form the subintima. Hence therapeutic strategies for transplant vasculopathy may have to focus on circulating progenitor cells. 4P-1071
BCG immunization is atherogenic in the chronically hypercholesterolaemic rabbit: Immune and vascular endothelial effects
D. Lamb, M. Tickner, A. Dreux, W. El-Sankary, S. Hourani, L.-J. Eales-Reynolds, G. Ferns. University of Surrey, Guildford, Surrey, United Kingdom A number of studies have reported associations between immune responses to heat shock protein-60/65 (HSP) and coronary disease. The aim of this study was to use a BCG immunisation model to investigate the relationship between anti-HSP immune responses, endothelial dysfunction and atherosclerosis in the chronic low-level hypercholesterolaemic rabbit model. Rabbits were injected with bacillus Calmette-Guerin (BCG) vaccine (n=12) or saline (n=12), boosted after 2 weeks and after a further 2 weeks fed either a 0.25-1% cholesterol diet or a chow diet for 16 weeks. Blood cholesterol levels were maintained at 10-12mmol/l by altering the dietary cholesterol content. Plasma levels of anti-mycobacterial antibodies rose following BCG immunisation, but anti-HSP antibodies developed only in the BCG-immunised, cholesterol-fed rabbits. BCG immunisation increased surface expression of CD11b on mononuclear cells and BCG immunised, cholesterol-fed rabbits developed more extensive aortic lesions (p<0.05) and thicker intima (p<0.01) compared to control animals. Lesions from BCG-immunised animals contained significantly more macrophages compared to controls (p<0.05). Aortic endothelium from cholesterol-fed, but not chow-fed, rabbits stained positively for HSP-60, independently of the immunisation protocol. Endothelial function was impaired in the BCG immunised, cholesterol-fed rabbits as measured by acetylcholine-mediated relaxation of isolated carotid artery rings (p<0.05). This impairment was positively associated with the level of plasma anti-HSP60 antibodies (p<0.01). These results suggest that BCG immunisation impairs endothelial responses and increases atherosclerosis in chronic low-level hypercholesterolaemic rabbits. This is mediated, at least in part, by immune responses against mycobacterial and vascular HSP. This work was sponsored by the BHF. 4P-1072
Research on the atherosclerotic lesions in coronary artery of apoE gene knockout mice
B. Xue 1 , L. Gao 2 , R. Chen 1 , J. Wang 1 , L. Li 1 , W. Hu 1 , P.D. Polinsky 3 , S.M. Schwartz 3 . 1 Dept. of Pathophysiology, School of Medicine, Shandong University; 2 Dept. of Physiology, Taishan Medical College, China; 3 Dept. of Pathology, University of Washington, USA Objective: Study the development of the atherosclerotic(AS) lesions in coronary artery(CA) of apoE knockout (-/-) mice. Methods: Make the successive sections of the hearts of apoE-/- mice and stain with Movat method. Use the electron microscope to identify the inflammatory cells at the adventitia. Results: There were lesions extending from the aorta directly into the trunks of CA. Extending lesions could only be found in trunks of CA with adventitial inflammation, no extending lesion could be found in trunks of CA without adventitia inflammation. The area of inflammatory infiltration in adventitia was larger than the area of extending lesions in intima. Electron
microscope showed that the inflammatory cells were mainly lymphocytes, macrophages, eosinophilic granulocytes and plasmacytes. Focal adventitial inflammation often exited in the small branch of vessels (including intramyocardial small branches), where some had in situ lesions in corresponding intima. Inflammatory cells aggregation could be seen in the corresponding adventitia with in situ lesions in the intima. The area of AS lesion in intima was smaller than the area of inflammatory cells infiltration in adventitia. The in situ lesions consisted of mainly proteoglycan and lipids. All the in situ lesions formed within the ventricular muscle. Most of them appeared in left ventricular wall. The lesions of intra-myocardial small branches tended to be located at the papillary muscles and near the bifurcation of vessels. Conclusion: There were extending lesions in the trunks of the CA of apoE-/- mice and in suit lesions in small branches. Adventitial inflammation induced the extending of the extending lesions and the formation of the in situ lesions. The formation of adventitia inflammation was related to the dynamic press upon the vessels by myocardial contraction. 4P-1073
15-Deoxy-∆12,14 -prostaglandin J2 inhibits the expression of fractalkine in endothelial cells
T. Imaizumi, H. Yoshida, K. Satoh. Hirosaki University School of Medicine, Hirosaki, Japan Objective: CX3CL1/fractalkine is a potent agonist for chemotaxis and adhesion of monocytes and lymphocytes. Endothelial cells produce fractalkine when stimulated with cytokines such as interleukin-1 (IL-1), tumor necrosis factor-α and interferon-γ (IFN-γ). 15-deoxy-12,14 -prostaglandin J2 (15dPGJ2 ) is a natural ligand of peroxisome proliferator-activated receptor-γ (PPAR-γ), and is known to regulate the expression of various genes which are related with inflammatory reactions. In this study, we examined the effect of 15d-PGJ2 on the expression of fractalkine in vascular endothelial cells. Methods: Human umbilical vein endothelial cells (HUVEC) were pretreated with 15d-PGJ2 and then stimulated with IL-1β or IFN-γ. Expression of fractalkine was examined by RT-PCR and western blot analyses. Adhesion of mononuclear cells (MNCs), isolated from human peripheral blood, to HUVEC was also examined. Results: Pretreatment of HUVEC with 15d-PGJ2 inhibited the induction of fractalkine mRNA and protein by IL-1β or IFN-γ. Adhesion of MNCs to HUVEC induced by IL-1β or IFN-γ was also inhibited by 15d-PGJ2 . Conclusion: 15d-PGJ2 regulates inflammatory reactions, at least in part, by inhibiting the expression of fractalkine in vascular endothelial cells. 4P-1074
Postprandial activation of neutrophilic granulocytes and monocytes in humans
A. Van Oostrom 1 , T. Rabelink 1 , C. Verseyden 1 , T. Plokker 2 , P. De Jaegere 2 , M. Castro Cabezas 1 . 1 Department of Vascular Medicine, UMC, Utrecht; 2 Heart Lung Institute, Utrecht, Netherlands Background: Low-grade inflammatory processes play an important role in atherosclerosis. Activated leukocytes in the blood adhering to the endothelium via specific ligands are obligatory for the development of atherosclerosis. Although in vitro studies have shown that triglycerides (TG) activate leukocytes, it is unknown whether this occurs in vivo. Methods: We studied the expression of leukocyte activation markers CD11A, CD11B, CD62L (all involved in endothelium adhesion) and CD66B (a neutrophil degranulation marker) during a 6 hours fat challenge (50 g/m2 ) in 20 healthy males (50±5 years) using flowcytometry. Results: Postprandially, monocyte counts did not change and neutrophils increased between t=1 and t=6h, with a maximum at t=3h (26% higher than t=0, p<0.005) while lymphocytes increased gradually to a maximum at t=6h (21% higher than baseline, p<0.001). A control test in 6 subjects who ingested water showed that only the neutrophil increment was fat-specific. After the fat load, the expression of activation markers on lymphocytes did not change. A time-dependent increase up to a maximum at t=6h was seen for CD11B on monocytes (35% increase) and neutrophils for CD11A (+14%), CD11B (+87%), CD62L (+10%) and CD66B (+24%, p<0.05 vs baseline, for all comparisons). The control test with water showed that these increments were fat-specific. The total incremental TG response was positively related to the increase of CD11B on monocytes and neutrophils (R=0.49 and R=0.58 respectively, p<0.05 for both). Conclusion: In the postprandial phase there is a TG-specific increase in monocyte activation and neutrophil cell count and activation. These results are suggestive of a postprandial pro-inflammatory situation and may represent increased adhesive capacity of neutrophils and monocytes to the endothelium.
XIIIth International Symposium on Atherosclerosis, September 28–October 2, 2003, Kyoto, Japan
Thursday October 2, 2003: Poster Session Immune response and inflammation
is known regarding the dynamics of host-derived cells in the graft media, we investigated this question and in this context the build up of the neointima after mouse carotid artery transplantation. Methods: C57BL/6 carotid arteries were orthotopically transplanted into BALB/c mice ubiquitously expressing enhanced green fluorescent protein (GFP). Grafts were harvested at 1, 2, 4 and 8 weeks after transplantation for histology. No immunosuppression was used. Results: Immunostaining and semiquantitative analysis of cross sections showed that donor medial smooth muscle cells (SMCs) decreased time dependently, whereas GFP-positive/α-actin positive cells, i.e. cells of host origin, increased over time. Interestingly, host cells were located only in the inner media and the neointima at 2 weeks, thereafter also in the outer media, indicating that the host cells entered the media from the luminal side of the vessel rather than from the adventitia. Conclusion: The medial cell replacement and the neointima formation by host cells expressing α-actin, probably derived from circulating precursor cells, might be a healing process to restore vascular function following transplantation. Our observations differ from the traditional view that the donor medial SMCs dedifferentiate and migrate to form the subintima. Hence therapeutic strategies for transplant vasculopathy may have to focus on circulating progenitor cells. 4P-1071
BCG immunization is atherogenic in the chronically hypercholesterolaemic rabbit: Immune and vascular endothelial effects
D. Lamb, M. Tickner, A. Dreux, W. El-Sankary, S. Hourani, L.-J. Eales-Reynolds, G. Ferns. University of Surrey, Guildford, Surrey, United Kingdom A number of studies have reported associations between immune responses to heat shock protein-60/65 (HSP) and coronary disease. The aim of this study was to use a BCG immunisation model to investigate the relationship between anti-HSP immune responses, endothelial dysfunction and atherosclerosis in the chronic low-level hypercholesterolaemic rabbit model. Rabbits were injected with bacillus Calmette-Guerin (BCG) vaccine (n=12) or saline (n=12), boosted after 2 weeks and after a further 2 weeks fed either a 0.25-1% cholesterol diet or a chow diet for 16 weeks. Blood cholesterol levels were maintained at 10-12mmol/l by altering the dietary cholesterol content. Plasma levels of anti-mycobacterial antibodies rose following BCG immunisation, but anti-HSP antibodies developed only in the BCG-immunised, cholesterol-fed rabbits. BCG immunisation increased surface expression of CD11b on mononuclear cells and BCG immunised, cholesterol-fed rabbits developed more extensive aortic lesions (p<0.05) and thicker intima (p<0.01) compared to control animals. Lesions from BCG-immunised animals contained significantly more macrophages compared to controls (p<0.05). Aortic endothelium from cholesterol-fed, but not chow-fed, rabbits stained positively for HSP-60, independently of the immunisation protocol. Endothelial function was impaired in the BCG immunised, cholesterol-fed rabbits as measured by acetylcholine-mediated relaxation of isolated carotid artery rings (p<0.05). This impairment was positively associated with the level of plasma anti-HSP60 antibodies (p<0.01). These results suggest that BCG immunisation impairs endothelial responses and increases atherosclerosis in chronic low-level hypercholesterolaemic rabbits. This is mediated, at least in part, by immune responses against mycobacterial and vascular HSP. This work was sponsored by the BHF. 4P-1072
Research on the atherosclerotic lesions in coronary artery of apoE gene knockout mice
B. Xue 1 , L. Gao 2 , R. Chen 1 , J. Wang 1 , L. Li 1 , W. Hu 1 , P.D. Polinsky 3 , S.M. Schwartz 3 . 1 Dept. of Pathophysiology, School of Medicine, Shandong University; 2 Dept. of Physiology, Taishan Medical College, China; 3 Dept. of Pathology, University of Washington, USA Objective: Study the development of the atherosclerotic(AS) lesions in coronary artery(CA) of apoE knockout (-/-) mice. Methods: Make the successive sections of the hearts of apoE-/- mice and stain with Movat method. Use the electron microscope to identify the inflammatory cells at the adventitia. Results: There were lesions extending from the aorta directly into the trunks of CA. Extending lesions could only be found in trunks of CA with adventitial inflammation, no extending lesion could be found in trunks of CA without adventitia inflammation. The area of inflammatory infiltration in adventitia was larger than the area of extending lesions in intima. Electron
microscope showed that the inflammatory cells were mainly lymphocytes, macrophages, eosinophilic granulocytes and plasmacytes. Focal adventitial inflammation often exited in the small branch of vessels (including intramyocardial small branches), where some had in situ lesions in corresponding intima. Inflammatory cells aggregation could be seen in the corresponding adventitia with in situ lesions in the intima. The area of AS lesion in intima was smaller than the area of inflammatory cells infiltration in adventitia. The in situ lesions consisted of mainly proteoglycan and lipids. All the in situ lesions formed within the ventricular muscle. Most of them appeared in left ventricular wall. The lesions of intra-myocardial small branches tended to be located at the papillary muscles and near the bifurcation of vessels. Conclusion: There were extending lesions in the trunks of the CA of apoE-/- mice and in suit lesions in small branches. Adventitial inflammation induced the extending of the extending lesions and the formation of the in situ lesions. The formation of adventitia inflammation was related to the dynamic press upon the vessels by myocardial contraction. 4P-1073
15-Deoxy-∆12,14 -prostaglandin J2 inhibits the expression of fractalkine in endothelial cells
T. Imaizumi, H. Yoshida, K. Satoh. Hirosaki University School of Medicine, Hirosaki, Japan Objective: CX3CL1/fractalkine is a potent agonist for chemotaxis and adhesion of monocytes and lymphocytes. Endothelial cells produce fractalkine when stimulated with cytokines such as interleukin-1 (IL-1), tumor necrosis factor-α and interferon-γ (IFN-γ). 15-deoxy-12,14 -prostaglandin J2 (15dPGJ2 ) is a natural ligand of peroxisome proliferator-activated receptor-γ (PPAR-γ), and is known to regulate the expression of various genes which are related with inflammatory reactions. In this study, we examined the effect of 15d-PGJ2 on the expression of fractalkine in vascular endothelial cells. Methods: Human umbilical vein endothelial cells (HUVEC) were pretreated with 15d-PGJ2 and then stimulated with IL-1β or IFN-γ. Expression of fractalkine was examined by RT-PCR and western blot analyses. Adhesion of mononuclear cells (MNCs), isolated from human peripheral blood, to HUVEC was also examined. Results: Pretreatment of HUVEC with 15d-PGJ2 inhibited the induction of fractalkine mRNA and protein by IL-1β or IFN-γ. Adhesion of MNCs to HUVEC induced by IL-1β or IFN-γ was also inhibited by 15d-PGJ2 . Conclusion: 15d-PGJ2 regulates inflammatory reactions, at least in part, by inhibiting the expression of fractalkine in vascular endothelial cells. 4P-1074
Postprandial activation of neutrophilic granulocytes and monocytes in humans
A. Van Oostrom 1 , T. Rabelink 1 , C. Verseyden 1 , T. Plokker 2 , P. De Jaegere 2 , M. Castro Cabezas 1 . 1 Department of Vascular Medicine, UMC, Utrecht; 2 Heart Lung Institute, Utrecht, Netherlands Background: Low-grade inflammatory processes play an important role in atherosclerosis. Activated leukocytes in the blood adhering to the endothelium via specific ligands are obligatory for the development of atherosclerosis. Although in vitro studies have shown that triglycerides (TG) activate leukocytes, it is unknown whether this occurs in vivo. Methods: We studied the expression of leukocyte activation markers CD11A, CD11B, CD62L (all involved in endothelium adhesion) and CD66B (a neutrophil degranulation marker) during a 6 hours fat challenge (50 g/m2 ) in 20 healthy males (50±5 years) using flowcytometry. Results: Postprandially, monocyte counts did not change and neutrophils increased between t=1 and t=6h, with a maximum at t=3h (26% higher than t=0, p<0.005) while lymphocytes increased gradually to a maximum at t=6h (21% higher than baseline, p<0.001). A control test in 6 subjects who ingested water showed that only the neutrophil increment was fat-specific. After the fat load, the expression of activation markers on lymphocytes did not change. A time-dependent increase up to a maximum at t=6h was seen for CD11B on monocytes (35% increase) and neutrophils for CD11A (+14%), CD11B (+87%), CD62L (+10%) and CD66B (+24%, p<0.05 vs baseline, for all comparisons). The control test with water showed that these increments were fat-specific. The total incremental TG response was positively related to the increase of CD11B on monocytes and neutrophils (R=0.49 and R=0.58 respectively, p<0.05 for both). Conclusion: In the postprandial phase there is a TG-specific increase in monocyte activation and neutrophil cell count and activation. These results are suggestive of a postprandial pro-inflammatory situation and may represent increased adhesive capacity of neutrophils and monocytes to the endothelium.
XIIIth International Symposium on Atherosclerosis, September 28–October 2, 2003, Kyoto, Japan