Multiple mechanisms operate to promote macrophage accumulation within the atherosclerotic intima. This proposed work stems in part from observations by others, including the finding that in apoE knockout mice which are deficient in the MCP-1 receptor CCR2, atherosclerosis is diminished, but lesion size and complexity still progress from 5-13 weeks. Also monocyte CCR2 expression in vitro is attenuated by several inflammatory mediators which can be in the plaque, or by differentiation to macrophages. They therefore propose that other mechanisms are at play in promoting macrophage retention and expansion within the atherosclerotic lesions. Base on their previous work, they have found that intimal macrophages express the chemokine receptor CXCR2, and the CXCR2 ligands, GROalpha, and IL-8. Initial data also indicate that leukocyte deficiency of the mouse CXCR2 homologue, mIL-8RH, arrested progression of early macrophage-rich lesions and decreased retention of lesion macrophages. They therefore hypothesize that specific CXCR2 effects mediate the intra-lesional growth, inflammatory differentiation and activation of recruited macrophages.
Their specific aims are as follows: Define the sequence of atherogenic events mediated by CXCR2, using two animal model systems. They will first transplant irradiated LDLR-/- mice with bone marrow from mIL-8/ CXCR2-/- or control mice under gnotobiotic conditions. Mice will be fed a chow or high cholesterol diet to define the role of mIL-8RH in mild vs. severe hypercholesterolemia-induced atherosclerosis. Second, to assess the role of mIL-8RH in cells other than leukocytes, they will study LDLR-/-, mIL-8RH-/- mice derived by crossbreeding. Early monocyte ingress, fatty streak formation, and lesion progression will be studied in both groups at 3 to 30 weeks. They will concurrently detect lesion mIL-8RH and macrophage markers including MOMA2 in each group of mice. They will also compare temporal and spatial macrophage accumulation, expression of mIL-8RH, KC/GROalpha, JE/MCP-1 and CCR2, the appearance of oxLDL epitopes and lipid deposition. Test the hypothesis that CXCR2 expression acts by promoting macrophage retention in early atherosclerotic lesions, and does so in part by increasing macrophage beta1 integrin-mediated adhesion. First they will determine if retention of lesion monocytes is greater in LDLR-/-, mIL-8RH+/+ than LDLR-/-, mIL-8RH-/- mice. To do so they will isolate, label (using [3H]-glycerol) and infuse bone marrow monocytes into LDLR-/- mice on a high fat diet. Migration and retention of labeled monocytes in vivo will be measured in aortic lesions at 2,4,7, and 14 days post infusion. They will then test the hypothesis that the generalized cell culture adhesion defect of macrophages from mIL-8RH-/- mice is associated with decreased activation of beta1 integrins, which bind fibronectin (VLA-4, VLA-5). They will also study CXCR2-mediated effects on adhesion of human peripheral blood monocytes using specific neutralizing antibodies for integrins and integrin ligands. They will also test the hypothesis that macrophage CXCR2 expression is necessary for macrophage matrix invasion. Test the hypothesis that CXCR2 expression, via effects on macrophage adhesion, mediates macrophage differentiation to a distinct, pro-atherogenic inflammatory phenotype. Here they will further study human peripheral monocyte-derived macrophages and mouse wild type and mIL-8RH-/- bone marrow derived macrophages under conditions where CXCR2 is normally expressed in vitro. They will specifically look at the effects of the CXCR2 ligands, GROalpha, and IL-8 on macrophage proliferation and expression of JE/MCP-1. They will also determine if CXCCR2-mediated adhesion modulates the production of apoE, oxidation of LDL, and accumulation of cholesteryl ester. Finally based on the results of Specific Aim 2, they will test the hypothesis that VLA-5, and VLA-4 activation, via stimulation by CXCR2 ligands, regulate one or more of these activities. They will also correlate these results with the features of the mouse atherosclerotic lesions of Specific Aim 1 (i.e. in vivo correlation studies between CXCR2 expression and PCNA, oxidized LDL antigens, and scavenger receptor expression).

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL061731-03
Application #
6390157
Study Section
Pathology A Study Section (PTHA)
Program Officer
Applebaum-Bowden, Deborah
Project Start
1999-09-01
Project End
2003-08-31
Budget Start
2001-09-01
Budget End
2002-08-31
Support Year
3
Fiscal Year
2001
Total Cost
$357,569
Indirect Cost
Name
Scripps Research Institute
Department
Type
DUNS #
City
La Jolla
State
CA
Country
United States
Zip Code
92037
Boisvert, William A; Rose, David M; Johnson, Kristen A et al. (2006) Up-regulated expression of the CXCR2 ligand KC/GRO-alpha in atherosclerotic lesions plays a central role in macrophage accumulation and lesion progression. Am J Pathol 168:1385-95
Liu, Ru; Liote, Frederic; Rose, David M et al. (2004) Proline-rich tyrosine kinase 2 and Src kinase signaling transduce monosodium urate crystal-induced nitric oxide production and matrix metalloproteinase 3 expression in chondrocytes. Arthritis Rheum 50:247-58
Rutsch, Frank; Terkeltaub, Robert (2003) Parallels between arterial and cartilage calcification: what understanding artery calcification can teach us about chondrocalcinosis. Curr Opin Rheumatol 15:302-10
Schraufstatter, Ingrid U; Trieu, Khanh; Zhao, Ming et al. (2003) IL-8-mediated cell migration in endothelial cells depends on cathepsin B activity and transactivation of the epidermal growth factor receptor. J Immunol 171:6714-22
Pinderski, Laura J; Fischbein, Michael P; Subbanagounder, Ganesamoorthy et al. (2002) Overexpression of interleukin-10 by activated T lymphocytes inhibits atherosclerosis in LDL receptor-deficient Mice by altering lymphocyte and macrophage phenotypes. Circ Res 90:1064-71
Johnson, K; Hashimoto, S; Lotz, M et al. (2001) Interleukin-1 induces pro-mineralizing activity of cartilage tissue transglutaminase and factor XIIIa. Am J Pathol 159:149-63
Rutsch, F; Vaingankar, S; Johnson, K et al. (2001) PC-1 nucleoside triphosphate pyrophosphohydrolase deficiency in idiopathic infantile arterial calcification. Am J Pathol 158:543-54
Schiller, N K; Kubo, N; Boisvert, W A et al. (2001) Effect of gamma-irradiation and bone marrow transplantation on atherosclerosis in LDL receptor-deficient mice. Arterioscler Thromb Vasc Biol 21:1674-80
Terkeltaub, R A (1999) IL-10: An ""immunologic scalpel"" for atherosclerosis? Arterioscler Thromb Vasc Biol 19:2823-5