Toll-like receptor 4-dependent and -independent cytokine secretion induced by minimally oxidized low-density lipoprotein in macrophages.
OBJECTIVE
Innate immune responses to oxidized low-density lipoprotein LDL (LDL) regulate the development of atherosclerosis. We demonstrated previously that an early form of oxidized LDL, minimally modified LDL (mmLDL), triggers cytoskeletal rearrangements in macrophages via CD14 and Toll-like receptor 4 (TLR4)/MD-2. Because lipopolysaccharide (LPS) activation of TLR4 leads to proinflammatory gene expression, in this study, we asked whether mmLDL also induced proinflammatory signaling.


RESULTS
We studied cytokine secretion and signaling in J774 and primary peritoneal macrophages stimulated with mmLDL, which was prepared by incubating LDL with cells expressing human 15-lipoxygenase. MmLDL stimulated robust phosphoinositide 3-kinase (PI3K) activation, and Akt and extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation, which exceeded that induced by LPS. On the other hand, although mmLDL induced nuclear factor kappaB (NF-kappaB) p65 translocation to the nucleus, there was no detectable NF-kappaB activation. However, mmLDL induced early mRNA and protein expression of the cytokines MIP-2, MCP-1, tumor necrosis factor-alpha, and interleukin-6. Chemokine MIP-2 but not MCP-1 secretion depended on TLR4/MyD88, ERK1/2, and PI3K signaling. In turn, TLR4 regulated phosphorylation of ERK1/2 but not of Akt, suggesting that mmLDL-induced PI3K activation is TLR4 independent.


CONCLUSIONS
In macrophages, mmLDL activates TLR4-dependent and -independent signaling pathways, resulting in secretion of proinflammatory cytokines. These results provide new insights into the inflammatory origins of atherosclerosis.
Miller
Yury I
YI
Division of Endocrinology and Metabolism, San Diego Healthcare System and Department of Pathology and Medicine, University of California, San Diego, La Jolla, CA 92093-0682, USA. yumiller@ucsd.edu
Viriyakosol
Suganya
S
Worrall
Dorothy S
DS
Boullier
Agnès
A
Butler
Susan
S
Witztum
Joseph L
JL
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics; D000818:Animals; D000943:Antigens, Differentiation / Q000235:genetics; D050197:Atherosclerosis / Q000276:immunology* / Q000378:metabolism; D016466:CHO Cells; D054407:Chemokine CCL4; D054426:Chemokine CXCL2; D018925:Chemokines / Q000235:genetics / Q000378:metabolism; D006224:Cricetinae; D016207:Cytokines / Q000235:genetics / Q000378:metabolism*; D005260:Female; D005347:Fibroblasts / Q000166:cytology; D008077:Lipoproteins, LDL / Q000378:metabolism*; D019402:Macrophage Inflammatory Proteins / Q000235:genetics / Q000378:metabolism; D017737:Macrophages, Peritoneal / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011971:Receptors, Immunologic / Q000235:genetics; D015398:Signal Transduction / Q000276:immunology; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D000943:Antigens, Differentiation; D054407:Chemokine CCL4; D054426:Chemokine CXCL2; D018925:Chemokines; C404915:Cxcl2 protein, mouse; D016207:Cytokines; D008077:Lipoproteins, LDL; C507779:MYD88 protein, human; D019402:Macrophage Inflammatory Proteins; C507780:Myd88 protein, mouse; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D011971:Receptors, Immunologic; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; D014409:Tumor Necrosis Factor-alpha; C099252:oxidized low density lipoprotein; D019869:Phosphatidylinositol 3-Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3