Inflammation and atherosclerosis Immune cells dominate early lesions Immune effector molecules accelerate lesion progression Activation of inflammation elicits acute coronary syndromes Atherosclerosis is an inflammatory disease in which immune mechanisms interact with metabolic risk factors to initiate, propagate and activate lesions in the arterial wall

The numbers CAD causes 38% of all deaths in North America Most common cause of death in men under 65 y.o. and second most common cause of women Dispite advances in control of hypercholesterolemia (statins),CVD expected to be the main cause of death globally over the next 15 years due to rapidly increasing prevalence of obesity and diabetes

Pathogenesis of coronary artery thrombosis

Cellular players in atherogenesis and plaque rupture

The early lesion: modified lipoproteins and foam cell formation

Role of macrophages MCSF induces monocytes entering lesion to differentiate into macrophages Macrophage differentiation associated with TLRs and scavenger receptors MCSF/apoE KO shows reduced atherosclerosis Scavenger receptors bind bacterial endotoxins, apoptotic cells and oxLDL SR/apoE KOs show reduced atherosclerosis

The macrophage as an inflammatory mediator

Toll-like GENES IL-1R TNF-R IFN-  R IRF NF  B MAP KINASE JAK/STAT Inflammation, Anti-Microbial Response, Apoptosis, …

TLR receptors and atherosclerosis 10 family members Recognize pathogen associated molecular patterns (e.g. LPS, dsRNA) as well as oxLDL, HSP60 etc. Initiate signaling cascades leading to production of inflammatory cytokines, proteases, reactive oxygen species In addition to macrophages, expressed by dendritic cells, mast cells, endothelial cells

Macrophage-T cell interactions in atherosclerosis

T cells and atherosclerosis Immune cells patrol tissues in search of antigen T cell infiltrate is common feature of atherosclerotic lesions Predominantly CD4+ cells, recognize antigen/MHC II CD4+ T cells reactive to oxLDL, HSP60, bacterial products detected in human lesions NK cells present in early lesions, recognize lipid antigens NK activation increases athero in apoE KO mice

T cell responses Th1 response activates macrophages and functions in the defense against intracellular pathogens Th2 response elicits allergic inflammation Atherosclerotic lesions contains cytokines that promote Th1 responses Activated Th1 effector cells in lesions produce macrophage activating cytokine IFN  IFN  improves efficiency of ag presentation and augments synthesis of TNF  and IL-1 IFN , TNF  and IL-1 in turn stimulate production of many other inflammatory mediators apoE mice lacking IFN  or downstream mediators such as IL-18 or T-bet show reduced athero

Anti-inflammatory factors and atherosclerosis Anti-inflammatory factors such as TGF  and IL-10 are protective IL-10 KO increases athero in mice and exacerbates thrombosis Abrogation of TGF signaling in T cells leads to large unstable atherosclerotic lesions

Immune cells and plaque rupture Preferentially occurs where fibrous cap is thin Active immune cells are abundant at site of rupture Immune cells produce inflammatory molecules and proteolytic enzymes that weaken cap, activate cells in the core and transform stable plaque into vulnerable, leading to plaque rupture MMPs likely to play important roles

Systemic inflammatory markers in atherosclerosis

Systemic indicators of inflammation Inflammatory process in lesions may lead to increased plasma levels of cytokines and acute phase proteins CRP and IL-6 are elevated in patients with unstable angina and MI Levels correlate with prognosis

TLR-3/4 IRF-3 NEMO IKK-  IKK  TANK TBK-1IKK i I  NF-  MyD88 “Independent” MyD88 “Dependent” Antiviral Response Inflammatory Response

MyD88/apoE KO mice show reduced athero

MyD88/apoE Ko mice show reduced chemokine expression

MyD88/apoE KO macs show reduced chemokine expression and recruitment in response to inflammatory cytokines

Unlikely to be caused by single organism Diverse pathogens have been detected within lesions Bacteria and viruses accelerate athero in murine models Links between infection and atherosclerosis

Potential mechanisms Stimulation of inflammatory cytokines Alteration of adhesion molecule expression ? Lipid metabolism ?

RXR LXR INFLAMMATION CHOLESTEROL EFFLUX CE oxLDLoxysterol cholesterol CD36  PLTP  ApoE/CII  ABCAI  LPL  iNOS  IL-6  IL-1   COX2  MMP9 TOLL4 IL-1R TNFR Inflammatory stimuli stimuli LPS, IL-1 , TNF  ABCA1 MACROPHAGE

p < Lesion area (  M 2 /section) + GW3965high fat LDLR-/- Aortic Sections 0 p < GW3965vehicle Lesion area (  M 2 /section) ApoE-/- Aortic Sections Synthetic LXR agonist reduces atherosclerosis inLDLR-/- and apoE-/- mice Joseph et al. PNAS 2002

C57Bl6BMT LXR  -/- BMT apoE -/- BMT Aortic Lesion Coverage (%) * * Loss of bone marrow LXR expression accelerates athero Tangirala et al. PNAS 2002

Bacterial or viral infection blocks expression of LXR target genes in macrophages ctrl GW3965T1317E. ColiE. Coli + GW E. Coli + T ABCA1 apoE 36B4 FAS SREBP-1c iNOS ctrl GW3965T1317Infl AInfl A + GWInfl A + T ABCA1 ABCG1 MX-1 FAS IFN-  36B4 +/- E. coli +/- Influenza A Castrillo et al., Molecular Cell 2003 Antonio Castrillo, Sean Joseph

TLR3 and TLR4 ligands inhibit cholesterol efflux from macrophages Ctrl LTA Poly IC Lipid A TNF  apoAI-dependent efflux (%) ** * * 14 Ctrl LTA Poly IC Lipid A TNF  Ctrl LXR ligand Cholesterol efflux

ABCA apoE ABCG IFN-  IP-10 Spleen * * * * mRNA ABCA1 apoE IFN-  IP-10 Vehicle Poly IC ABCG Aorta 19 * * * mRNA Vehicle Poly IC Vehicle Poly IC Vehicle Poly IC vehicle Poly IC Activation of TLR3 inhibits LXR signaling in vivo GW3965 Both

Foam cell formation bacteri a IRF-3 virus cholesterol efflux C C C C C C ABCA1 TLR-3/4 ABCA1, ABCG1, apoE LXRRXR TLR-LXR crosstalk: potential implications for atherosclerosis  Foam cell formation other signals TANK TBK-1 IKK i ?

Immunization with pneumococus induces oxLDL-specific IgM

Immunization with pneumococus induces oxLDL-specific IgM s. pneumo s. pneumo + oxLDL block s. pneumo + s. pneumo block E06 (ox PL)

Plasma from mice immunized with pneumococus inhibits macrophage binding of oxLDL

Immunization with pneumococus reduces atherosclerosis in LDLR-/- mice