Serum amyloid A and lipoprotein retention in murine models of atherosclerosis.

OBJECTIVE: Elevated serum amyloid A (SAA) levels are associated with increased cardiovascular risk in humans. Because SAA associates primarily with lipoproteins in plasma and has proteoglycan binding domains, we postulated that SAA might mediate lipoprotein retention on atherosclerotic extracellular matrix. METHODS AND RESULTS: Immunohistochemistry was performed for SAA, apolipoprotein A-I (apoA-I), apolipoprotein B (apoB), and perlecan on proximal aortic lesions from chow-fed low-density lipoprotein receptor (LDLR)-/- and apoE-/- mice euthanized at 10, 50, and 70 weeks. SAA was detected on atherosclerotic lesion extracellular matrix at all time points in both strains. SAA area correlated highly with lesion areas (apoE-/-, r=0.76; LDLR-/-, r=0.86), apoA-I areas (apoE-/-, r=0.88; LDLR-/-, r=0.80), apoB areas (apoE-/-, r=0.74; LDLR-/-, r=0.89), and perlecan areas (apoE-/-, r=0.83; LDLR-/-, r=0.79) (all P<0.0001). In vitro, SAA enrichment increased high-density lipoprotein (HDL) binding to heparan sulfate proteoglycans, and immunoprecipitation experiments using plasma from apoE-/- and LDLR-/- mice demonstrated that SAA was present on both apoA-I-containing and apoB-containing lipoproteins. CONCLUSIONS: In chow-fed apoE-/- and LDLR-/- mice, SAA is deposited in murine atherosclerosis at all stages of lesion development, and SAA immunoreactive area correlates highly with lesion area, apoA-I area, apoB area, and perlecan area. These findings are consistent with a possible role for SAA-mediated lipoprotein retention in atherosclerosis.

Serum amyloid P colocalizes with apolipoproteins in human atheroma: functional implications.

Serum amyloid P (SAP) is a common component of human amyloid deposits and has been identified in atherosclerotic lesions. We investigated the extent of the colocalization of SAP with apolipoprotein A-I (apoA-I), apoB, apoC-II, and apoE in human coronary arteries and explored potential roles for SAP in these regions, specifically the effect of SAP on the rate of formation and macrophage recognition of amyloid fibrils composed of apoC-II. Analysis of 42 human arterial sections by immunohistochemistry and double label fluorescence microscopy demonstrated that SAP and apoA-I, apoB, apoC-II, and apoE were increased significantly in atherosclerotic lesions compared with nonatherosclerotic segments. SAP colocalized with all four apolipoproteins to a similar extent, whereas plaque macrophages were found to correlate most strongly with apoC-II and apoB. In vitro studies showed that SAP accelerated the formation of amyloid fibrils by purified apoC-II. Furthermore, SAP strongly inhibited the phagocytosis of apoC-II amyloid fibrils by primary macrophages and macrophage cell lines and blocked the resultant production of reactive oxygen species. The ability of SAP to accelerate apoC-II amyloid fibril formation and inhibit macrophage recognition of apoC-II fibrils suggests that SAP may modulate the inflammatory response to amyloid fibrils in atherosclerosis.

Fibrillar amyloid protein present in atheroma activates CD36 signal transduction.

The self-association of proteins to form amyloid fibrils has been implicated in the pathogenesis of a number of diseases including Alzheimer's, Parkinson's, and Creutzfeldt-Jakob diseases. We recently reported that the myeloid scavenger receptor CD36 initiates a signaling cascade upon binding to fibrillar beta-amyloid that stimulates recruitment of microglia in the brain and production of inflammatory mediators. This receptor plays a key role in the pathogenesis of atherosclerosis, prompting us to evaluate whether fibrillar proteins were present in atherosclerotic lesions that could initiate signaling via CD36. We show that apolipoprotein C-II, a component of very low and high density lipoproteins, readily forms amyloid fibrils that initiate macrophage inflammatory responses including reactive oxygen production and tumor necrosis factor alpha expression. Using macrophages derived from wild type and Cd36(-/-) mice to distinguish CD36-specific events, we show that fibrillar apolipoprotein C-II activates a signaling cascade downstream of this receptor that includes Lyn and p44/42 MAPKs. Interruption of this signaling pathway through targeted deletion of Cd36 or blocking of p44/42 MAPK activation inhibits macrophage tumor necrosis factor alpha gene expression. Finally, we demonstrate that apolipoprotein C-II in human atheroma co-localizes to regions positive for markers of amyloid and macrophage accumulation. Together, these data characterize a CD36-dependent signaling cascade initiated by fibrillar amyloid species that may promote atherogenesis.