Abstract Body (Do not enter title and authors here): Background
Atherosclerosis is a chronic vascular disease characterized by lipid accumulation within the artery wall. While scavenger receptor CD36 helps macrophages remove oxidized low-density lipoprotein (oxLDL), excessive oxLDL engulfment promotes macrophage foam cell formation. Focal adhesion kinase (FAK), an integrin-associated tyrosine kinase, promotes receptor signaling, but its role in CD36 signaling in macrophages remains unclear. We found FAK activation promotes Filamin A (FLNA) phosphorylation and enhances oxLDL uptake via CD36 in macrophages; however, the mechanism by which FLNA increases oxLDL uptake remains unknown.
Hypothesis
FAK and FLNA crosstalk promotes oxLDL-CD36 endocytosis, leading to macrophage foam cell formation.
Methods
We evaluated FAK and FLNA roles in CD36 mediated-oxLDL uptake and internalization using cell surface biotinylation and dil-oxLDL uptake in Raw264.7 and murine bone marrow-derived macrophages. FAK, FLNA, and CD36 expression and interactions were evaluated by co-immunoprecipitation, immunoblotting, and staining. In mouse atherosclerosis models, plaques were examined using Oil Red O (ORO) staining.
Results
FAK inhibition (FAK-I) blocked oxLDL-induced FAK activation and CD36 internalization reducing foam cell formation. OxLDL increased FLNA tyrosine phosphorylation and FAK-FLNA -CD36 complex formation but was blocked by FAK-I. FLNA shRNA (shFLNA) reduced oxLDL-induced FAK activation and internalization of oxLDL and CD36. shFLNA also reduced CD36 protein expression and FAK-CD36 interaction, suggesting FLNA-FAK crosstalk may regulate CD36 protein stability. In vivo, FAK inhibition in Apoe^−/− mice reduced atherosclerosis and FLNA-CD36 colocalization. Using macrophage-specific FAK kinase-dead (KD) Apoe^−/− mice, ORO staining of carotid arteries showed that FAK-KD mice had reduced atherosclerotic lesions compared to FAK wild type (WT) mice upon partial carotid ligation. We also verified increased FAK activation and FLNA-CD36 colocalization in human atherosclerotic tissues.
Conclusions
These data demonstrate that oxLDL stimulation of macrophages triggers the formation of a FAK-FLNA-CD36 complex, essential for oxLDL uptake and CD36 internalization. Disrupting this complex-either through FAK inhibition or FLNA knockdown-reduces foam cell formation. Thus, targeting the FAK-FLNA-CD36 axis may provide a potential new therapeutic strategy for treating or preventing macrophage foam cells in atherosclerosis.