Abstract Body (Do not enter title and authors here): Background: Inflammation underpins atherosclerosis (AS) and its life-threaten complications caused by the rupture ofvulnerable plaques. AS is characterized by excessive accumulation of foam cells and deficient efferocytosis in the lesions, both of which contribute significantly to plaque vulnerability. EGLN3 is emerging as an important inflammatory regulator. However, virtually nothing is known about the role and mechanism for EGLN3 in modulating plaque stability.
Objective: This study aimed to determine the role and mechanism of EGLN3 in regulating foam cell formation, efferocytosis, atherosclerosis initiation and plaque stability.
Methods: Bone marrow-derived macrophages prepared from EGLN3^+/+ and EGLN3^-/- mice were used for foam cell formation and efferocytosis assays. ApoE^-/- and EGLN3^-/-;ApoE^-/- mice were placed on a high-fat diet to induce atherosclerosis. Lipid uptake and cholesterol efflux were detected by cell fluorescence and NBD-cholesterol. In vitro and in vivo efferocytosis assays were carried out to analyze efferocytosis. Oil red O staining was conducted to detect foam cell formation and the size of aortic plaque. HE and Sirius red staining were performed to estimate the size of the necrotic core and the thickness of the fibrous cap. Gene expression in macrophages and the lesions were determined by qPCR, immunoblotting and immunostaining. The content of inflammatory mediators was measured by ELISA.
Results: EGLN3 deficiency promotes macrophage foam cell formation. EGLN3 depletion increases CD36 expression and lipid uptake, but impedes ABCA1 expression and lipid efflux. EGLN3 deletion impairs macrophage efferocytosis in vitro and in vivo. EGLN3 elimination blunts the expression of efferocytotic receptor MerTK in cultured and lesional macrophages. EGLN3 ablation in ApoE^-/- mice increases atherosclerotic plaque formation, lesional lipid deposit, and the plaque vulnerability. EGLN3 deficiency in ApoE^-/- mice enhances lesional macrophage content, the expression of inflammatory mediators in the lesions, and the systemic inflammatory response.
Conclusion: EGLN3 restrains macrophage foam cell formation, potentiates macrophage efferocytosis, and strengths plaque stability. This study provides novel mechanistic insights into the athero-protective and plaque-stabilizing roles of EGLN3, which will help develop new strategy to combat atherosclerotic cardiovascular disease.
Keywords: EGLN3, foam cell, efferocytosis, atherosclerosis, plaque stability.