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The phosphoinositide 3-kinase signaling pathway is involved in the control of modified low-density lipoprotein uptake by human macrophages.

ABSTRACT: The transformation of macrophages into lipid-loaded foam cells is a critical early event in the pathogenesis of atherosclerosis. Both receptor-mediated uptake of modified LDL, mediated primarily by scavenger receptors-A (SR-A) and CD36 along with other proteins such as lipoprotein lipase (LPL), and macropinocytosis contribute to macrophage foam cell formation. The signaling pathways that are involved in the control of foam cell formation are not fully understood. In this study, we have investigated the role of phosphoinositide 3-kinase (PI3K) in relation to foam cell formation in human macrophages. The pan PI3K inhibitor LY294002 attenuated the uptake of modified LDL and macropinocytosis, as measured by Lucifer Yellow uptake, by human macrophages. In addition, the expression of SR-A, CD36 and LPL was attenuated by LY294002. The use of isoform-selective PI3K inhibitors showed that PI3K-β, -γ and -δ were all required for the expression of SR-A and CD36 whereas only PI3K-γ was necessary in the case of LPL. These studies reveal a pivotal role of PI3K in the control of macrophage foam cell formation and provide further evidence for their potential as therapeutic target against atherosclerosis.

SUBMITTER: Michael DR

PROVIDER: S-EPMC4339697 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND:Macrophage accumulation in the vascular wall is a hallmark of atherosclerosis. Studies showed that shifting of oxidized lipids-induced inflammatory macrophages towards an anti-inflammatory phenotype by promoting oxidative metabolism attenuated atherosclerosis progression. Therefore, this study aimed to investigate whether metformin, which has ameliorated atherosclerosis in animal models and clinical trials, modulated oxidized low-density lipoprotein (Ox-LDL) induced inflammatory status in macrophages by regulating cellular oxidative metabolism. METHODS:Murine raw264.7 macrophages were incubated with Ox-LDL (50 μg/mL) in the presence or absence of metformin (15 μmol/L) for 24 h. Real-time polymerase chain reaction was used to quantify the transcription of classically activated (M1) pro-inflammatory and alternatively activated (M2) anti-inflammatory markers and mitochondrial DNA copy numbers. Cellular reactive oxygen species (ROS) production and mitochondrial membrane potential were detected by immunofluorescence. Cellular adenosine triphosphate (ATP) synthesis, glucose uptake, and lactic acid production were measured by commercial kit and normalized to cellular lysates. Western blotting analysis was performed to detect the expression of mitochondrial fusion/fission related proteins, enzymes mediating lipid metabolism and signaling pathway of glucose transport. Differences between groups were analyzed using one-way analysis of variance. RESULTS:Metformin improved Ox-LDL-impaired anti-inflammatory phenotype in raw264.7 macrophages as shown by up-regulated transcription of anti-inflammatory markers including interleukin 10 (0.76 ± 0.04 vs. 0.94 ± 0.01, P = 0.003) and Resistin-like molecule alpha (0.67 ± 0.08 vs. 1.78 ± 0.34, P = 0.030). Conversely, Ox-LDL-diminished phosphorylation of Akt was up-regulated by metformin treatment (0.47 ± 0.05 vs. 1.02 ± 0.08, P = 0.040), associated with an improvement of mitochondrial function, characterized by decreased ROS generation (2.50 ± 0.07 vs. 2.15 ± 0.04, P = 0.040), increased lipid oxidation, and elevated cellular ATP production (0.026 ± 0.001 vs. 0.035 ± 0.003, P = 0.020). Moreover, metformin-mediated Akt activation increased Akt substrate of 160 kDa (AS160) phosphorylation (0.51 ± 0.04 vs. 1.03 ± 0.03, P = 0.0041), promoted membrane translocation of glucose transporter 1, and increased glucose influx into the cells (4.78 ± 0.04 vs. 5.47 ± 0.01, P < 0.001). CONCLUSION:This study suggested that targeting macrophage metabolism with new or existing drugs had therapeutic potential for the prevention and treatment of diabetes-accelerated atherosclerosis.

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The phosphoinositide 3-kinase signaling pathway is involved in the control of modified low-density lipoprotein uptake by human macrophages.

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