Journal: Arteriosclerosis, thrombosis, and vascular biology

Article Title: Smooth Muscle Cells Contribute the Majority of Foam Cells in Apolipoprotein E-Deficient Mouse Atherosclerosis

doi: 10.1161/ATVBAHA.119.312434

Figure Lengend Snippet: (A) ABCA1 expression of CD45+ and CD45− foam cells from ApoE−/− mice fed a WD for 6 weeks (left panel) or 27-week-old male ApoE−/− mice fed a chow diet (right panel) determined as the median fluorescence of ABCA1 intensity by flow cytometry. For WD-fed mice, **P < 0.01 using two-way ANOVA, n=9 mice/group; there was no statistical difference between the two sexes. For chow-fed mice, **P < 0.01 using Mann–Whitney U test, n=5 mice/group. (B) Fold change of ABCA1 mRNA level in RAW 264.7 macrophages and MASMCs treated with 100 g/ml agLDL as compared to fatty acid-free albumin alone for 24 h. *P < 0.05, using Welch’s t test. (C) ABCA1 protein levels and cholesterol efflux from RAW 264.7 macrophages and MASMCs treated with 100 μg/mL agLDL for 48 hrs followed by incubation with 0.3 mM 8Br-cAMP ± 10 μg/mL (for efflux) for 24h. Cholesterol efflux was determined by LC-MS and normalized by total cell protein levels *P < 0.05, **P < 0.01 using two-tailed Mann–Whitney U test, N=5.

Article Snippet: BV421 rat anti-mouse CD45, V500 rat anti-mouse I-A/I-E (BD Pharmingen), and Alexa Fluor647 rat anti-mouse ABCA1 (AbD Serotec) antibodies were use according to manufacturer’s instructions.

Techniques: Expressing, Fluorescence, Flow Cytometry, MANN-WHITNEY, Incubation, Liquid Chromatography with Mass Spectroscopy, Two Tailed Test

Journal: Arteriosclerosis, thrombosis, and vascular biology

Article Title: Smooth Muscle Cells Contribute the Majority of Foam Cells in Apolipoprotein E-Deficient Mouse Atherosclerosis

doi: 10.1161/ATVBAHA.119.312434

Figure Lengend Snippet: In humans, a thick diffuse intimal thickening (DIT) layer of SMCs is present in atherosclerosis-prone arteries beginning from birth8. At the onset of atherosclerosis lipoproteins that diffuse into the artery wall are trapped through a charge-charge interaction with SMC-secreted proteoglycans, primarily in the deep intima7. These trapped lipoproteins are modified and aggregated, becoming a substrate for uptake by the surrounding SMCs. At this stage monocyte/macrophages are located primarily in the subendothelial region away from the deposited lipids9. Over time both SMCs and macrophages take up modified lipoproteins to become foam cells independently and/or interactively. ABCA1 (red dots), the rate-limiting cholesterol efflux promoter, is robustly upregulated in macrophage foam cells, but less so in SMC foam cells. Increased ABCA1 expression facilitates cholesterol removal out of macrophage-derived foam cells, while impaired ABCA1 expression reduces cholesterol efflux from SMC foam cells, resulting in SMCs being the predominant lineage among total foam cells. In ApoE−/− mice, no DIT SMC layer is present. Atherogenic lipoproteins induce endothelial injury and monocyte binding and infiltration into the intima, where they mature into macrophages that take up the modified intimal lipids. Cytokines released by injured endothelial cells and monocytes/macrophages, including PDGF, TNF-α, and IL-1β induce SMC migration from the media into the intima. SMCs proliferate in the intima and take up modified lipoproteins to become foam cells. Similar to human lesions, the SMC foam cells exhibit reduced ABCA1 expression compared to macrophages. Over time the total contribution to foam cells shifts from macrophages to SMCs, due at least in part to impaired ABCA1 expression by the SMC foam cells.

Article Snippet: BV421 rat anti-mouse CD45, V500 rat anti-mouse I-A/I-E (BD Pharmingen), and Alexa Fluor647 rat anti-mouse ABCA1 (AbD Serotec) antibodies were use according to manufacturer’s instructions.

Techniques: Modification, Expressing, Derivative Assay, Binding Assay, Migration