CD163+ macrophages promote angiogenesis and vascular permeability accompanied by inflammation in atherosclerosis
Liang Guo, Hirokuni Akahori, Emanuel Harari, Samantha L. Smith, Rohini Polavarapu, Vinit Karmali, Fumiyuki Otsuka, Rachel L. Gannon, Ryan E. Braumann, Megan H. Dickinson, Anuj Gupta, Audrey L. Jenkins, Michael J. Lipinski, Johoon Kim, Peter Chhour, Paul S. de Vries, Hiroyuki Jinnouchi, Robert Kutys, Hiroyoshi Mori, Matthew D. Kutyna, Sho Torii, Atsushi Sakamoto, Cheol Ung Choi, Qi Cheng, Megan L. Grove, Mariem A. Sawan, Yin Zhang, Yihai Cao, Frank D. Kolodgie, David P. Cormode, Dan E. Arking, Eric Boerwinkle, Alanna C. Morrison, Jeanette Erdmann, Nona Sotoodehnia, Renu Virmani, Aloke V. Finn
Liang Guo, Hirokuni Akahori, Emanuel Harari, Samantha L. Smith, Rohini Polavarapu, Vinit Karmali, Fumiyuki Otsuka, Rachel L. Gannon, Ryan E. Braumann, Megan H. Dickinson, Anuj Gupta, Audrey L. Jenkins, Michael J. Lipinski, Johoon Kim, Peter Chhour, Paul S. de Vries, Hiroyuki Jinnouchi, Robert Kutys, Hiroyoshi Mori, Matthew D. Kutyna, Sho Torii, Atsushi Sakamoto, Cheol Ung Choi, Qi Cheng, Megan L. Grove, Mariem A. Sawan, Yin Zhang, Yihai Cao, Frank D. Kolodgie, David P. Cormode, Dan E. Arking, Eric Boerwinkle, Alanna C. Morrison, Jeanette Erdmann, Nona Sotoodehnia, Renu Virmani, Aloke V. Finn
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Research Article Angiogenesis Vascular biology

CD163+ macrophages promote angiogenesis and vascular permeability accompanied by inflammation in atherosclerosis

Abstract

Intake of hemoglobin by the hemoglobin-haptoglobin receptor CD163 leads to a distinct alternative non–foam cell antiinflammatory macrophage phenotype that was previously considered atheroprotective. Here, we reveal an unexpected but important pathogenic role for these macrophages in atherosclerosis. Using human atherosclerotic samples, cultured cells, and a mouse model of advanced atherosclerosis, we investigated the role of intraplaque hemorrhage on macrophage function with respect to angiogenesis, vascular permeability, inflammation, and plaque progression. In human atherosclerotic lesions, CD163+ macrophages were associated with plaque progression, microvascularity, and a high level of HIF1α and VEGF-A expression. We observed irregular vascular endothelial cadherin in intraplaque microvessels surrounded by CD163+ macrophages. Within these cells, activation of HIF1α via inhibition of prolyl hydroxylases promoted VEGF-mediated increases in intraplaque angiogenesis, vascular permeability, and inflammatory cell recruitment. CD163+ macrophages increased intraplaque endothelial VCAM expression and plaque inflammation. Subjects with homozygous minor alleles of the SNP rs7136716 had elevated microvessel density, increased expression of CD163 in ruptured coronary plaques, and a higher risk of myocardial infarction and coronary heart disease in population cohorts. Thus, our findings highlight a nonlipid-driven mechanism by which alternative macrophages promote plaque angiogenesis, leakiness, inflammation, and progression via the CD163/HIF1α/VEGF-A pathway.

Authors

Liang Guo, Hirokuni Akahori, Emanuel Harari, Samantha L. Smith, Rohini Polavarapu, Vinit Karmali, Fumiyuki Otsuka, Rachel L. Gannon, Ryan E. Braumann, Megan H. Dickinson, Anuj Gupta, Audrey L. Jenkins, Michael J. Lipinski, Johoon Kim, Peter Chhour, Paul S. de Vries, Hiroyuki Jinnouchi, Robert Kutys, Hiroyoshi Mori, Matthew D. Kutyna, Sho Torii, Atsushi Sakamoto, Cheol Ung Choi, Qi Cheng, Megan L. Grove, Mariem A. Sawan, Yin Zhang, Yihai Cao, Frank D. Kolodgie, David P. Cormode, Dan E. Arking, Eric Boerwinkle, Alanna C. Morrison, Jeanette Erdmann, Nona Sotoodehnia, Renu Virmani, Aloke V. Finn

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Figure 5

Deletion of CD163 in mice reduces intraplaque neovascularization and plaque progression.

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Deletion of CD163 in mice reduces intraplaque neovascularization and pla...
(A) Immunoblotting of mouse macrophages with quantitation of densitometry for HIF1α in macrophages isolated from WT or CD163–/– mice stimulated with Hb (n = 4 per group). (B) Analysis of macrophage supernatant for VEGF-A by ELISA (n = 4 per group). (1: WT macrophages, 2: Hb-stimulated WT macrophages, 3: CD163–/– macrophages, 4: Hb-stimulated CD163–/– macrophages). (C) Representative H&E staining of BCA plaque with a of gross inset photograph of the aortic arch from 1-year-old ApoE–/– and ApoE–/– CD163–/– mice. Scale bars: 100 μm. (D–G) Quantitative measurements of lesion size, percentage of stenosis, lesion pathological scores, and necrotic areas in the BCA plaque (n = 8–12 per group). (H) Representative immunofluorescence confocal microscopic images of BCA plaque stained with VE-cadherin (green), CD163 (red), and DAPI (blue). Scale bars: 50 μm and 10 μm (enlarged images of boxed areas on the left). (I) Microvessel density quantification calculated by the number of microvessels per plaque area identified by VE-cadherin immunofluorescence confocal microscopy (n = 6–7 per group). (J) Representative immunofluorescence confocal microscopic images of intraplaque FITC-dextran (green) as a marker for permeability. Scale bars: 100 μm. Total intraplaque FITC fluorescence was quantified from confocal images of BCA plaques perfused with FITC-dextran to determine permeability (n = 7 per group). Bars and plots indicate the mean ± SEM (A and B) or the mean ± SD (D–G, I, and J). (A and B) *P < 0.05 versus other groups, by 1-way ANOVA , and, if the ratio test (F test) was significant, a more detailed post-hoc analysis of differences between groups was performed using a Tukey-Kramer honest significant difference test. (D–G, I, and J) P < 0.01 and P < 0.05, by 2-sided Student’s t test.