Chronic skin inflammation accelerates macrophage cholesterol crystal formation and atherosclerosis
Yvonne Baumer, Qimin Ng, Gregory E. Sanda, Amit K. Dey, Heather L. Teague, Alexander V. Sorokin, Pradeep K. Dagur, Joanna I. Silverman, Charlotte L. Harrington, Justin A. Rodante, Shawn M. Rose, Nevin J. Varghese, Agastya D. Belur, Aditya Goyal, Joel M. Gelfand, Danielle A. Springer, Christopher K.E. Bleck, Crystal L. Thomas, Zu-Xi Yu, Mårten C.G. Winge, Howard S. Kruth, M. Peter Marinkovich, Aditya A. Joshi, Martin P. Playford, Nehal N. Mehta
Yvonne Baumer, Qimin Ng, Gregory E. Sanda, Amit K. Dey, Heather L. Teague, Alexander V. Sorokin, Pradeep K. Dagur, Joanna I. Silverman, Charlotte L. Harrington, Justin A. Rodante, Shawn M. Rose, Nevin J. Varghese, Agastya D. Belur, Aditya Goyal, Joel M. Gelfand, Danielle A. Springer, Christopher K.E. Bleck, Crystal L. Thomas, Zu-Xi Yu, Mårten C.G. Winge, Howard S. Kruth, M. Peter Marinkovich, Aditya A. Joshi, Martin P. Playford, Nehal N. Mehta
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Research Article Cardiology Inflammation

Chronic skin inflammation accelerates macrophage cholesterol crystal formation and atherosclerosis

Abstract

Inflammation is critical to atherogenesis. Psoriasis is a chronic inflammatory skin disease that accelerates atherosclerosis in humans and provides a compelling model to understand potential pathways linking these diseases. A murine model capturing the vascular and metabolic diseases in psoriasis would accelerate our understanding and provide a platform to test emerging therapies. We aimed to characterize a new murine model of skin inflammation (Rac1V12) from a cardiovascular standpoint to identify novel atherosclerotic signaling pathways modulated in chronic skin inflammation. The RacV12 psoriasis mouse resembled the human disease state, including presence of systemic inflammation, dyslipidemia, and cardiometabolic dysfunction. Psoriasis macrophages had a proatherosclerotic phenotype with increased lipid uptake and foam cell formation, and also showed a 6-fold increase in cholesterol crystal formation. We generated a triple-genetic K14-RacV12–/+/Srb1–/–/ApoER61H/H mouse and confirmed psoriasis accelerates atherogenesis (~7-fold increase). Finally, we noted a 60% reduction in superoxide dismutase 2 (SOD2) expression in human psoriasis macrophages. When SOD2 activity was restored in macrophages, their proatherogenic phenotype reversed. We demonstrate that the K14-RacV12 murine model captures the cardiometabolic dysfunction and accelerates vascular disease observed in chronic inflammation and that skin inflammation induces a proatherosclerotic macrophage phenotype with impaired SOD2 function, which associated with accelerated atherogenesis.

Authors

Yvonne Baumer, Qimin Ng, Gregory E. Sanda, Amit K. Dey, Heather L. Teague, Alexander V. Sorokin, Pradeep K. Dagur, Joanna I. Silverman, Charlotte L. Harrington, Justin A. Rodante, Shawn M. Rose, Nevin J. Varghese, Agastya D. Belur, Aditya Goyal, Joel M. Gelfand, Danielle A. Springer, Christopher K.E. Bleck, Crystal L. Thomas, Zu-Xi Yu, Mårten C.G. Winge, Howard S. Kruth, M. Peter Marinkovich, Aditya A. Joshi, Martin P. Playford, Nehal N. Mehta

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Figure 2

Psoriatic mouse macrophages display proatherosclerotic characteristics.

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Psoriatic mouse macrophages display proatherosclerotic characteristics. ...
(A) BMDM were analyzed for the presence of Rac1 and the attached Myc tag. No differences in expression of Rac1 or Myc were detected. (B) Equal amounts of BM cells were plated for differentiation. Cell count after differentiation was determined. Resulting cell counts are increased in psoriatic mice, possibly due to differences in adhesion and proliferation. (C) Gene expression of macrophage polarization was determined using qPCR and is altered in psoriatic BMDMs as compared with LMC BMDMs. (D and E) Scanning electron microscopy analysis of BMDM phenotype with subsequent quantification of cells presenting a round phenotype (n = 6, each with 5 fields imaged and quantified). (F) Phenotypically, BMDMs were characterized using scanning electron microscopy and immunofluorescence analysis for CD206 (red) and CD68 (green) (nuclei: DAPI, blue) (representative images of n = 3). (G and H) Modified lipoprotein uptake by BMDM was determined after treatment with 20 μg/ml DiI-labeled AcLDL or OxLDL for 2h and 4h of incubation. Five images per condition were taken at equal exposure and MFI of 100 individual cells per image analyzed using ImageJ. (I) Differences in modified LDL receptors between LMC and K14-Rac1V12–/+ BMDM were detected using qPCR. (J) Macrophage foam cell formation upon 24h 50μg/ml AcLDL or OxLDL treatment was detected after subsequent Bodipy493/503 labeling (green, nuclei: DAPI, blue) and ImageJ analysis as described in E and F. (K and L) Presence of cholesterol crystals was visualized using polarized light microscopy of mouse (K) and human (L) macrophages, displaying increased cholesterol crystal presence under baseline and lipid treated conditions (50 μg/ml) (BMDM n = 6/6, HMDM n = 5/10). (Data are expressed as mean ± SEM; 2-way ANOVA with Bonferroni correction [H and J] with P < 0.007; duplicates for H and J; Mann-Whitney test [B, C, E, and I] with P < 0.05; n ≥ 6/6, *P < 0.05, **P < 0.005, ***P < 0.0005 indicates significance to LMC control; ^P < 0.05, ^^P < 0.005, ^^^P < 0.0005 indicates significance between indicated groups; scale bars: 50 μm [F and G],100 μm [J]) (LMC, littermate control; AcLDL, acetylated low-density lipoprotein; OxLDL, oxidized LDL)