We report the first systems biology investigation of regulators controlling arterial plaque macrophage transcriptional changes in response to lipid lowering in vivo in two distinct mouse models of atherosclerosis regression. Transcriptome measurements from plaque macrophages from the Reversa mouse were integrated with measurements from an aortic transplant-based mouse model of plaque regression. Functional relevance of the genes detected as differentially expressed in plaque macrophages in response to lipid lowering in vivo was assessed through analysis of gene functional annotations, overlap with in vitro foam cell studies, and overlap of associated eQTLs with human atherosclerosis/CAD risk SNPs. To identify transcription factors that control plaque macrophage responses to lipid lowering in vivo , we used an integrative strategy – leveraging macrophage epigenomic measurements – to detect enrichment of transcription factor binding sites upstream of genes that are differentially expressed in plaque macrophages during regression. The integrated analysis uncovered eight transcription factor binding site elements that were statistically overrepresented within the 5′ regulatory regions of genes that were upregulated in plaque macrophages in the Reversa model under maximal regression conditions and within the 5′ regulatory regions of genes that were upregulated in the aortic transplant model during regression. Of these, the TCF/LEF binding site was present in promoters of upregulated genes related to cell motility, suggesting that the canonical Wnt signaling pathway may be activated in plaque macrophages during regression. We validated this network-based prediction by demonstrating that β-catenin expression is higher in regressing (vs. control group) plaques in both regression models, and we further demonstrated that stimulation of canonical Wnt signaling increases macrophage migration in vitro . These results suggest involvement of canonical Wnt signaling in macrophage emigration from the plaque during lipid lowering-induced regression, and they illustrate the discovery potential of an epigenome-guided, systems approach to understanding atherosclerosis regression. Author Summary Atherosclerosis, a progressive accumulation of lipid-rich plaque within arteries, is an inflammatory disease in which the response of macrophages (a key cell type of the innate immune system) to plasma lipoproteins plays a central role. In humans, the goal of significantly reducing already-established plaque through drug treatments, including statins, remains elusive. In mice, atherosclerosis can be reversed by experimental manipulations that lower circulating lipid levels. A common feature of many regression models is that macrophages transition to a less inflammatory state and emigrate from the plaque. While the molecular regulators that control these responses are largely unknown, we hypothesized that by integrating global measurements of macrophage gene expression in regressing plaques with measurements of the macrophage chromatin landscape, we could identify key molecules that control macrophage responses to the lowering of circulating lipid levels. Our systems biology analysis of plaque macrophages yielded a network in which the Wnt signaling pathway emerged as a candidate upstream regulator. Wnt signaling is known to affect both inflammation and the ability of macrophages to migrate from one location to another, and our targeted validation studies provide evidence that Wnt signaling is increased in plaque macrophages during regression. Our findings both demonstrate the power of a systems approach to uncover candidate regulators of regression and to identify a potential new therapeutic target.
【초록키워드】 Transcriptome, Inflammation, Macrophage, Gene Expression, macrophages, Lipoproteins, eQTL, SNPs, risk, Systems biology, in vitro, binding site, innate immune system, Migration, mice, humans, Regression model, response, Atherosclerosis, network, molecular, in vivo, mouse model, expression, statins, Stimulation, Evidence, power, Analysis, Regression, inflammatory state, therapeutic target, transcription factor, transcription factors, drug treatments, cell type, cell motility, control group, annotations, overlap, enrichment, promoters, validation study, circulating, lowering, regression models, inflammatory disease, plasma lipoproteins, arteries, macrophage responses, plaque, plaque macrophage responses, plaque macrophages, plaque regression, plaques, regressing plaques, upregulated genes, Wnt signaling, Wnt signaling pathway, β-catenin, chromatin, element, promoter, upstream, regulatory region, while, upregulated gene, Affect, Macrophage response, approach, Cell, identify, detect, significantly, eight, functional, activated, condition, increase, less, reducing, demonstrated, upregulated, differentially expressed, reversed, canonical, statistically, transcriptional change, arterial plaque macrophage, epigenomic, foam cell, plaque macrophage, plaque macrophage response, plasma lipoprotein, regressing plaque, TCF/LEF, 【제목키워드】 Transcriptome, macrophages, Atherosclerosis, Wnt,