Lymphocytic tumor necrosis factor receptor superfamily co-stimulatory molecules in the pathogenesis of atherosclerosis

Purpose of reviewThe role of lymphocytes in the chronic inflammatory disease atherosclerosis has emerged over the past decade. Co-stimulatory molecules of the heterogeneous tumor necrosis factor receptor superfamily play a pivotal role in lymphocyte activation, proliferation and differentiation. Here we describe the immune modulatory properties and mechanisms of four tumor necrosis factor receptor superfamily members in atherosclerosis.Recent findingsCD40/CD40L, OX40L/OX40, CD70/CD27 and CD137/CD137L are present in human atherosclerotic plaques and have shown strong immune modulatory functions in atherosclerosis, resulting in either atherogenic or atheroprotective effects in mouse models of atherosclerosis.SummaryInsight into the immune modulatory mechanisms of co-stimulatory interactions in atherosclerosis can contribute to clinical exploitation of these interactions in the treatment of cardiovascular disease

Lymphocytic tumor necrosis factor receptor superfamily co-stimulatory molecules in the pathogenesis of atherosclerosis

Purpose of reviewThe role of lymphocytes in the chronic inflammatory disease atherosclerosis has emerged over the past decade. Co-stimulatory molecules of the heterogeneous tumor necrosis factor receptor superfamily play a pivotal role in lymphocyte activation, proliferation and differentiation. Here we describe the immune modulatory properties and mechanisms of four tumor necrosis factor receptor superfamily members in atherosclerosis.Recent findingsCD40/CD40L, OX40L/OX40, CD70/CD27 and CD137/CD137L are present in human atherosclerotic plaques and have shown strong immune modulatory functions in atherosclerosis, resulting in either atherogenic or atheroprotective effects in mouse models of atherosclerosis.SummaryInsight into the immune modulatory mechanisms of co-stimulatory interactions in atherosclerosis can contribute to clinical exploitation of these interactions in the treatment of cardiovascular disease

Targeting macrophage Histone deacetylase 3 stabilizes atherosclerotic lesions

Macrophages are key immune cells found in atherosclerotic plaques and critically shape atherosclerotic disease development. Targeting the functional repertoire of macrophages may hold novel approaches for future atherosclerosis management. Here, we describe a previously unrecognized role of the epigenomic enzyme Histone deacetylase 3 (Hdac3) in regulating the atherosclerotic phenotype of macrophages. Using conditional knockout mice, we found that myeloid Hdac3 deficiency promotes collagen deposition in atherosclerotic lesions and thus induces a stable plaque phenotype. Also, macrophages presented a switch to anti-inflammatory wound healing characteristics and showed improved lipid handling. The pro-fibrotic phenotype was directly linked to epigenetic regulation of the Tgfb1 locus upon Hdac3 deletion, driving smooth muscle cells to increased collagen production. Moreover, in humans, HDAC3 was the sole Hdac upregulated in ruptured atherosclerotic lesions, Hdac3 associated with inflammatory macrophages, and HDAC3 expression inversely correlated with pro-fibrotic TGFB1 expression. Collectively, we show that targeting the macrophage epigenome can improve atherosclerosis outcome and we identify Hdac3 as a potential novel therapeutic target in cardiovascular disease

The role of dendritic cells and their chemokine TARC in the pathogenesis of atherosclerosis

In this study, the DC-derived chemokine CCL17 and its role in the pathogenesis of atherosclerosis was investigated. By using a ‘knock-in’ mice expressing a targeted replacement of the Ccl17 gene by eGFP, we observed that CCL17+ DCs belong to a mature subpopulation of cDCs related to the myeloid lineage. Microarray profiling showed no further difference in gene regulation or even intrinsic defects of EGFP+ Ccl17E/E compared with EGFP+ Ccl17E/+ BMDCs. Additionally, normal DC functions such as phagocytosis or migration were not altered in Ccl17 knock-out DCs. In vivo, a network of CD11c+ DCs could be detected in the intima of the vessel wall of naïve wt mice, but these cells did not express CCL17, in contrast to a prominent expression of MHC-II. However, during plaque development CCL17+ DCs accumulate within inflamed atherosclerotic lesions in Apoe-/- mice. In addition, CCL17+ DCs were detected in lesions of bone marrow transplanted mice, indicating that these DCs are recruited from the bone marrow to the site of inflammation. Furthermore, we demonstrated that CCL17-deficiency reduces atherosclerotic plaque formation with a decreased macrophage content in the plaque, while the content of SMCs was increased, indicating a more stable plaque phenotype compared to the phenotype observed in Ccl17+/+ Apoe-/- mice. In line with the findings that CCL17 attracts T cells, we observed that CCL17+ DCs attract CD4+ T cells in vitro and in vivo. In addition, increased numbers of CD3+ T cells were detected in atherosclerotic plaques of Ccl17+/+ Apoe-/- compared with Ccl17E/E Apoe-/- mice. Furthermore, CCL17 seems to be necessary for efficient T cell activation, as higher IL-2 levels were observed in the supernatants of T cells cocultured with CCL17+ BMDCs. Moreover IFN-gamma levels were significantly higher in those cultures, indicating that T cells were primed and polarized in a Th1 specific manner. The difference in cytokine profile was also observed in vivo. In LNs of Ccl17+/+ Apoe-/- mice, Ifn-gamma and Il-17 transcription levels were up-regulated, while the typical Th2 cytokines Il-4 and Il-10 were down-regulated in comparison to Ccl17E/E Apoe-/- mice. Finally, adoptive transfer experiments demonstrated that T cells isolated from Ccl17E/E Apoe-/- mice and injected into Apoe-/- recipient mice act atheroprotective. Lesion development in these animals was significantly reduced when compared with Apoe-/- mice receiving T cells from Ccl17+/+ Apoe-/-. These data open the possibility of achieving therapeutic regression and stabilization of advanced atherosclerotic lesions by targeting CCL17

The role of dendritic cells and their chemokine TARC in the pathogenesis of atherosclerosis

In this study, the DC-derived chemokine CCL17 and its role in the pathogenesis of atherosclerosis was investigated. By using a ‘knock-in’ mice expressing a targeted replacement of the Ccl17 gene by eGFP, we observed that CCL17+ DCs belong to a mature subpopulation of cDCs related to the myeloid lineage. Microarray profiling showed no further difference in gene regulation or even intrinsic defects of EGFP+ Ccl17E/E compared with EGFP+ Ccl17E/+ BMDCs. Additionally, normal DC functions such as phagocytosis or migration were not altered in Ccl17 knock-out DCs. In vivo, a network of CD11c+ DCs could be detected in the intima of the vessel wall of naïve wt mice, but these cells did not express CCL17, in contrast to a prominent expression of MHC-II. However, during plaque development CCL17+ DCs accumulate within inflamed atherosclerotic lesions in Apoe-/- mice. In addition, CCL17+ DCs were detected in lesions of bone marrow transplanted mice, indicating that these DCs are recruited from the bone marrow to the site of inflammation. Furthermore, we demonstrated that CCL17-deficiency reduces atherosclerotic plaque formation with a decreased macrophage content in the plaque, while the content of SMCs was increased, indicating a more stable plaque phenotype compared to the phenotype observed in Ccl17+/+ Apoe-/- mice. In line with the findings that CCL17 attracts T cells, we observed that CCL17+ DCs attract CD4+ T cells in vitro and in vivo. In addition, increased numbers of CD3+ T cells were detected in atherosclerotic plaques of Ccl17+/+ Apoe-/- compared with Ccl17E/E Apoe-/- mice. Furthermore, CCL17 seems to be necessary for efficient T cell activation, as higher IL-2 levels were observed in the supernatants of T cells cocultured with CCL17+ BMDCs. Moreover IFN-gamma levels were significantly higher in those cultures, indicating that T cells were primed and polarized in a Th1 specific manner. The difference in cytokine profile was also observed in vivo. In LNs of Ccl17+/+ Apoe-/- mice, Ifn-gamma and Il-17 transcription levels were up-regulated, while the typical Th2 cytokines Il-4 and Il-10 were down-regulated in comparison to Ccl17E/E Apoe-/- mice. Finally, adoptive transfer experiments demonstrated that T cells isolated from Ccl17E/E Apoe-/- mice and injected into Apoe-/- recipient mice act atheroprotective. Lesion development in these animals was significantly reduced when compared with Apoe-/- mice receiving T cells from Ccl17+/+ Apoe-/-. These data open the possibility of achieving therapeutic regression and stabilization of advanced atherosclerotic lesions by targeting CCL17

MicroRNA 302a is a novel modulator of cholesterol homeostasis and atherosclerosis

Macrophage foam cell formation is a key feature of atherosclerosis. Recent studies have shown that specific microRNAs (miRs) are regulated in modified low-density lipoprotein-treated macrophages, which can affect the cellular cholesterol homeostasis. Undertaking a genome-wide screen of miRs regulated in primary macrophages by modified low-density lipoprotein, miR-302a emerged as a potential candidate that may play a key role in macrophage cholesterol homeostasis. The objective of this study was to assess the involvement of miR-302a in macrophage lipid homeostasis and if it can influence circulating lipid levels and atherosclerotic development when it is inhibited in a murine atherosclerosis model. We found that transfection of primary macrophages with either miR-302a or anti-miR-302a regulated the expression of ATP-binding cassette (ABC) transporter ABCA1 mRNA and protein. Luciferase reporter assays showed that miR-302a repressed the 3' untranslated regions (UTR) activity of mouse Abca1 by 48% and human ABCA1 by 45%. In addition, transfection of murine macrophages with miR-302a attenuated cholesterol efflux to apolipoprotein A-1 (apoA-1) by 38%. Long-term in vivo administration of anti-miR-302a to mice with low-density lipoprotein receptor deficiency (Ldlr(-/-)) fed an atherogenic diet led to an increase in ABCA1 in the liver and aorta as well as an increase in circulating plasma high-density lipoprotein levels by 35% compared with that of control mice. The anti-miR-302a-treated mice also displayed reduced atherosclerotic plaque size by ≈25% and a more stable plaque morphology with reduced signs of inflammation. These studies identify miR-302a as a novel modulator of cholesterol efflux and a potential therapeutic target for suppressing atherosclerosi

Constitutive GITR Activation Reduces Atherosclerosis by Promoting Regulatory CD4+ T-Cell Responses-Brief Report

Glucocorticoid-induced tumor necrosis factor receptor family-related protein (GITR) is expressed on CD4(+) effector memory T cells and regulatory T cells; however, its role on these functionally opposing cell types in atherosclerosis is not fully understood. Low-density lipoprotein receptor-deficient mice (Ldlr(-/-)) were lethally irradiated and reconstituted with either bone marrow from B-cell-restricted Gitrl transgenic mice or from wild-type controls and fed a high-cholesterol diet for 11 weeks. Chimeric Ldlr(-/-) Gitrl(tg) mice showed a profound increase in both CD4(+) effector memory T cells and regulatory T cells in secondary lymphoid organs. Additionally, the number of regulatory T cells was significantly enhanced in the thymus and aorta of these mice along with increased Gitrl and Il-2 transcript levels. Atherosclerotic lesions of Ldlr(-/-) Gitrl(tg) chimeras contained more total CD3(+) T cells as well as Foxp3(+) regulatory T cells overall, leading to significantly less severe atherosclerosis. These data indicate that continuous GITR stimulation through B cell Gitrl acts protective in a mouse model of atherosclerosis by regulating the balance between regulatory and effector memory CD4(+) T cell

Hypercholesterolemia-induced priming of hematopoietic stem and progenitor cells aggravates atherosclerosis

Modulation of hematopoietic stem and progenitor cells (HSPCs) determines immune cell function. In this study, we investigated how hypercholesterolemia affects HSPC biology and atherosclerosis. Hypercholesterolemia induced loss of HSPC quiescence, characterized by increased proliferation and expression of cyclin B1, C1, and D1, and a decreased expression of Rb, resulting in a 3.6- fold increase in the number of HSPCs in hypercholesterolemic Ldlr(-/-) mice. Competitive bone marrow (BM) transplantations showed that a hypercholesterolemic BM microenvironment activates HSPCs and skews their development toward myeloid lineages. Conversely, hypercholesterolemia-primed HSPCs acquired an enhanced propensity to generate myeloid cells, especially granulocytes and Ly6C(high) monocytes, even in a normocholesterolemic BM microenvironment. In conformity, macrophages differentiated from hypercholesterolemia-primed HSPCs produced 17.0% more TNF-α, 21.3% more IL-6, and 10.5% more MCP1 than did their normocholesterolemic counterparts. Hypercholesterolemia-induced priming of HSPCs generated leukocytes that more readily migrated into the artery, which resulted in a 2.1-fold increase in atherosclerotic plaque size. In addition, these plaques had a more advanced phenotype and exhibited a 1.2-fold increase in macrophages and 1.8-fold increase in granulocytes. These results identify hypercholesterolemia-induced activation and priming of HSPCs as a novel pathway in the development of atherosclerosis. Inhibition of this proinflammatory differentiation pathway on the HSPC level has the potential to reduce atherosclerosi