MARK4 (Microtubule Affinity-Regulating Kinase 4)-Dependent Inflammasome Activation Promotes Atherosclerosis-Brief Report.

OBJECTIVE: MARK4 (microtubule affinity-regulating kinase 4) regulates NLRP3 (nucleotide-binding oligomerization domain, leucine-rich repeat, and pyrin domain containing 3) inflammasome activation. The aim of the study is to examine the role of MARK4 in hematopoietic cells during atherosclerosis. METHODS AND RESULTS: We show increased MARK4 expression in human atherosclerotic lesions compared with adjacent areas. MARK4 is coexpressed with NLRP3, and they colocalize in areas enriched in CD68-positive but α-SMA (α-smooth muscle actin)-negative cells. Expression of MARK4 and NLRP3 in the atherosclerotic lesions is associated with the production of active IL (interleukin)-1ß and IL-18. To directly assess the role of hematopoietic MARK4 in NLRP3 inflammasome activation and atherosclerotic plaque formation, Ldlr (low-density lipoprotein receptor)-deficient mice were lethally irradiated and reconstituted with either wild-type or Mark4-deficient bone marrow cells, and were subsequently fed a high-fat diet and cholesterol diet for 9 weeks. Mark4 deficiency in bone marrow cells led to a significant reduction of lesion size, together with decreased circulating levels of IL-18 and IFN-γ (interferon-γ). Furthermore, Mark4 deficiency in primary murine bone marrow-derived macrophages prevented cholesterol crystal-induced NLRP3 inflammasome activation, as revealed by reduced caspase-1 activity together with reduced production of IL-1ß and IL-18. CONCLUSIONS: MARK4-dependent NLRP3 inflammasome activation in the hematopoietic cells regulates the development of atherosclerosis.

X-Box Binding Protein-1 Dependent Plasma Cell Responses Limit the Development of Atherosclerosis.

RATIONALE: Diverse B cell responses and functions may be involved in atherosclerosis. Protective antibody responses, such as those against oxidized lipid epitopes, are thought to mainly derive from T cell-independent innate B cell subsets. In contrast, both pathogenic and protective roles have been associated with T cell-dependent antibodies, and their importance in both humans and mouse models is still unclear. OBJECTIVE: To specifically target antibody production by plasma cells and determine the impact on atherosclerotic plaque development in mice with and without CD4+ T cells. METHODS AND RESULTS: We combined a model of specific antibody deficiency, B cell-specific CD79a-Cre x XBP1 (X-box binding protein-1) floxed mice (XBP1-conditional knockout), with antibody-mediated depletion of CD4+ T cells. Ldlr knockout mice transplanted with XBP1-conditional knockout (or wild-type control littermate) bone marrow were fed western diet for 8 weeks with or without anti-CD4 depletion. All groups had similar levels of serum cholesterol. In Ldlr/XBP1-conditional knockout mice, serum levels of IgG, IgE, and IgM were significantly attenuated, and local antibody deposition in atherosclerotic plaque was absent. Antibody deficiency significantly accelerated atherosclerosis at both the aortic root and aortic arch. T cell and monocyte responses were not modulated, but necrotic core size was greater, even when adjusting for plaque size, and collagen deposition significantly lower. Anti-CD4 depletion in Ldlr/wild-type mice led to a decrease of serum IgG1 and IgG2c but not IgG3, as well as decreased IgM, associated with increased atherosclerosis and necrotic cores, and a decrease in plaque collagen. The combination of antibody deficiency and anti-CD4 depletion has no additive effects on aortic root atherosclerosis. CONCLUSIONS: The endogenous T cell-dependent humoral response can be protective. This has important implications for novel vaccine strategies for atherosclerosis and in understanding the impacts of immunotherapies used in patients at high risk for cardiovascular disease.

Type-2 innate lymphoid cells control the development of atherosclerosis in mice.

Type-2 innate lymphoid cells (ILC2) are a prominent source of type II cytokines and are found constitutively at mucosal surfaces and in visceral adipose tissue. Despite their role in limiting obesity, how ILC2s respond to high fat feeding is poorly understood, and their direct influence on the development of atherosclerosis has not been explored. Here, we show that ILC2 are present in para-aortic adipose tissue and lymph nodes and display an inflammatory-like phenotype atypical of adipose resident ILC2. High fat feeding alters both the number of ILC2 and their type II cytokine production. Selective genetic ablation of ILC2 in Ldlr-/- mice accelerates the development of atherosclerosis, which is prevented by reconstitution with wild type but not Il5-/- or Il13-/- ILC2. We conclude that ILC2 represent a major innate cell source of IL-5 and IL-13 required for mounting atheroprotective immunity, which can be altered by high fat diet.

Regulatory B cell-specific interleukin-10 is dispensable for atherosclerosis development in mice.

OBJECTIVE: To determine the role of regulatory B cell-derived interleukin (IL)-10 in atherosclerosis. APPROACH AND RESULTS: We created chimeric Ldlr(-/-) mice with a B cell-specific deficiency in IL-10, and confirmed that purified B cells stimulated with lipopolysaccharide failed to produce IL-10 compared with control Ldlr(-/-) chimeras. Mice lacking B-cell IL-10 demonstrated enhanced splenic B-cell numbers but no major differences in B-cell subsets, T cell or monocyte distribution, and unchanged body weights or serum cholesterol levels compared with control mice. After 8 weeks on high-fat diet, there were no differences in aortic root or aortic arch atherosclerosis. In addition to plaque size, plaque composition (macrophages, T cells, smooth muscle cells, and collagen) was similar between groups. CONCLUSIONS: In contrast to its prominent regulatory role in many immune-mediated diseases and its proposed modulatory role in atherosclerosis, B cell-derived IL-10 does not alter atherosclerosis in mice.

MHC Class II-restricted antigen presentation by plasmacytoid dendritic cells drives proatherogenic T cell immunity.

BACKGROUND: Plasmacytoid dendritic cells (pDCs) bridge innate and adaptive immune responses and are important regulators of immuno-inflammatory diseases. However, their role in atherosclerosis remains elusive. METHODS AND RESULTS: Here, we used genetic approaches to investigate the role of pDCs in atherosclerosis. Selective pDC deficiency in vivo was achieved using CD11c-Cre × Tcf4(-/flox) bone marrow transplanted into Ldlr(-/-) mice. Compared with control Ldlr(-/-) chimeric mice, CD11c-Cre × Tcf4(-/flox) mice had reduced atherosclerosis levels. To begin to understand the mechanisms by which pDCs regulate atherosclerosis, we studied chimeric Ldlr(-/-) mice with selective MHCII deficiency on pDCs. Significantly, these mice also developed reduced atherosclerosis compared with controls without reductions in pDC numbers or changes in conventional DCs. MHCII-deficient pDCs showed defective stimulation of apolipoprotein B100-specific CD4(+) T cells in response to native low-density lipoprotein, whereas production of interferon-α was not affected. Finally, the atheroprotective effect of selective MHCII deficiency in pDCs was associated with significant reductions of proatherogenic T cell-derived interferon-γ and lesional T cell infiltration, and was abrogated in CD4(+) T cell-depleted animals. CONCLUSIONS: This study supports a proatherogenic role for pDCs in murine atherosclerosis and identifies a critical role for MHCII-restricted antigen presentation by pDCs in driving proatherogenic T cell immunity.