IL-17 and Th17 cells have quickly become an important paradigm in immunology and the IL-23-IL-17 axis has emerged as a critical regulatory system that bridges the innate and adaptive arms of the immune system and have been linked to the pathogenesis of several chronic inflammatory diseases (39
). The role of Th17 cells and IL-17 in various stages of atherogenesis remains controversial and is only beginning to be elucidated. While IL-17 is predominantly a proinflammatoryy cytokine, it has pleotrophic and regulatory functions as well and has been implicated both as an instigator in the pathogenesis of inflammatory disorders as well as protective in certain inflammatory disease models (17
). Here we show that genetic deficiency of IL-17A reduces the size of atherosclerotic plaques in the aortic sinus and the aorta en face preparations in C57BL/6 mice under high fat diet. Furthermore, IL-17A deficiency was associated with significant reductions in lipid composition of the plaques, the number of infiltrating macrophages as well as expression of the proinflammatory enzyme, COX-2 and T cell infiltration in the aortic sinus lesions. The atheroprotective effect of IL-17A deficiency was not due to altered serum cholesterol or lipoproteins. We observed significantly reduced circulating proinflammatory cytokines IL-12p40, CCL2 and IFNγ in IL-17A−/− mice, suggesting that the atheroprotective effects of IL-17A deficiency may result in part from reduced systemic inflammation. Collectively, our data obtained from IL-17-deficient mice indicate that IL-17A is proatherogenic in the hypercholesterolemic mouse model, and that Th17 cells and IL-17A contribute to atherogenesis by both systemic and local effects.
Consistent with previous studies (25
), we also found that C. pneumoniae
infection of hypercholesterolemic C57BL/6 mice (WT) resulted in accelerated atherosclerosis associated with significantly increased lesion size, lipid content, numbers of macrophage and COX-2 in lesions, and serum levels of proinflammatory cytokines (IL-12p40, CCL2) compared with mock-infected controls. In contrast, while C. pneumoniae
infection was still able to accelerate atherosclerosis in IL-17A−/−, the infection-mediated acceleration of lesion size and lipid composition of the aorta as well as serum levels of IL-12p40 and CCL2 increases were significantly inhibited in IL-17A-deficient mice compared to infected WT mice, suggesting that C. pneumoniae
infection-induced acceleration in lesion development in hypercholesterolemic mice, is at least partially driven by an IL-17A-dependent manner. However, the data also suggest that pathogen-induced acceleration can still occur in an IL-17A-independent manner. Importantly, since we did not observe any influence of IL-17A gene deficiency on bacterial replication and clearance in the lungs during Cpn infection (data not shown), we believe that the residual infection-induced acceleration of atherosclerosis in IL-17A-deficient mice is not due to a significantly altered course of infection.
In this study we used a well-described and widely-used high-fat diet that contains cholate (Paigen diet) (44
) that has been shown to induce atherosclerotic lesions in C57BL/6 mice by several investigators (45
), While there have been some concerns that cholate containing high-fat diets may be associated with induction of fibrosis related genes in the liver in addition to inflammation genes induced by non-cholate containing high-fat diets (30
), we did not observed any liver fibrosis in our groups and lipoprotein levels were similar in both WT and IL-17-deficient mice that received the same diet. The hypercholesterolemia induced by this atherogenic diet in C57BL/C mice is milder than in genetically altered mice such as ApoE-deficient mice (48
). Therefore, this model seems to be a useful model for studying the accelerating role of C. pneumoniae
in atherosclerosis at an earlier stage.
Our data showing that IL-17A is proatherogenic, using IL-17A-deficient mice, is in agreement with several previous studies (21
). Kuiper van Es et al observed that irradiated LDLR-deficient recipient mice transplanted with IL-17RA-deficient bone marrow resulted in a 46% reduction in lesion size in the aortic root plaque under a Western-type diet (21
). However, one limitation to this model is that stromal cells are known to be important responders to IL-17A (49
), thus this model should be considered only as a partial phenotype. Erbel et al. administered blocking antbody agianst IL-17A that resulted in reduced atherosclerotic lesion development and decreased plaque vulnerability, cellular infiltration, and tissue activation in ApoE-deficient mice (22
). One limitation of this study is that blocking IL-17A will not prevent the formation of Th17 cells, as well as other IL-17 producing cell types, which may also have effects aside from IL-17A production. Additionally, as with all blocking strategies, the prospect of leaky signaling, and or cross reactivity of the antibody cannot be ignored. Another recent study by Smith et al found that IL-17A treatment of whole aorta isolated from ApoE−/− mice promots aortic CXCL1 expression and monocyte adhesion in an ex- vivo adhesion assay(23
), also supporting a proatherogenic role for IL-17A. However, in contrast to our results and these earlier studies, Taleb et al published that loss of suppressor of cytokine signaling-3 (SOCS-3) in mouse T cells increases IL-17A production, inducing an anti-inflammatory macrophage phenotype, which results in a reduction in lesion development and vascular inflammation, suggesting that IL-17 may have a protective role in atherogenesis (24
). Additionally, these investigators have also shown that in vivo administration of rIL-17A to LDLR−/− mice resulted in reduced endothelial VCAM-1 expression, as well as reduced vascular T cell infiltration and atherosclerotic lesion development (24
). These investigators concluded that endogenous expression of SOCS3 in T cells interrupts a major regulatory pathway in atherosclerosis through inhibition of IL-17A production and that IL-17A functions as an atheroprotective cytokine. However, one caveat to this study is the fact that in addition to upregulated IL-17A, the authors also found an increase in IL-10 production, a potent regulatory cytokine, which is atheroprotective (51
). Additionally, SOCS-3 plays a role in several different signaling cascades, thus it is difficult to assign cause and effect for the role of IL-17A in the SOCS-3 knockout model (56
). In summary while the precise role of IL-17 in various stages of atherogenesis remains controversial, our study together with recently published studies (21
) now provide a more direct evidence that IL-17A maybe predominantly proatherogenic. Nevertheless, IL-17 is a pleiotropic cytokine with environment-specific inflammatory or protective and regulatory functions(17
). Therefore, the role of IL-17 in various stages of lesion development are probably complex and needs to be further evaluated.
Many studies have documented the importance of Th1 responses and atherogenesis (57
). We have gone over in some detail the current literature on this subject and reviewed the current controversy on the role of Th17 cells and atherosclerosis(63
). It has become increasingly clear that Th17 and Th1 cells regulate each other, apparently in both directions (16
). Th1 cells have been found to be necessary for the downstream accumulation of Th17 cells (67
). In a recent study published in Immunity, Lin Y et al found that IL-17A is required for induction of IL-12, a critical cytokine in Th1 skewing, and host resistance to infection (16
). If IL-17 was required for proper Th1 responses in atherogenesis, then one would predict that IL-17A−/− mice would have reduced atherosclerotic lesions, as we have seen in our study. We also found that IL-17A could induce proinflammatory cytokine production (CCL2, IL-6) in primary aortic endothelial cells, and that these supernatants could then drive foam cell formation and contribute to atherogenesis. Taken together with the previous study that showed IL-17A could induce monocyte adhesion and CXCL1 expression on aortic endothelial cells, one could argue that IL-17A most likely also has a direct effect on lesion development (23
Adding to the complexity in assessing our study and the other investigations for the role of IL-17 is the fact that IL-17 is a family of cytokines, including various isoforms. Our study utilized the IL-17A knockout mouse, but presumably IL-17F would be unaffected in this mouse model. Additionally, IL-17E (IL-25), an anti-inflammatory cytokine, was found to be expressed in normal and atherosclerotic vessels, and might play a role in regulating inflammatory processes in the vessel wall (68
). Therefore further work investigations should be done to define the specific role of the different IL-17 isoforms (such as IL-17F or IL-17E) in atherosclerosis.