Objective-Degradation of extracellular matrix plays an important role in growth and destabilization of atherosclerotic plaques.Cystatin C, inhibitor of the collagen-and elastin-degrading cysteine proteases of the cathepsin family, is produced by virtually all cell types. It is present in the normal artery wall but severely reduced in human atherosclerotic lesions. Methods and Results-To determine the functional role of cystatin C in atherosclerosis, we crossed cystatin C-deficient (cysC Ϫ/Ϫ ) mice with apolipoprotein E-deficient (apoE Ϫ/Ϫ ) mice. After 25 weeks of atherogenic diet, mice lacking apoE and cystatin C (cysC Ϫ/Ϫ apoE Ϫ/Ϫ ) had larger subvalvular plaques compared with cysC ϩ/ϩ apoE Ϫ/Ϫ mice (766 000Ϯ20 000 m 2 per section versus 662 000Ϯ19 000 m 2 per section; Pϭ0.001), suggesting an atheroprotective role of cystatin C. The plaques from cysC Ϫ/Ϫ apoE Ϫ/Ϫ mice were characterized by increased total macrophage content. To determine which cellular source is important for the antiatherosclerotic effect of cystatin C, we performed bone marrow transplantations. ApoE Ϫ/Ϫ mice were transplanted with either cysC Ϫ/Ϫ apoE Ϫ/Ϫ or cysC ϩ/ϩ apoE Ϫ/Ϫ bone marrow. No significant differences in plaque area, macrophage, collagen, or lipid content of subvalvular lesions between the 2 groups were detected. Conclusions-The result suggests that the protective role of cystatin C in atherosclerosis is dependent primarily on its expression in nonhematopoietic cell types.

ObjectiveIL-25 has been implicated in the initiation of type 2 immunity and in the protection against autoimmune inflammatory diseases. Recent studies have identified the novel innate lymphoid type 2 cells (ILC2s) as an IL-25 target cell population. The purpose of this study was to evaluate if IL-25 has any influence on atherosclerosis development in mice.Methods and ResultsAdministration of 1 μg IL-25 per day for one week to atherosclerosis-prone apolipoprotein (apo)E deficient mice, had limited effect on the frequency of T cell populations, but resulted in a large expansion of ILC2s in the spleen. The expansion was accompanied by increased levels of anti-phosphorylcholine (PC) natural IgM antibodies in plasma and elevated levels of IL-5 in plasma and spleen. Transfer of ILC2s to apoE deficient mice elevated the natural antibody-producing B1a cell population in the spleen. Treatment of apoE/Rag-1 deficient mice with IL-25 was also associated with extensive expansion of splenic ILC2s and increased plasma IL-5, suggesting ILC2s to be the source of IL-5. Administration of IL-25 in IL-5 deficient mice resulted in an expanded ILC2 population, but did not stimulate generation of anti-PC IgM, indicating that IL-5 is not required for ILC2 expansion but for the downstream production of natural antibodies. Additionally, administration of 1 μg IL-25 per day for 4 weeks in apoE deficient mice reduced atherosclerosis in the aorta both during initiation and progression of the disease.ConclusionsThe present findings demonstrate that IL-25 has a protective role in atherosclerosis mediated by innate responses, including ILC2 expansion, increased IL-5 secretion, B1a expansion and natural anti-PC IgM generation, rather than adaptive Th2 responses.

Objective of the StudyDiabetic patients have a much more widespread and aggressive form of atherosclerosis and therefore, higher risk for myocardial infarction, peripheral vascular disease and stroke, but the molecular mechanisms leading to accelerated damage are still unclear. Recently, we showed that hyperglycemia activates the transcription factor NFAT in the arterial wall, inducing the expression of the pro-atherosclerotic protein osteopontin. Here we investigate whether NFAT activation may be a link between diabetes and atherogenesis.Methodology and Principal FindingsStreptozotocin (STZ)-induced diabetes in apolipoprotein E−/− mice resulted in 2.2 fold increased aortic atherosclerosis and enhanced pro-inflammatory burden, as evidenced by elevated blood monocytes, endothelial activation- and inflammatory markers in aorta, and pro-inflammatory cytokines in plasma. In vivo treatment with the NFAT blocker A-285222 for 4 weeks completely inhibited the diabetes-induced aggravation of atherosclerosis, having no effect in non-diabetic mice. STZ-treated mice exhibited hyperglycemia and higher plasma cholesterol and triglycerides, but these were unaffected by A-285222. NFAT-dependent transcriptional activity was examined in aorta, spleen, thymus, brain, heart, liver and kidney, but only augmented in the aorta of diabetic mice. A-285222 completely blocked this diabetes-driven NFAT activation, but had no impact on the other organs or on splenocyte proliferation or cytokine secretion, ruling out systemic immunosuppression as the mechanism behind reduced atherosclerosis. Instead, NFAT inhibition effectively reduced IL-6, osteopontin, monocyte chemotactic protein 1, intercellular adhesion molecule 1, CD68 and tissue factor expression in the arterial wall and lowered plasma IL-6 in diabetic mice.ConclusionsTargeting NFAT signaling may be a novel and attractive approach for the treatment of diabetic macrovascular complications.