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The gut microbiota were shown to play critical roles in the development of atherosclerosis, but the detailed mechanism is limited. The purpose of this study is to clarify the influence of gut microbiota on atherogenesis via lipid metabolism and systemic inflammation. Germ-free or conventionally raised (Conv) ApoE-deficient (ApoE−/−) mice were fed chow diet and euthanized at 20 weeks of age. We found that the lack of gut microbiota in ApoE−/−mice caused a significant increase in the plasma and hepatic cholesterol levels compared with Conv ApoE−/−mice. The absence of gut microbiota changed the bile acid composition in the ileum, which was associated with activation of the enterohepatic fibroblast growth factor 15, fibroblast growth factor receptor 4 axis, and reduction of cholesterol 7α-hydroxylase and hepatic bile acid synthesis, resulting in the accumulation of liver cholesterol content. However, we found that the lack of microbiota caused a significant reduction in atherosclerotic lesion formation compared with Conv ApoE−/−mice, which might be associated with the attenuation of lipopolysaccharide-mediated inflammatory responses. Our findings indicated that the gut microbiota affected both hypercholesterolemia and atherogenesis in mice.

Supplemental Digital Content is available in the text.

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Introduction:Expansion and rupture of abdominal aortic aneurysms (AAA) are clinically associated with high morbidity and mortality. Cigarrete smoke (CS) is the strongest risk factor, yet direct evidence that CS causes AAA is missing. Our aim is to uncover a previously unknown capacity for CS induced-atherosclerosis to directly injure the aorta causing development of AAA.Methods and Results:High cholesterol diet- fed Apoe-/- mice revealed the development of saccular lesions and enlargement of the abdominal aortae of animals exposed to CS twice daily for 5 days per week, but not room air. Lesion incidence increased with duration of cigarette smoke exposure, appearing as early as 4 weeks in the supra-renal abdominal aorta (4% at 4 week, 15% at 8 week and 29% at 12 week; AAA Incidence). Aortic wall rupture resulted in death in 11% of CS exposed mice. Histological examination revealed that aortic wall degenerations were associated with overlying atherosclerotic (AS) plaques (96%) and that the elastic lamina was frequently disrupted in CS exposed mice, even at 4 week time point. BrdU pulse chase and flow cytometry analyses showed more monocytes were recruited in CS exposed aortae. RNA-sequencing of 5159 single cells showed 2 monocyte and 8 macrophage populations were clustered according to considerable differences in gene expression and revealed that hypercholesterolemia determined monocyte and macrophage diversity. Anti colony stimulating factor-1 (?CSF-1) antibody treatment decreased macrophage accumulation and plaque development and led to less damage of the elastic lamina in abdominal aorta, suggesting that macrophages play a critical role in AS and AAA (60% isotype, 19% ?CSF-1, p<0.05; histological sections with elastin fragmentation).Conclusions:We show that interaction of bona fide risk factors of AAA such as CS and hypercholesterolemia led to degeneration of the aortic wall, rupture and death.This exploration of mechanisms unleashed by known risk factors for AAA augments understanding of the mechanisms whereby atherosclerotic plaque components can directly injure the aorta and identifies recruitment of mononuclear phagocytes as a therapeutic target in preventing degeneration of the arterial wall.

Introduction:Increasing evidence has suggested that a strong correlation exists between the gut microbiota and development of coronary artery disease (CAD). However, how specific gut bacteria contribute to CAD and how we can manipulate their abundance remains to be determined. Here, we aimed to identify the gut bacterial species involved in CAD and determine whether resistant starch (RS) could prevent CAD by acting on gut microbiota.Methods:We recruited 30 CAD patients and 30 non-CAD controls with coronary risk factors and compared the gut microbiota composition in fecal samples to identify bacterial species with differential abundance between the groups. We then used mouse models of atherosclerosis to study the mechanisms underlying the relationship between these differentially expressed bacterial species and atherosclerosis. Finally, we investigated whether RS could manipulate the gut microbiota composition using a single-batch fermentation system.Results:A significantly lower abundance of Bacteroides vulgatusand B. doreiwas observed in CAD patients (9.5% [range, 3.8?16.5%] vs. 3.7% [range, 1.9?11.1%]; p < 0.05). Gavage with B. vulgatusand B. doreiattenuated atherosclerotic lesion formation in atherosclerosis-prone mice, and markedly ameliorated endotoxemia. These mice also showed decreased lipopolysaccharide (LPS) production by the gut microbiota, which effectively suppressed pro-inflammatory immune responses. Furthermore, significantly higher gut microbial LPS production was observed in CAD patients (1.8 ? 105[range, 1.0?3.6 ? 105] endotoxin unit (EU)/g vs. 3.4 ? 105[range, 1.9?6.6 ? 105] EU/g; p < 0.05). Fecal LPS levels in CAD patients were significantly and negatively correlated with the abundance of two species of Bacteroides(r = -0.29; p < 0.05). Finally, RS significantly increased the absolute number of B. vulgatusand B. doreiamong gut bacteria in the fermentation system.Conclusions:We identified two species of Bacteroidesthat may serve as novel, inexpensive therapeutic targets to prevent CAD. RS enhanced their growth in the human gut microbiota, suggesting that this treatment could be used in CAD patients to restore the gut microbiota composition to symbiotic state.

Introduction:Aortic aneurysms are characterized by macrophage mediated inflammation. Lyve-1+resident aortic macrophages independently self-renew, and are functionally distinct from bone marrow derived macrophages that are recruited during inflammation.Hypothesis:We hypothesized that Lyve-1+and Lyve-1-macrophages differentially contribute to aortic aneurysm/dissection.Methods:The contribution of Lyve-1 expression to aortic and cardiac macrophage populations was characterized using conditional depletion with Lyve-1cre/+Csf1rfl/flmice. Abdominal aortic aneurysms (AAA) were induced in C57BL6/J mice by administering ?-Aminopropionitrile monofumarate in drinking water and osmotic infusion of angiotensin II. In order to assess the contribution of macrophages, we depleted monocytes and macrophages by infusion of anti- Colony Stimulating Factor 1 Receptor (CSF1R) neutralizing antibody. Lyve-1cre/+Csf1rfl/flmice and parabiosis was used to determine the contribution of resident and monocyte-derived macrophage populations, respectively. Aortas were analyzed using histology and flow cytometry.Results:Lyve-1cre/+Csf1rfl/flmice had selective depletion of aortic and cardiac tissue resident macrophages (P<0.01). AAA is associated with increased CD11B+F4/80+macrophages (P<0.0001), and disease severity correlated with relative depletion of Lyve-1+macrophages (P<0.05). Parabionts with aneurysm had increased chimerism in aortic macrophages compared to na?ve, suggesting contribution of circulating cells. Whilst anti-CSF1R neutralization protected against AAA, conditional depletion of Lyve-1 macrophages in Lyve-1cre/+Csf1rfl/fldid not.Conclusions:These data suggest that Lyve-1-bone marrow-derived aortic macrophages drive abdominal aortic aneurysm/dissection.

Mechanisms that govern transcriptional regulation of inflammation in atherosclerosis remain largely unknown. Here, we identify the nuclear transcription factor c-Myb as an important mediator of atherosclerotic disease in mice. Atherosclerosis-prone animals fed a diet high in cholesterol exhibit increased levels of c-Mybin the bone marrow. Use of mice that either harbor a c-Mybhypomorphic allele or where c-Mybhas been preferentially deleted in B cell lineages revealed that c-Myb potentiates atherosclerosis directly through its effects on B lymphocytes. Reduced c-Myb activity prevents the expansion of atherogenic B2 cells yet associates with increased numbers of IgM-producing antibody-secreting cells (IgM-ASCs) and elevated levels of atheroprotective oxidized low-density lipoprotein (OxLDL)-specific IgM antibodies. Transcriptional profiling revealed that c-Myb has a limited effect on B cell function but is integral in maintaining B cell progenitor populations in the bone marrow. Thus, targeted disruption of c-Myb beneficially modulates the complex biology of B cells in cardiovascular disease.