Impact of Gut Microbiota and Diet on the Development of Atherosclerosis in Apoe-/- Mice.

Objective- To investigate the effect of gut microbiota and diet on atherogenesis. Approach and Results- Here, we investigated the interaction between the gut microbiota and diet on atherosclerosis by feeding germ-free or conventionally raised Apoe-/- mice chow or Western diet alone or supplemented with choline (which is metabolized by the gut microbiota and host enzymes to trimethylamine N-oxide) for 12 weeks. We observed smaller aortic lesions and lower plasma cholesterol levels in conventionally raised mice compared with germ-free mice on a chow diet; these differences were not observed in mice on a Western diet. Choline supplementation increased plasma trimethylamine N-oxide levels in conventionally raised mice but not in germ-free mice. However, this treatment did not affect the size of aortic lesions or plasma cholesterol levels. Gut microbiota composition was analyzed by sequencing of 16S rRNA genes. As expected, the global community structure and relative abundance of many taxa differed between mice fed chow or a Western diet. Choline supplementation had minor effects on the community structure although the relative abundance of some taxa belonging to Clostridiales was altered. Conclusions- In conclusion, the impact of the gut microbiota on atherosclerosis is dietary dependent and is associated with plasma cholesterol levels. Furthermore, the microbiota was required for trimethylamine N-oxide production from dietary choline, but this process could not be linked to increased atherosclerosis in this model.

Bacterial profile in human atherosclerotic plaques.

BACKGROUND AND AIMS: Several studies have confirmed the presence of bacterial DNA in atherosclerotic plaques, but its contribution to plaque stability and vulnerability is unclear. In this study, we investigated whether the bacterial plaque-profile differed between patients that were asymptomatic or symptomatic and whether there were local differences in the microbial composition within the plaque. METHODS: Plaques were removed by endarterectomy from asymptomatic and symptomatic patients and divided into three different regions known to show different histological vulnerability: A, upstream of the maximum stenosis; B, site for maximum stenosis; C, downstream of the maximum stenosis. Bacterial DNA composition in the plaques was determined by performing 454 pyrosequencing of the 16S rRNA genes, and total bacterial load was determined by qPCR. RESULTS: We confirmed the presence of bacterial DNA in the atherosclerotic plaque by qPCR analysis of the 16S rRNA gene but observed no difference (n.s.) in the amount between either asymptomatic and symptomatic patients or different plaque regions A, B and C. Unweighted UniFrac distance metric analysis revealed no distinct clustering of samples by patient group or plaque region. Operational taxonomic units (OTUs) from 5 different phyla were identified, with the majority of the OTUs belonging to Proteobacteria (48.3%) and Actinobacteria (40.2%). There was no difference between asymptomatic and symptomatic patients, or plaque regions, when analyzing the origin of DNA at phylum, family or OTU level (n.s.). CONCLUSIONS: There were no major differences in bacterial DNA amount or microbial composition between plaques from asymptomatic and symptomatic patients or between different plaque regions, suggesting that other factors are more important in determining plaque vulnerability.

Melanocortin 1 receptor agonist protects podocytes through catalase and RhoA activation.

Drugs containing adrenocorticotropic hormone have been used as therapy for patients with nephrotic syndrome. We have previously shown that adrenocorticotropic hormone and a selective agonist for the melanocortin 1 receptor (MC1R) exert beneficial actions in experimental membranous nephropathy with reduced proteinuria, reduced oxidative stress, and improved glomerular morphology and function. Our hypothesis is that MC1R activation in podocytes elicits beneficial effects by promoting stress fibers and maintaining podocyte viability. To test the hypothesis, we cultured podocytes and used highly specific agonists for MC1R. Podocytes were subjected to the nephrotic-inducing agent puromycin aminonucleoside, and downstream effects of MC1R activation on podocyte survival, antioxidant defense, and cytoskeleton dynamics were studied. To increase the response and enhance intracellular signals, podocytes were transduced to overexpress MC1R. We showed that puromycin promotes MC1R expression in podocytes and that activation of MC1R promotes an increase of catalase activity and reduces oxidative stress, which results in the dephosphorylation of p190RhoGAP and formation of stress fibers through RhoA. In addition, MC1R agonists protect against apoptosis. Together, these mechanisms protect the podocyte against puromycin. Our findings strongly support the hypothesis that selective MC1R-activating agonists protect podocytes and may therefore be useful to treat patients with nephrotic syndromes commonly considered as podocytopathies.

Effects of melanocortin 1 receptor agonists in experimental nephropathies.

Nephrotic syndrome, characterized by massive proteinuria, is caused by a large group of diseases including membranous nephropathy (MN) and focal segmental glomerulosclerosis (FSGS). Although the underlying mechanisms are beginning to unravel, therapy is unspecific and far from efficient. It has been suggested that adrenocorticotropic hormone (ACTH) has beneficial effects in patients with MN and possibly in other nephrotic diseases. We have previously reported that ACTH may act directly on podocytes through the melanocortin 1 receptor (MC1R). In the present study, we evaluate the effect of highly specific MC1R agonists in two different nephrotic disease models. Experimental MN: Passive Heymann nephritis (PHN) was induced in rats that were treated for four weeks with MS05, a selective MC1R agonist, or saline. The degree of albuminuria was significantly reduced over time and the effect was sustained one week after treatment withdrawal (p<0.05). Experimental FSGS: Based on a dose-response study, two doses of adriamycin were used for induction of nephropathy in Balb/c mice. Mice were treated with either a synthetic MC1R agonist (BMS-470539), with α-melanocyte stimulating hormone (α-MSH) or with saline. There was no beneficial effect of treatment. In summary, MC1R agonists reduce albuminuria and improve morphology in experimentally induced MN whereas they have no effect in experimental FSGS. The results illustrate the differences in these podocytopathies in terms of signaling mechanisms underlying proteinuria, and progression of disease.