Lack of ApoA-I in ApoEKO Mice Causes Skin Xanthomas, Worsening of Inflammation, and Increased Coronary Atherosclerosis in the Absence of Hyperlipidemia.

BACKGROUND: HDL (high-density lipoprotein) and its major protein component, apoA-I (apolipoprotein A-I), play a unique role in cholesterol homeostasis and immunity. ApoA-I deficiency in hyperlipidemic, atheroprone mice was shown to drive cholesterol accumulation and inflammatory cell activation/proliferation. The present study was aimed at investigating the impact of apoA-I deficiency on lipid deposition and local/systemic inflammation in normolipidemic conditions. METHODS: ApoE deficient mice, apoE/apoA-I double deficient (DKO) mice, DKO mice overexpressing human apoA-I, and C57Bl/6J control mice were fed normal laboratory diet until 30 weeks of age. Plasma lipids were quantified, atherosclerosis development at the aortic sinus and coronary arteries was measured, skin ultrastructure was evaluated by electron microscopy. Blood and lymphoid organs were characterized through histological, immunocytofluorimetric, and whole transcriptome analyses. RESULTS: DKO were characterized by almost complete HDL deficiency and by plasma total cholesterol levels comparable to control mice. Only DKO showed xanthoma formation and severe inflammation in the skin-draining lymph nodes, whose transcriptome analysis revealed a dramatic impairment in energy metabolism and fatty acid oxidation pathways. An increased presence of CD4+ T effector memory cells was detected in blood, spleen, and skin-draining lymph nodes of DKO. A worsening of atherosclerosis at the aortic sinus and coronary arteries was also observed in DKO versus apoE deficient. Human apoA-I overexpression in the DKO background was able to rescue the skin phenotype and halt atherosclerosis development. CONCLUSIONS: HDL deficiency, in the absence of hyperlipidemia, is associated with severe alterations of skin morphology, aortic and coronary atherosclerosis, local and systemic inflammation.

The transcriptome profiling of diseased mouse aortas discloses a dysregulation of the sympathetic neurotransmission in atherosclerosis.

Previous reports suggest an association between the development of atherosclerosis and alterations in the aortic sympathetic nervous system, but there is no agreement on whether atherosclerotic plaques are accompanied by increased or decreased sympathetic innervation in the arterial wall. In the present study, the aortic transcriptional profile of mice with different predisposition to atherosclerosis was investigated to clarify how the expression of genes involved in sympathetic neurotransmission varied. Eight-week-old C57Bl/6J control mice, Apoe knockout mice (EKO), EKO mice overexpressing human apoA-I (EKO/hA-I) and double Apoe/Apoa1 knockout mice (DKO) mice were fed either a standard rodent diet or a Western-type diet for 22 weeks. Atherosclerosis was quantified, and the aortic transcriptome was analyzed by RNAseq. Western-type diet administration deeply modified the aortic transcriptome. In the genetically modified atherosclerosis-prone mouse lines, an upregulated expression of genes associated with the immunomodulatory response was observed, paralleled by a downregulated expression of the genes related to sympathetic nervous system. Functional enrichment analysis indicated that the presence of advanced atherosclerosis was accompanied by reduced neuronal generation, modulation of synapse chemical transmission, and catecholamine biosynthesis, supporting a relationship between atherosclerosis, dyslipidemia, and sympathetic neurotransmission.

Effect of Diets on Plasma and Aorta Lipidome: A Study in the apoE Knockout Mouse Model.

SCOPE: Specific lipid molecules circulating in plasma at low concentrations have emerged as biomarkers of atherosclerotic risk. The aim of the present study is that of evaluating, in an athero-prone mouse model, how different diets can affect plasma and aorta lipidome. METHODS AND RESULTS: Thirty-six apoE knockout mice are divided in three groups and feed 12 weeks with diets differing for cholesterol and fatty acid content. Atherosclerosis is measured at the aortic sinus and aorta. Lipids are quantified in plasma and aorta with mass spectrometry. The cholesterol content of the diets is the main driver of lipid accumulation in plasma and aorta. The fatty acid composition of the diets affects plasma levels both of essential (linoleic acid) and nonessential (myristic and arachidonic acid) ones. Lipidomics show a comparable distribution, in plasma and aorta, of the main lipid components of oxidized LDL, including cholesteryl esters and lysophosphatidylcholines. Interestingly, lactosylceramide, glucosyl/galactosylceramide, and individual ceramide species are found to accumulate in diseased aortic segments. CONCLUSION: Both the cholesterol and fatty acid content of the diets profoundly affect plasma lipidome. Aorta lipidome is likewise affected with the accumulation of specific lipids known as markers of atherosclerosis.

Effect of diet and genotype on the miRNome of mice with altered lipoprotein metabolism.

The molecular mechanism by which lipid/lipoprotein biosynthesis is regulated in mammals involves a very large number of genes that are subject to multiple levels of regulation. miRNAs are recognized contributors to lipid homeostasis at the post-transcriptional level, although the elucidation of their role is made difficult by the multiplicity of their targets and the ability of more miRNAs to affect the same mRNAs. In this study, an evaluation of how miRNA expression varies in organs playing a key role in lipid/lipoprotein metabolism was conducted in control mice and in two mouse models carrying genetic ablations which differently affect low-density lipoprotein metabolism. Mice were fed a lipid-poor standard diet and a diet enriched in cholesterol and saturated fat. The results obtained showed that there are no miRNAs whose expression constantly vary with dietary or genetic changes. Furthermore, it appears that diet, more than genotype, impacts on organ-specific miRNA expression profiles.

reString: an open-source Python software to perform automatic functional enrichment retrieval, results aggregation and data visualization.

Functional enrichment analysis is an analytical method to extract biological insights from gene expression data, popularized by the ever-growing application of high-throughput techniques. Typically, expression profiles are generated for hundreds to thousands of genes/proteins from samples belonging to two experimental groups, and after ad-hoc statistical tests, researchers are left with lists of statistically significant entities, possibly lacking any unifying biological theme. Functional enrichment tackles the problem of putting overall gene expression changes into a broader biological context, based on pre-existing knowledge bases of reference: database collections of known expression regulation, relationships and molecular interactions. STRING is among the most popular tools, providing both protein-protein interaction networks and functional enrichment analysis for any given set of identifiers. For complex experimental designs, manually retrieving, interpreting, analyzing and abridging functional enrichment results is a daunting task, usually performed by hand by the average wet-biology researcher. We have developed reString, a cross-platform software that seamlessly retrieves from STRING functional enrichments from multiple user-supplied gene sets, with just a few clicks, without any need for specific bioinformatics skills. Further, it aggregates all findings into human-readable table summaries, with built-in features to easily produce user-customizable publication-grade clustermaps and bubble plots. Herein, we outline a complete reString protocol, showcasing its features on a real use-case.