High miR-133a levels in the circulation anticipates presentation of clinical events in familial hypercholesterolaemia patients.

AIMS: Presentation of acute events in patients with atherosclerosis remains unpredictable even after controlling for classical risk factors. MicroRNAs (miRNAs) measured in liquid biopsies could be good candidate biomarkers to improve risk prediction. Here, we hypothesized that miRNAs could predict atherosclerotic plaque progression and clinical event presentation in familial hypercholesterolaemia (FH) patients. METHODS AND RESULTS: Circulating miRNAs (plasma, exosomes, and microvesicles) were investigated by TaqMan Array and RT-qPCR assays. Patients with genetic diagnosis of FH and healthy relatives from the SAFEHEART cohort were included. A differential signature of 10 miRNA was obtained by comparing two extreme phenotypes consisting of FH patients suffering a cardiovascular event (CVE) within a 8-year follow-up period (FH-CVE, N = 42) and non-FH hypercholesterolaemic relatives from the same cohort, matched for age and treatment, without CVE during the same period (nFH-nCVE, N = 30). The validation studies included two independent groups of patients with FH background (discovery group, N = 89, validation group N = 196), developing a future CVE (FH-CVE) or not (FH-nCVE) within the same time period of follow-up. Of the 10 miRNAs initially selected, miR-133a was significantly higher in FH-CVE than in FH-nCVE patients. Receiver operating characteristic analysis confirmed miR-133a as the best microRNA for predicting CVE in FH patients (0.76 ± 0.054; P < 0.001). Furthermore, Kaplan-Meier and COX analysis showed that high plasma miR-133a levels associated to the higher risk of presenting a CVE within the next 8 years (hazard ratio 3.89, 95% confidence interval 1.88-8.07; P < 0.001). In silico analysis of curate biological interactions related miR-133a with target genes involved in regulation of the cell-membrane lipid-receptor LRP6 and inflammatory cytokines (CXCL8, IL6, and TNF). These predictions were experimentally proven in human macrophages and endothelial cells transfected with agomiR-133a. CONCLUSION: Elevated levels of miR-133a in the circulation anticipate those FH patients that are going to present a clinical CVE within the next 2 years (average). Mechanistically, miR-133a is directly related with lipid- and inflammatory signalling in key cells for atherosclerosis progression.

Detrimental Effect of Hypercholesterolemia on High-Density Lipoprotein Particle Remodeling in Pigs.

BACKGROUND: Beneficial effects of high-density lipoproteins (HDL) seem altered in patients with symptomatic cardiovascular disease. We recently demonstrated in a swine model of ischemia-reperfusion (IR) that hypercholesterolemia abolishes HDL-related cardioprotection. OBJECTIVES: This study sought to investigate, using the same animal model, whether the reported impairment of HDL cardioprotective function was associated with alterations in HDL remodeling and functionality. METHODS: Pigs were fed a normocholesterolemic (NC) or hypercholesterolemic (HL) diet for 10 days, reaching non-HDL cholesterol concentrations of 38.2 ± 3.5 mg/dl and 218.6 ± 27.6 mg/dl, respectively (p < 0.0001). HDLs were isolated, and lipidomics and differential proteomics tests were performed to determine HDL molecular changes. HDL functionality and particle size were determined. RESULTS: Using principal component analysis, we identified 255 molecular lipid species differentially clustered in NC-HDL and HL-HDL. Ninety lipid metabolites were differentially expressed, and 50 showed at least 1.5-fold variation (false discovery rate adjustment q value <0.05). HL-HDLs presented a core enriched in cholesteryl esters and a surface depleted of phosphatidylcholine species containing polyunsaturated and long-chain fatty acids, indicating the presence of mature HDL particles with low surface fluidity. Hypercholesterolemia induced an important change in HDL-transported proteins (576 spots in HL-HDL vs. 621 spots in NC-HDL). HL-HDLs showed a reduced content of lipocalin retinol binding protein 4 and apolipoprotein M and in the retinoic acid-transporter cellular retinoic acid binding protein 1 (p < 0.05 vs. NC-HDL). No changes were observed in apolipoprotein A-I content and profile. Functionally, HL-HDL showed lower antioxidant activity (-35%) and a reduced capacity to efflux cholesterol (-60%) compared to NC-HDL (p < 0.05). Hypercholesterolemia induced larger HDL particles. CONCLUSIONS: We demonstrate that hypercholesterolemia induces HDL lipidomic changes, losing phosphatidylcholine-lipid species and gaining cholesteryl esters, and proteomic changes, with losses in cardioprotective proteins. These remodeling changes shifted HDL particles toward a dysfunctional state.

Lactobacillus plantarum CECT 7315/7316 intake modulates the acute and chronic innate inflammatory response.

PURPOSE: Probiotics may confer health benefits for the host. Although Lactobacillus has demonstrated to stimulate the immune response, only a few strains have demonstrated immunomodulatory properties. The newly identified Lactobacillus plantarum strains CECT7315 and CECT7316 (LP3457) seem to boost the immune system in individuals that immune decline. We aimed to investigate whether LP3457 protects against inflammation and the mechanism behind. METHODS: LP3457 potential anti-inflammatory effects were assessed in an acute model LPS-induced inflammation in healthy rats and in a chronic model of low-grade inflammation in Zucker diabetic fatty (ZDF) rats. Wistar rats received LP3457 or placebo control for 20 days. Lipopolysaccharide (LPS; 1 mg/kg) was injected intraperitoneally at day 14, and animals were sacrificed 6 days after. Blood was collected at baseline (day 0) and consecutively at day 7, 14, 17, and 20 for haematological evaluation and assessment of anti-inflammatory/pro-inflammatory systemic markers. Myeloperoxidase activity was investigated in the ileum. ZDF rats received LP3457 or placebo control during 8 weeks, and changes in inflammasome-related transcripts were assessed in the ileum. RESULTS: LPS induced a comparable and significant leucocytosis 3 days post-injection (day 17) in both LP3457-treated and LP3457-untreated rats. However, the probiotic supplementation attenuated IL-1ß, IL-6, and CRP release and increased anti-inflammatory IL-10 levels 6 days post-LPS induction (p < 0.05 vs. placebo). LP3457-supplemented animals also displayed lower intestinal myeloperoxidase activity (p < 0.05 vs. placebo). Chronic administration of LP3457 to ZDF rats resulted in a significant downregulation of the inflammasome signalling pathway (p < 0.05 vs. placebo). CONCLUSIONS: Intake of LP3457 attenuates both acute endotoxemia-induced and chronic metabolically induced inflammatory reactions and the inflammasome signalling pathway. The stabilization and regulation of the gut microbiota is an important target for reducing the impact of organ-related inflammatory reactions.