VEGF-A Regulates Cellular Localization of SR-BI as Well as Transendothelial Transport of HDL but Not LDL.

OBJECTIVE: Low- and high-density lipoproteins (LDL and HDL) must pass the endothelial layer to exert pro- and antiatherogenic activities, respectively, within the vascular wall. However, the rate-limiting factors that mediate transendothelial transport of lipoproteins are yet little known. Therefore, we performed a high-throughput screen with kinase drug inhibitors to identify modulators of transendothelial LDL and HDL transport. APPROACH AND RESULTS: Microscopy-based high-content screening was performed by incubating human aortic endothelial cells with 141 kinase-inhibiting drugs and fluorescent-labeled LDL or HDL. Inhibitors of vascular endothelial growth factor (VEGF) receptors (VEGFR) significantly decreased the uptake of HDL but not LDL. Silencing of VEGF receptor 2 significantly decreased cellular binding, association, and transendothelial transport of 125I-HDL but not 125I-LDL. RNA interference with VEGF receptor 1 or VEGF receptor 3 had no effect. Binding, uptake, and transport of HDL but not LDL were strongly reduced in the absence of VEGF-A from the cell culture medium and were restored by the addition of VEGF-A. The restoring effect of VEGF-A on endothelial binding, uptake, and transport of HDL was abrogated by pharmacological inhibition of phosphatidyl-inositol 3 kinase/protein kinase B or p38 mitogen-activated protein kinase, as well as silencing of scavenger receptor BI. Moreover, the presence of VEGF-A was found to be a prerequisite for the localization of scavenger receptor BI in the plasma membrane of endothelial cells. CONCLUSIONS: The identification of VEGF as a regulatory factor of transendothelial transport of HDL but not LDL supports the concept that the endothelium is a specific and, hence, druggable barrier for the entry of lipoproteins into the vascular wall.

Antiatherogenic effects of ellagic acid and urolithins in vitro.

Atherosclerosis, one of the leading causes of death worldwide, is characterized by impaired endothelial function and lipid metabolism, among other factors. Ellagitannins are a class of phenolic compounds that may play a role in cardiovascular health. This work aimed to study the potential atheroprotective effects of urolithins, ellagitannin-derived gut microbiota metabolites, on different key factors in atherosclerosis development: the ability of monocytes to adhere to endothelial cells and the uptake and efflux of cholesterol by macrophages. The biotransformations urolithins undergo in peripheral cells were also evaluated. Results indicated that some urolithins and ellagic acid were able to reduce the adhesion of THP-1 monocytes to human umbilical vein endothelial cells (HUVECs) and the secretion of a cellular adhesion molecule (sVCAM-1) and pro-inflammatory cytokine (IL-6). Urolithin C, a combination of urolithins A and B, and ellagic acid also decreased the accumulation of cholesterol in THP-1-derived macrophages, but they were not able to promote cholesterol efflux. The analysis of cell media by UHPLC-ESI-MS(n) indicated urolithins and ellagic underwent extensive metabolism, with sulfate and methyl conjugation. This evidence indicates that atherosclerotic processes may be attenuated by urolithins, but future human intervention trials are required to establish if is translated in vivo.

Evidence for cholesterol-lowering activity by Bifidobacterium bifidum PRL2010 through gut microbiota modulation.

Bifidobacteria are members of the human gut microbiota, which are known to influence the metabolic abilities of their host. Here, we investigated the capabilities of bifidobacteria to reduce cholesterol levels in synthetic growth media, clearly demonstrating assimilation of this molecule by particular bifidobacterial strains, including Bifidobacterium bifidum PRL2010 (LMG S-28692). The transcriptomic analysis of PRL2010 cells cultivated in the presence of cholesterol revealed a significantly increased transcription level of genes encoding putative transporters and reductases, indicative of specific mechanisms for cholesterol assimilation as well as cholesterol conversion to coprostanol. Cholesterol lowering activity of B. bifidum PRL2010 cells was further evaluated by means of an in vivo murine model, showing that the fecal microbiota of mice is modified toward those bacteria involved in the metabolism of cholesterol.

Anti-Atherosclerotic Effect of a Polyphenol-Rich Ingredient, Oleactiv®, in a Hypercholesterolemia-Induced Golden Syrian Hamster Model.

The development of nutraceutical ingredients has risen as a nutritional solution for health prevention. This study evaluated the effects of Oleactiv®, an ingredient developed for the prevention of atherogenesis, in hypercholesterolemic hamsters. Oleactiv® is a polyphenol-rich ingredient obtained from artichoke, olive and grape extracts as part of fruit and vegetables commonly consumed within the Mediterranean diet. A total of 21 Golden Syrian hamsters were divided into three groups. The standard group (STD) was fed a normolipidemic diet for 12 weeks, while the control group (CTRL) and Oleactiv® goup (OLE) were fed a high-fat diet. After sacrifice, the aortic fatty streak area (AFSA), plasmatic total cholesterol (TC), high-density lipoproteins (HDL-C), non-HDL-C and triglycerides (TG), were assessed. The cholesterol efflux capacity (CEC) of hamster plasma was quantified using a radiolabeled technique in murine macrophages J774. OLE administration induced a significant reduction of AFSA (-69%, p < 0.0001). Hamsters of the OLE group showed a significant decrease of both non-HDL-C (-173 mmol/L, p < 0.05) and TG (-154 mmol/L, p < 0.05). Interestingly, OLE induced a significant increase of total CEC (+17,33%, p < 0,05). Oleactiv® supplementation prevented atheroma development and had positive effects on the lipid profile of hypercholesterolemic hamsters. The increased CEC underlines the anti-atherosclerotic mechanism at the root of the atheroma reduction observed.

Alcohol Pattern Consumption Differently Affects the Efficiency of Macrophage Reverse Cholesterol Transport in Vivo.

It has been well established that moderate alcohol consumption inversely correlates with cardiovascular morbidity and mortality, whereas binge alcohol drinking increases cardiovascular disease risk. The aim of this study was to assess in vivo the impact of different drinking patterns on reverse cholesterol transport (RCT); the atheroprotective process leading to the removal of excess cholesterol from the body. RCT was measured with a standardized, radioisotope-based technique in three groups of atherosclerosis-prone apolipoprotein E knock out mice: Placebo group, receiving water, which would mimic the abstainers; moderate group, receiving 0.8 g/kg alcohol/day for 28 days, which would mimic a moderate intake; binge group, receiving 0.8 g/kg alcohol/day for 5 days/week, followed by the administration of 2.8 g/kg alcohol/day for 2 days/week, which would mimic a heavy intake in a short period. Mice in the binge drinking group displayed an increase in total cholesterol, high density lipoprotein cholesterol (HDL-c) and non-HDL-c (all p < 0.0001 vs. placebo), and a significantly reduced elimination of fecal cholesterol. The moderate consumption did not lead to any changes in circulating lipids, but slightly improved cholesterol mobilization along the RCT pathway. Overall, our data confirm the importance of considering not only the total amount, but also the different consumption patterns to define the impact of alcohol on cardiovascular risk.

Oncocytic modifications in rectal adenocarcinomas after radio and chemotherapy.

The purpose of the study is to highlight oncocytic modifications in rectal adenocarcinomas and evaluate a possible correlation with preoperative radiochemotherapy (RCT). Twenty-eight cases of advanced rectal carcinoma, treated preoperatively by 5-fluorouracil (200-225 mg/m(2)) and 44-46 Gy in 22-23 fractions, were studied. All patients underwent biopsy before RCT. Surgery was performed within 6 weeks after RCT. In all cases oncocytic modifications were searched for on hematoxylin and eosin (H&E) and at immunohistochemistry using an antimitochondrial antibody. In addition, in two cases, both pre- and post-RCT tissues were examined at electron microscopy. All tumors were adenocarcinomas. In pre-RCT biopsies, oncocytic changes were difficult to find on H&E, while the antimitochondrial antibody strongly stained numerous neoplastic cells (mean 48.4%). In post-RCT surgical specimens, oncocytic changes were detected in 24 out of 28 cases on H&E and the antimitochondrial antibody stained most of the residual neoplastic cells (mean 76.7%). Ultrastructural examination revealed large and bizarre mitochondria inside tumor cells both in pre- and post-RCT tissues. In conclusion, the present data suggest that rectal adenocarcinomas are "mitochondrion-rich" tumors. After preoperative RCT, residual neoplastic cells acquire a definite oncocytic phenotype.

High MUC2 immunohistochemical expression is a predictor of poor response to preoperative radiochemotherapy (RCT) in rectal adenocarcinoma.

The purpose of this study is to establish if mucoid differentiation is associated with responsiveness to preoperative radiochemotherapy (RCT) in rectal adenocarcinomas. Thirty-two patients with rectal adenocarcinomas were preoperatively treated with 44 to 46 Gy in 22 to 23 fractions and with 5-fluorouracil (200 to 225 mg/m) before surgery. Mucoid differentiation was searched for both in pre-RCT biopsies with anti-MUC2 antiserum and in postoperative specimens. To evaluate the responsiveness to preoperative RCT, a regression grading was used (grades 0 to 4). Statistical analysis showed a significant negative correlation between immunohistochemical expression of MUC2 in pre-RCT biopsies and regression grade in postoperative specimens (r=-0.529; P=0.002). A significant cutoff value of 60% of MUC2 positive neoplastic cells in pre-RCT biopsies was observed (P=0.018): 13 cases with more than 60% exhibited a poor response to RCT (grade 0 in 5/13, grade 1 in 4/13, grade 2 in 4/13), whereas 19 cases with less than 60% showed a better response to RCT (grade 1 in 6/19, grade 2 in 9/19, grade 3 in 3/19, grade 4 in 1/19).