Novel Associations between Related Proteins and Cellular Effects of High-Density Lipoprotein.

BACKGROUND AND OBJECTIVES: Recent studies have examined the structure-function relationship of high-density lipoprotein (HDL). This study aimed to identify and rank HDL-associated proteins involved in several biological function of HDL. METHODS: HDLs isolated from 48 participants were analyzed. Cholesterol efflux capacity, effect of HDL on nitric oxide production, and vascular cell adhesion molecule-1 expression were assessed. The relative abundance of identified proteins in the highest vs. lowest quartile was expressed using the normalized spectral abundance factor ratio. RESULTS: After adjustment by multiple testing, six proteins, thyroxine-binding globulin, alpha-1B-glycoprotein, plasma serine protease inhibitor, vitronectin, angiotensinogen, and serum amyloid A-4, were more abundant (relative abundance ratio ≥2) in HDLs with the highest cholesterol efflux capacity. In contrast, three proteins, complement C4-A, alpha-2-macroglobulin, and immunoglobulin mu chain C region, were less abundant (relative abundance ratio <0.5). In terms of nitric oxide production and vascular cell adhesion molecule-1 expression, no proteins showed abundance ratios ≥2 or <0.5 after adjustment. Proteins correlated with the functional parameters of HDL belonged to diverse biological categories. CONCLUSIONS: In summary, this study ranked proteins showing higher or lower abundance in HDLs with high functional capacities and newly identified multiple proteins linked to cholesterol efflux capacity.

Novel association between CDKAL1 and cholesterol efflux capacity: Replication after GWAS-based discovery.

BACKGROUND AND AIMS: Although the importance of the functional properties of high-density lipoprotein (HDL) has been increasingly emphasized, studies on the genetic factors associated with HDL function are highly limited. The aim of this study was to identify genetic variants associated with an individual's cholesterol efflux capacity (CEC) using a genome-wide association study approach. METHODS: This study included a discovery group of 607 subjects with coronary artery disease and an independent replication group of 158 subjects. CEC was assessed using a radioisotope and ApoB-depleted serum. Genome-wide associations between the adjusted CEC and genotyped and imputed variants were examined with linear regression, assuming an additive genetic model. Finally, adjustments were made for confounding parameters to assess the independence of associations and to determine R2 of overall model on CEC. RESULTS: In the discovery group, 631 variants showed significant association with CEC, and five of them were found to correlate with CEC in the replication group. One of them was located near LOC541471 in 2q13, whereas the other four (rs117835232, rs117252933, rs118064592, and rs150434350) were located in CDKAL1 in 6p22.3. The association between the presence of any CDKAL1 variant and CEC was significant after adjustment for clinical and laboratory variables. High-density lipoprotein-cholesterol levels also showed a very significant association with CEC. Body mass index, current alcohol use, triglycerides levels, low-density lipoprotein-cholesterol levels and statin use showed borderline associations with CEC. CONCLUSIONS: We identified and replicated genetic variants associated with CEC using a genome-wide association study-based approach. CDKAL1 variants showed correlations with CEC independent of HDL-cholesterol levels and other clinical characteristics.

Palmitate and minimally-modified low-density lipoprotein cooperatively promote inflammatory responses in macrophages.

Increased consumption of Western-type diets and environmental insults lead to wide-spread increases in the plasma levels of saturated fatty acids and lipoprotein oxidation. The aim of this study is to examine whether palmitate and minimally modified low-density lipoprotein (mmLDL) exert an additive effect on macrophage activation. We found that CXCL2 and TNF-α secretion as well as ERK and p38 phosphorylation were additively increased by co-treatment of J774 macrophages with palmitate and mmLDL in the presence of lipopolysaccharide (LPS). Furthermore, the analysis of differentially expressed genes using the KEGG database revealed that several pathways, including cytokine-cytokine receptor interaction, and genes were significantly altered. These results were validated with real-time PCR, showing upregulation of Il-6, Csf3, Il-1ß, and Clec4d. The present study demonstrated that palmitate and mmLDL additively potentiate the LPS-induced activation of macrophages. These results suggest the existence of synergistic mechanisms by which saturated fatty acids and oxidized lipoproteins activate immune cells.

Effect of two lipid-lowering strategies on high-density lipoprotein function and some HDL-related proteins: a randomized clinical trial.

BACKGROUND: The influence of lipid-lowering therapy on high-density lipoprotein (HDL) is incompletely understood. We compared the effect of two lipid-lowering strategies on HDL functions and identified some HDL-related proteins. METHODS: Thirty two patients were initially screened and HDLs of 21 patients were finally analyzed. Patients were randomized to receive atorvastatin 20 mg (n = 11) or atorvastatin 5 mg/ezetimibe 10 mg combination (n = 10) for 8 weeks. The cholesterol efflux capacity and other anti-inflammatory functions were assessed based on HDLs of the participants before and after treatment. Pre-specified HDL proteins of the same HDL samples were measured. RESULTS: The post-treatment increase in cholesterol efflux capacities was similar between the groups (35.6% and 34.6% for mono-therapy and combination, respectively, p = 0.60). Changes in nitric oxide (NO) production, vascular cell adhesion molecule-1 (VCAM-1) expression, and reactive oxygen species (ROS) production were similar between the groups. The baseline cholesterol efflux capacity correlated positively with apolipoprotein (apo)A1 and C3, whereas apoA1 and apoC1 showed inverse associations with VCAM-1 expression. The changes in the cholesterol efflux capacity were positively correlated with multiple HDL proteins, especially apoA2. CONCLUSIONS: Two regimens increased the cholesterol efflux capacity of HDL comparably. Multiple HDL proteins, not limited to apoA1, showed a correlation with HDL functions. These results indicate that conventional lipid therapy may have additional effects on HDL functions with changes in HDL proteins. TRIAL REGISTRATION: ClinicalTrials.gov, number NCT02942602 .

Discovery and characterization of novel microRNAs during endothelial differentiation of human embryonic stem cells.

MicroRNAs (miRNAs) are small RNAs that participate in the regulation of genes associated with the differentiation and proliferation. In this study, 5 novel miRNAs were identified from human mesenchymal stem cells and characterized using various analyses. To investigate the potential functions associated with the regulation of cell differentiation, the differences in miRNA expression were examined in undifferentiated and differentiated human embryonic stem (ES) cells using reverse transcription (RT)-PCR analysis. Specifically, 3 miRNAs exhibited decreased expression levels in human umbilical vein endothelial cells (HUVECs) and endothelial cells derived from human ES cells. Putative target genes related to differentiation or maturation of endothelial cells were predicted by seed sequences of 2 novel miRNAs and analyzed for their expression via miRNA-mediated regulation using a luciferase assay. In HUVECs, CDH5 gene expression was directly repressed by hsa-miR-6086. Similarly, hsa-miR-6087 significantly downregulated endoglin expression. Therefore, the roles of these 2 miRNAs may be to directly suppress their target genes, popularly known as endothelial cell markers. Taken together, our results demonstrate that several novel miRNAs perform critical roles in human endothelial cell development.