Recent studies indicate high density lipoproteins (HDL) and their major structural protein, apolipoprotein A1 (apoA1), recovered from human atheroma, are dysfunctional and extensively oxidized by myeloperoxidase (MPO), while in vitro oxidation of apoA1/HDL by MPO impairs its cholesterol acceptor function. We developed a high affinity monoclonal antibody (mAb) that specifically recognizes apoA1/HDL modified by the MPO/H2O2/Cl-system using phage display affinity maturation. An oxindolyl alanine (2-OH-Trp) moiety at tryptophan 72 of apoA1 is the immunogenic epitope. Mutagenesis studies confirm a critical role for apoA1 Trp72 in MPO-mediated inhibition of ABCA1-dependent cholesterol acceptor activity of apoA1 in vitro and in vivo. ApoA1 containing a 2-OH-Trp72 group (oxTrp72-apoA1) is in low abundance within the circulation, but accounts for 20% of the apoA1 in atherosclerotic plaque. OxTrp72-apoA1 recovered from human atheroma or plasma was lipid-poor, virtually devoid of cholesterol acceptor activity, and demonstrated both potent pro-inflammatory activities on endothelial cells and impaired HDL biogenesis activity in vivo. Elevated oxTrp72-apoA1 levels in subjects presenting to a cardiology clinic (n=627) were associated with increased cardiovascular disease risk. Circulating oxTrp72-apoA1 levels may serve as a way to monitor a pro-atherogenic process in the artery wall.

Objective Preclinical and clinical studies have shown beneficial effects of infusions of apolipoprotein A-I (ApoA-I) on atherosclerosis. ApoA-I is also a target for myeloperoxidase (MPO)-mediated oxidation, leading in vitro to a loss of its ability to promote ABCA1-dependent macrophage cholesterol efflux. Therefore, we hypothesized that MPO-mediated ApoA-I oxidation would impair its promotion of reverse cholesterol transport (RCT) in vivo and the beneficial effects on atherosclerotic plaques. Approach and Results ApoA-I−/− or ApoE−/− mice were subcutaneously injected with native human ApoA-I, oxidized human ApoA-I (oxApoAI; MPO/hydrogen peroxide/chloride treated) or carrier. While early post injection (8 hours) levels of total ApoA-I in plasma were similar for native versus oxApoA-I, native ApoA-I primarily resided within the HDL fraction, whereas the majority of oxApoA-I was highly cross-linked and not HDL particle associated, consistent with impaired ABCA1 interaction. In ApoA-I−/− mice, ApoA-I oxidation significantly impaired RCT in vivo. In advanced aortic root atherosclerotic plaques of ApoE−/− mice, native ApoA-I injections led to significant decreases in lipid content, macrophage number, and an increase in collagen content; in contrast, oxApoA-I failed to mediate these changes. The decrease in plaque macrophages with native ApoA-I was accompanied by significant induction of their chemokine receptor CCR7. Furthermore, only native ApoA-I injections led to a significant reduction of inflammatory M1 and increase in anti-inflammatory M2 macrophage markers in the plaques. Conclusions MPO-mediated oxidation renders ApoA-I dysfunctional and unable to: (i) promote RCT; (ii) mediate beneficial changes in the composition of atherosclerotic plaques; and (iii) pacify the inflammatory status of plaque macrophages.

BackgroundA pilot study was performed in order to investigate the effects of bariatric surgery on whole blood gene expression profiles in obese subjects with type 2 diabetes.Methodology/Principal FindingsWhole blood from eleven obese subjects with type 2 diabetes was collected in PAXgene tubes prior to and 6–12 months after bariatric surgery. Total RNA was isolated, amplified, labeled and hybridized to Illumina gene expression microarrays. Clinical and expression data were analyzed using a paired t-test, and correlations between changes in clinical trait and transcript levels were calculated. Pathways were identified using Ingenuity Pathway Analysis and DAVID gene ontology software. Overall, bariatric surgery resulted in significant reduction of body mass index, fasting plasma glucose, fasting plasma insulin, and normalization of glycosylated hemoglobin levels. The expression levels of 204 transcripts, representing 200 unique genes, were significantly altered after bariatric surgery. Among the significantly regulated genes were GGT1, CAMP, DEFA1, LCN2, TP53, PDSS1, OLR1, CNTNAP5, DHCR24, HHAT and SARDH, which have been previously implicated in lipid metabolism, obesity and/or type 2 diabetes. Selected findings were replicated by quantitative real-time-PCR. The changes in expression of seven transcripts, WDR35, FLF45244, DHCR24, TIGD7, TOPBP1, TSHZ1, and FAM8A1 were strongly correlated with the changes in body weight, fasting plasma glucose and glycosylated hemoglobin content. The top pathways associated with gene expression changes after bariatric surgery was lipid metabolism, small molecule biochemistry and gene expression. Two antimicrobial peptides were among the transcripts with the largest changes in gene expression after bariatric surgery.Conclusions/SignificanceData from this pilot study suggest that whole blood expression levels of specific transcripts may be useful as biomarkers associated with susceptibility for type 2 diabetes and/or therapeutic response.