Mitochondrial Isolevuglandins Contribute to Vascular Oxidative Stress and Mitochondria-Targeted Scavenger of Isolevuglandins Reduces Mitochondrial Dysfunction and Hypertension.

Hypertension remains a major health problem in Western Societies, and blood pressure is poorly controlled in a third of patients despite use of multiple drugs. Mitochondrial dysfunction contributes to hypertension, and mitochondria-targeted agents can potentially improve treatment of hypertension. We have proposed that mitochondrial oxidative stress produces reactive dicarbonyl lipid peroxidation products, isolevuglandins, and that scavenging of mitochondrial isolevuglandins improves vascular function and reduces hypertension. To test this hypothesis, we have studied the accumulation of mitochondrial isolevuglandins-protein adducts in patients with essential hypertension and Ang II (angiotensin II) model of hypertension using mass spectrometry and Western blot analysis. The therapeutic potential of targeting mitochondrial isolevuglandins was tested by the novel mitochondria-targeted isolevuglandin scavenger, mito2HOBA. Mitochondrial isolevuglandins in arterioles from hypertensive patients were 250% greater than in arterioles from normotensive subjects, and ex vivo mito2HOBA treatment of arterioles from hypertensive subjects increased deacetylation of a key mitochondrial antioxidant, SOD2 (superoxide dismutase 2). In human aortic endothelial cells stimulated with Ang II plus TNF (tumor necrosis factor)-α, mito2HOBA reduced mitochondrial superoxide and cardiolipin oxidation, a specific marker of mitochondrial oxidative stress. In Ang II-infused mice, mito2HOBA diminished mitochondrial isolevuglandins-protein adducts, raised Sirt3 (sirtuin 3) mitochondrial deacetylase activity, reduced vascular superoxide, increased endothelial nitric oxide, improved endothelium-dependent relaxation, and attenuated hypertension. Mito2HOBA preserved mitochondrial respiration, protected ATP production, and reduced mitochondrial permeability pore opening in Ang II-infused mice. These data support the role of mitochondrial isolevuglandins in endothelial dysfunction and hypertension. We conclude that scavenging of mitochondrial isolevuglandins may have therapeutic potential in treatment of vascular dysfunction and hypertension.

Scavenging of reactive dicarbonyls with 2-hydroxybenzylamine reduces atherosclerosis in hypercholesterolemic Ldlr-/- mice.

Lipid peroxidation generates reactive dicarbonyls including isolevuglandins (IsoLGs) and malondialdehyde (MDA) that covalently modify proteins. Humans with familial hypercholesterolemia (FH) have increased lipoprotein dicarbonyl adducts and dysfunctional HDL. We investigate the impact of the dicarbonyl scavenger, 2-hydroxybenzylamine (2-HOBA) on HDL function and atherosclerosis in Ldlr-/- mice, a model of FH. Compared to hypercholesterolemic Ldlr-/- mice treated with vehicle or 4-HOBA, a nonreactive analogue, 2-HOBA decreases atherosclerosis by 60% in en face aortas, without changing plasma cholesterol. Ldlr-/- mice treated with 2-HOBA have reduced MDA-LDL and MDA-HDL levels, and their HDL display increased capacity to reduce macrophage cholesterol. Importantly, 2-HOBA reduces the MDA- and IsoLG-lysyl content in atherosclerotic aortas versus 4-HOBA. Furthermore, 2-HOBA reduces inflammation and plaque apoptotic cells and promotes efferocytosis and features of stable plaques. Dicarbonyl scavenging with 2-HOBA has multiple atheroprotective effects in a murine FH model, supporting its potential as a therapeutic approach for atherosclerotic cardiovascular disease.

Safety, tolerability, and pharmacokinetics of repeated oral doses of 2-hydroxybenzylamine acetate in healthy volunteers: a double-blind, randomized, placebo-controlled clinical trial.

BACKGROUND: 2-Hydroxybenzylamine (2-HOBA) is a selective dicarbonyl electrophile scavenger being developed as a nutritional supplement to help protect against the development of conditions associated with dicarbonyl electrophile formation, such as the cognitive decline observed with Mild Cognitive Impairment or Alzheimer's disease. METHODS: This study evaluated the safety, tolerability, and pharmacokinetics of repeated oral doses of 2-HOBA acetate (500 or 750 mg) administered to healthy volunteers every eight hours for two weeks. The effects of 2-HOBA on cyclooxygenase function and cerebrospinal fluid penetrance of 2-HOBA were also investigated. RESULTS: Repeated oral administration of 2-HOBA was found to be safe and well-tolerated up to 750 mg TID for 15 days. 2-HOBA was absorbed within 2 h of administration, had a half-life of 2.10-3.27 h, and an accumulation ratio of 1.38-1.52. 2-HOBA did not interfere with cyclooxygenase function and was found to be present in cerebrospinal fluid 90 min after dosing. CONCLUSIONS: Repeated oral administration of 2-HOBA was found to be safe and well-tolerated. These results support continued development of 2-HOBA as a nutritional supplement. TRIAL REGISTRATION: Studies are registered at ClinicalTrials.gov (NCT03555682 Registered 13 June 2018, NCT03554096 Registered 12 June 18).

Arachidonic Acid Kills Staphylococcus aureus through a Lipid Peroxidation Mechanism.

Staphylococcus aureus infects every niche of the human host. In response to microbial infection, vertebrates have an arsenal of antimicrobial compounds that inhibit bacterial growth or kill bacterial cells. One class of antimicrobial compounds consists of polyunsaturated fatty acids, which are highly abundant in eukaryotes and encountered by S. aureus at the host-pathogen interface. Arachidonic acid (AA) is one of the most abundant polyunsaturated fatty acids in vertebrates and is released in large amounts during the oxidative burst. Most of the released AA is converted to bioactive signaling molecules, but, independently of its role in inflammatory signaling, AA is toxic to S. aureus Here, we report that AA kills S. aureus through a lipid peroxidation mechanism whereby AA is oxidized to reactive electrophiles that modify S. aureus macromolecules, eliciting toxicity. This process is rescued by cotreatment with antioxidants as well as in a S. aureus strain genetically inactivated for lcpA (USA300 ΔlcpA mutant) that produces lower levels of reactive oxygen species. However, resistance to AA stress in the USA300 ΔlcpA mutant comes at a cost, making the mutant more susceptible to ß-lactam antibiotics and attenuated for pathogenesis in a murine infection model compared to the parental methicillin-resistant S. aureus (MRSA) strain, indicating that resistance to AA toxicity increases susceptibility to other stressors encountered during infection. This report defines the mechanism by which AA is toxic to S. aureus and identifies lipid peroxidation as a pathway that can be modulated for the development of future therapeutics to treat S. aureus infections.IMPORTANCE Despite the ability of the human immune system to generate a plethora of molecules to control Staphylococcus aureus infections, S. aureus is among the pathogens with the greatest impact on human health. One class of host molecules toxic to S. aureus consists of polyunsaturated fatty acids. Here, we investigated the antibacterial properties of arachidonic acid, one of the most abundant polyunsaturated fatty acids in humans, and discovered that the mechanism of toxicity against S. aureus proceeds through lipid peroxidation. A better understanding of the molecular mechanisms by which the immune system kills S. aureus, and by which S. aureus avoids host killing, will enable the optimal design of therapeutics that complement the ability of the vertebrate immune response to eliminate S. aureus infections.

First-in-human study assessing safety, tolerability, and pharmacokinetics of 2-hydroxybenzylamine acetate, a selective dicarbonyl electrophile scavenger, in healthy volunteers.

BACKGROUND: 2-Hydroxybenzylamine (2-HOBA) is a selective scavenger of dicarbonyl electrophiles that protects proteins and lipids from being modified by these electrophiles. It is currently being developed for use as a nutritional supplement to help maintain good health and protect against the development of conditions associated with dicarbonyl electrophile formation, such as the cognitive decline associated with Mild Cognitive Impairment and Alzheimer's disease. METHODS: In this first-in-human study, the safety, tolerability, and pharmacokinetics of six ascending single oral doses of 2-HOBA acetate were tested in eighteen healthy human volunteers. RESULTS: Reported adverse events were mild and considered unlikely to be related to 2-HOBA. There were no clinically significant changes in vital signs, ECG recordings, or clinical laboratory parameters. 2-HOBA was fairly rapidly absorbed, with a tmax of 1-2 h, and eliminated, with a t1/2 of approximately 2 h. Both tmax and t1/2 were independent of dose level, while Cmax and AUC increased proportionally with dose level. CONCLUSIONS: 2-HOBA acetate was safe and well-tolerated at doses up to 825 mg in healthy human volunteers, positioning it as a good candidate for continued development as a nutritional supplement. TRIAL REGISTRATION: This study is registered at ClinicalTrials.gov (NCT03176940).

Simplified LC/MS assay for the measurement of isolevuglandin protein adducts in plasma and tissue samples.

Isolevuglandins (IsoLGs) are a family of highly reactive 4-ketoaldehydes formed by lipid peroxidation that modify the lysyl residues of cellular proteins. Modification of proteins by IsoLGs have been shown to contribute to disease processes such as the development of hypertension. Accurate quantitation of the extent of protein modification by IsoLGs is essential for understanding the mechanisms whereby these modifications contribute to disease and the efficacy of interventions designed to prevent this modification. The previously described LC/MS assay to quantitate IsoLG protein adducts was extremely labor-intensive and time consuming, and while it offered reasonably low intra-day variation for replicate samples, variation when replicate samples were processed on separate days was significant. These limitations significantly restricted utilization of this approach. We therefore performed a series of studies to optimize the assay. We now report a significantly simplified LC/MS assay for measurement of IsoLG protein adducts with increased sensitivity and lower intra-day and inter-day variability.

Bid maintains mitochondrial cristae structure and function and protects against cardiac disease in an integrative genomics study.

Bcl-2 family proteins reorganize mitochondrial membranes during apoptosis, to form pores and rearrange cristae. In vitro and in vivo analysis integrated with human genetics reveals a novel homeostatic mitochondrial function for Bcl-2 family protein Bid. Loss of full-length Bid results in apoptosis-independent, irregular cristae with decreased respiration. Bid-/- mice display stress-induced myocardial dysfunction and damage. A gene-based approach applied to a biobank, validated in two independent GWAS studies, reveals that decreased genetically determined BID expression associates with myocardial infarction (MI) susceptibility. Patients in the bottom 5% of the expression distribution exhibit >4 fold increased MI risk. Carrier status with nonsynonymous variation in Bid's membrane binding domain, BidM148T, associates with MI predisposition. Furthermore, Bid but not BidM148T associates with Mcl-1Matrix, previously implicated in cristae stability; decreased MCL-1 expression associates with MI. Our results identify a role for Bid in homeostatic mitochondrial cristae reorganization, that we link to human cardiac disease.

Dysfunctional BMPR2 signaling drives an abnormal endothelial requirement for glutamine in pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is increasingly recognized as a systemic disease driven by alteration in the normal functioning of multiple metabolic pathways affecting all of the major carbon substrates, including amino acids. We found that human pulmonary hypertension patients (WHO Group I, PAH) exhibit systemic and pulmonary-specific alterations in glutamine metabolism, with the diseased pulmonary vasculature taking up significantly more glutamine than that of controls. Using cell culture models and transgenic mice expressing PAH-causing BMPR2 mutations, we found that the pulmonary endothelium in PAH shunts significantly more glutamine carbon into the tricarboxylic acid (TCA) cycle than wild-type endothelium. Increased glutamine metabolism through the TCA cycle is required by the endothelium in PAH to survive, to sustain normal energetics, and to manifest the hyperproliferative phenotype characteristic of disease. The strict requirement for glutamine is driven by loss of sirtuin-3 (SIRT3) activity through covalent modification by reactive products of lipid peroxidation. Using 2-hydroxybenzylamine, a scavenger of reactive lipid peroxidation products, we were able to preserve SIRT3 function, to normalize glutamine metabolism, and to prevent the development of PAH in BMPR2 mutant mice. In PAH, targeting glutamine metabolism and the mechanisms that underlie glutamine-driven metabolic reprogramming represent a viable novel avenue for the development of potentially disease-modifying therapeutics that could be rapidly translated to human studies.

Assessment of complementary feeding of Canadian infants: effects on microbiome & oxidative stress, a randomized controlled trial.

BACKGROUND: The World Health Organization recommends exclusive breastfeeding until 6 months followed by introduction of iron-rich complementary foods (CFs). The aim of this study was to determine the impact of different iron-rich CFs on infant gut inflammation and microbiota. METHODS: Eighty-seven exclusively breastfed infants were randomly assigned to receive one of the following as their first CF: iron-fortified cereal (Cer), iron-fortified cereal with fruit (Cer + Fr), or meat (M). Urine and stool samples were collected to assess reactive oxygen species (ROS) generation, gut microbiota and inflammation. RESULTS: Fecal iron differed across feeding groups (p < 0.001); levels were highest in the Cer group and lowest in M group. A significant increase of fecal ROS formation (p < 0.002) after the introduction of CFs was observed, but did not differ across feeding groups. Fecal calprotectin increased within all groups after the introduction of CFs (p = 0.004). Gut microbiota richness increased after introduction of M or Cer + Fr. Regardless of feeding group, Coriobacteriaceae were positively correlated with ROS and Staphylococcaceae were negatively correlated with calprotectin. CONCLUSIONS: Choice of first CF may influence gut inflammation and microbiota, potentially due to variations in iron absorption from different foods. Further research is warranted to fully characterize these associations and to establish implications for infant health. This study was registered in the ClinicalTrial.gov registry (Identifier No. NCT01790542 ). TRIAL REGISTRATION: This study was registered in the ClinicalTrial.gov registry under the name "Assessment of Complementary Feeding of Canadian Infants" (Identifier No. NCT01790542 ) February 6, 2013.

Intraoperative cerebral oxygenation, oxidative injury, and delirium following cardiac surgery.

BACKGROUND: Delirium affects 20-30% of patients after cardiac surgery and is associated with increased mortality and persistent cognitive decline. Hyperoxic reperfusion of ischemic tissues increases oxidative injury, but oxygen administration remains high during cardiac surgery. We tested the hypothesis that intraoperative hyperoxic cerebral reperfusion is associated with increased postoperative delirium and that oxidative injury mediates this association. METHODS: We prospectively measured cerebral oxygenation with bilateral oximetry monitors in 310 cardiac surgery patients, quantified intraoperative hyperoxic cerebral reperfusion by measuring the magnitude of cerebral oxygenation above baseline after any ischemic event, and assessed patients for delirium twice daily in the ICU following surgery using the confusion assessment method for ICU (CAM-ICU). We examined the association between hyperoxic cerebral reperfusion and postoperative delirium, adjusted for the extent of cerebral hypoxia, the extent of cerebral hyperoxia prior to any ischemia, and additional potential confounders and risk factors for delirium. To assess oxidative injury mediation, we examined the association between hyperoxic cerebral reperfusion and delirium after further adjusting for plasma levels of F2-isoprostanes and isofurans at baseline and ICU admission, the association between hyperoxic cerebral reperfusion and these markers of oxidative injury, and the association between these markers and delirium. RESULTS: Ninety of the 310 patients developed delirium following surgery. Every 10%·hour of intraoperative hyperoxic cerebral reperfusion was independently associated with a 65% increase in the odds of delirium (OR, 1.65 [95% CI, 1.12-2.44]; P=0.01). Hyperoxia prior to ischemia was also independently associated with delirium (1.10 [1.01-1.19]; P=0.02), but hypoxia was not (1.12 [0.97-1.29]; P=0.11). Increased hyperoxic cerebral reperfusion was associated with increased concentrations of F2-isoprostanes and isofurans at ICU admission, increased concentrations of these markers were associated with increased delirium, and the association between hyperoxic cerebral reperfusion and delirium was weaker after adjusting for these markers of oxidative injury. CONCLUSIONS: Intraoperative hyperoxic cerebral reperfusion was associated with increased postoperative delirium, and increased oxidative injury following hyperoxic cerebral reperfusion may partially mediate this association. Further research is needed to assess the potential deleterious role of cerebral hyper-oxygenation during surgery.

Scavenging of highly reactive gamma-ketoaldehydes attenuates cognitive dysfunction associated with epileptogenesis.

Cognitive dysfunction is a major comorbidity of the epilepsies; however, treatments targeting seizure-associated cognitive dysfunction, particularly deficits in learning and memory are not available. Isoketals and neuroketals, collectively known as gamma-ketoaldehydes are formed via the non-enzymatic, free radical catalyzed oxidation of arachidonic acid and docosahexaenoic acid, respectively. They are attractive candidates for oxidative protein damage and resultant cognitive dysfunction due to their formation within the plasma membrane and their high proclivity to form cytotoxic adducts on protein lysine residues. We tested the hypothesis that gamma-ketoaldehydes mechanistically contribute to seizure-associated memory impairment using a specific gamma-ketoaldehyde scavenger, salicylamine in the kainic acid and pilocarpine rat models of temporal lobe epilepsy. We show that gamma-ketoaldehydes are increased following epileptogenic injury in hippocampus and perirhinal cortex, two brain regions imperative for learning and memory. Treatment with an orally bioavailable, brain permeable scavenger, salicylamine attenuated 1) spatial memory deficits 2) reference memory deficits and 3) neuronal loss and astrogliosis in two mechanistically distinct models of epilepsy without affecting the epileptogenic injury or the development of chronic epilepsy. We have previously demonstrated that reactive oxygen species and the lipid peroxidation biomarkers, F2-isoprostanes are produced following status epilepticus. However, which reactive species specifically mediate oxidative damage to cellular macromolecules remains at large. We provide novel data suggesting that memory impairment occurs via gamma-ketoaldehyde production in two models of epilepsy and that treatment with a gamma-ketoaldehyde scavenger can protect vulnerable neurons. This work suggests a novel target and therapy to treat seizure-induced memory deficits in epilepsy.

Scavengers of reactive γ-ketoaldehydes extend Caenorhabditis elegans lifespan and healthspan through protein-level interactions with SIR-2.1 and ETS-7.

Isoketals (IsoKs) are highly reactive γ-ketoaldehyde products of lipid peroxidation that covalently adduct lysine side chains in proteins, impairing their function. Using C. elegans as a model organism, we sought to test the hypothesis that IsoKs contribute to molecular aging through adduction and inactivation of specific protein targets, and that this process can be abrogated using salicylamine (SA), a selective IsoK scavenger. Treatment with SA extends adult nematode longevity by nearly 56% and prevents multiple deleterious age-related biochemical and functional changes. Testing of a variety of molecular targets for SA's action revealed the sirtuin SIR-2.1 as the leading candidate. When SA was administered to a SIR-2.1 knockout strain, the effects on lifespan and healthspan extension were abolished. The SIR-2.1-dependent effects of SA were not mediated by large changes in gene expression programs or by significant changes in mitochondrial function. However, expression array analysis did show SA-dependent regulation of the transcription factor ets-7 and associated genes. In ets-7 knockout worms, SA's longevity effects were abolished, similar to sir-2.1 knockouts. However, SA dose-dependently increases ets-7 mRNA levels in non-functional SIR-2.1 mutant, suggesting that both are necessary for SA's complete lifespan and healthspan extension.

Controlled moderate hypovolaemia in healthy volunteers is not associated with the development of oxidative stress assessed by plasma F2-isoprostanes and isofurans.

Hypovolaemia can be associated with substantial morbidity, particularly when it occurs in the setting of trauma and in patients with comorbid diseases. Hypovolaemia and inflammation such as occur in the setting of trauma and surgery, are associated with systemic oxidative stress and free-radical injury. Free-radical injury that results from hypovolaemia-induced organ reperfusion may further augment inflammatory processes. It is unknown exactly what proportion of free-radical injury is associated with isolated hypovolaemia as opposed to the contribution from inflammation from surgery or trauma. In the first human study of its kind, we exposed 8 adult male volunteers to venesection-induced hypovolaemia in progressive aliquots of 5% of total blood volume until 20% had been removed. This blood was subsequently reinfused. Plasma F2-isoprostanes and isofurans, markers of in vivo lipid oxidation, were measured by gas chromatography-mass spectrometry at each 5% aliquot venesected and at each 5% reinfused. Between baseline and maximal blood loss there was a minor fall in haemoglobin concentration from 143.9g/l to 138.8g/l (p=0.004, 95% CI 2.2, 8.0g/L). No significant change from baseline occurred in the concentrations of either plasma F2-isoprostanes or isofurans during venesection (p=0.116 and p=0.152, respectively) or blood reinfusion (p=0.553 and p=0.736, respectively). We can conclude that in healthy adult volunteers, isolated hypovolaemia to 20% total blood volume loss is not associated with detectable systemic oxidative stress. The free-radical injury identified in surgical and trauma patients may represent the effects of tissue damage and inflammation, with an uncertain contribution from tissue ischemia as may occur with hypovolaemia.

Mitochondrial dysfunction and oxidative stress in patients with chronic kidney disease.

Mitochondria abnormalities in skeletal muscle may contribute to frailty and sarcopenia, commonly present in patients with chronic kidney disease (CKD). Dysfunctional mitochondria are also a major source of oxidative stress and may contribute to cardiovascular disease in CKD We tested the hypothesis that mitochondrial structure and function worsens with the severity of CKD Mitochondrial volume density, mitochondrial DNA (mtDNA) copy number, BNIP3, and PGC1α protein expression were evaluated in skeletal muscle biopsies obtained from 27 subjects (17 controls and 10 with CKD stage 5 on hemodialysis). We also measured mtDNA copy number in peripheral blood mononuclear cells (PBMCs), plasma isofurans, and plasma F2-isoprostanes in 208 subjects divided into three groups: non-CKD (eGFR>60 mL/min), CKD stage 3-4 (eGFR 60-15 mL/min), and CKD stage 5 (on hemodialysis). Muscle biopsies from patients with CKD stage 5 revealed lower mitochondrial volume density, lower mtDNA copy number, and higher BNIP3 content than controls. mtDNA copy number in PBMCs was decreased with increasing severity of CKD: non-CKD (6.48, 95% CI 4.49-8.46), CKD stage 3-4 (3.30, 95% CI 0.85-5.75, P = 0.048 vs. non-CKD), and CKD stage 5 (1.93, 95% CI 0.27-3.59, P = 0.001 vs. non-CKD). Isofurans were higher in patients with CKD stage 5 (median 59.21 pg/mL, IQR 41.76-95.36) compared to patients with non-CKD (median 49.95 pg/mL, IQR 27.88-83.46, P = 0.001), whereas F2-isoprostanes did not differ among groups. Severity of CKD is associated with mitochondrial dysfunction and markers of oxidative stress. Mitochondrial abnormalities, which are common in skeletal muscle from patients with CKD stage 5, may explain the muscle dysfunction associated with frailty and sarcopenia in CKD Further studies are required to evaluate mitochondrial function in vivo in patients with different CKD stages.

Cardiolipin fatty acid remodeling regulates mitochondrial function by modifying the electron entry point in the respiratory chain.

Modifications of cardiolipin (CL) levels or compositions are associated with changes in mitochondrial function in a wide range of pathologies. We have made the discovery that acetaminophen remodels CL fatty acids composition from tetralinoleoyl to linoleoyltrioleoyl-CL, a remodeling that is associated with decreased mitochondrial respiration. Our data show that CL remodeling causes a shift in electron entry from complex II to the ß-oxidation electron transfer flavoprotein quinone oxidoreductase (ETF/QOR) pathway. These data demonstrate that electron entry in the respiratory chain is regulated by CL fatty acid composition and provide proof-of-concept that pharmacological intervention can be used to modify CL composition.

Effect of Drug Therapy on Net Cholesterol Efflux Capacity of High-Density Lipoprotein-Enriched Serum in Rheumatoid Arthritis.

OBJECTIVE: Patients with rheumatoid arthritis (RA) have an increased risk of coronary heart disease (CHD). Some RA therapies may modify this risk, but the underlying mechanisms are unclear. The cholesterol efflux capacity of high-density lipoprotein (HDL) is associated with a reduced CHD risk in non-RA populations; however, inflammation may impair the function of HDL. The aim of this study was to evaluate whether reduced inflammation resulting from treatment with methotrexate (MTX), adalimumab (ADA), or tocilizumab (TCZ) would increase the net cholesterol efflux capacity of HDL in patients with RA. METHODS: A longitudinal multicenter study repository (Treatment Efficacy and Toxicity in Rheumatoid Arthritis Database and Repository) provided clinical information for and serum samples from 70 patients with RA before and 6 months after starting treatment with a new drug (MTX [n = 23], ADA [n = 22], or TCZ [n = 25]). Disease activity was measured using the Disease Activity Score in 28 joints using the erythrocyte sedimentation rate (DAS28-ESR). The net cholesterol efflux capacity was measured in paired serum samples using THP-1 macrophages, and total cellular cholesterol was measured by fluorometric assay. RESULTS: The DAS28-ESR decreased with all treatments (P < 0.001). Net cholesterol efflux capacity was not significantly changed after 6 months of new RA therapy (mean ± SD 36.9 ± 17.3% units at baseline versus 38.0% ± 16.9% units at 6 months [P = 0.58]). However, change in net cholesterol efflux capacity was associated with change in the DAS28-ESR (ρ = -0.25, P = 0.04). In a post hoc analysis of patients with impaired net cholesterol efflux capacity at baseline, treatment with TCZ resulted in significant improvement in net cholesterol efflux capacity (21.9 ± 14.7% units at baseline versus 31.3% ± 12.8% units at 6 months [P < 0.02]), but this was not observed with MTX or ADA. CONCLUSION: Net cholesterol efflux capacity of HDL cholesterol did not change significantly after 6 months of new RA therapy, except in patients with impaired baseline cholesterol efflux capacity who were receiving TCZ. Change in disease activity was associated with change in the net cholesterol efflux capacity.

The relationship between plasma lipid peroxidation products and primary graft dysfunction after lung transplantation is modified by donor smoking and reperfusion hyperoxia.

BACKGROUND: Donor smoking history and higher fraction of inspired oxygen (FIO2) at reperfusion are associated with primary graft dysfunction (PGD) after lung transplantation. We hypothesized that oxidative injury biomarkers would be elevated in PGD, with higher levels associated with donor exposure to cigarette smoke and recipient hyperoxia at reperfusion. METHODS: We performed a nested case-control study of 72 lung transplant recipients from the Lung Transplant Outcomes Group cohort. Using mass spectroscopy, F2-isoprostanes and isofurans were measured in plasma collected after transplantation. Cases were defined in 2 ways: grade 3 PGD present at day 2 or day 3 after reperfusion (severe PGD) or any grade 3 PGD (any PGD). RESULTS: There were 31 severe PGD cases with 41 controls and 35 any PGD cases with 37 controls. Plasma F2-isoprostane levels were higher in severe PGD cases compared with controls (28.6 pg/ml vs 19.8 pg/ml, p = 0.03). Plasma F2-isoprostane levels were higher in severe PGD cases compared with controls (29.6 pg/ml vs 19.0 pg/ml, p = 0.03) among patients reperfused with FIO2 >40%. Among recipients of lungs from donors with smoke exposure, plasma F2-isoprostane (38.2 pg/ml vs 22.5 pg/ml, p = 0.046) and isofuran (66.9 pg/ml vs 34.6 pg/ml, p = 0.046) levels were higher in severe PGD compared with control subjects. CONCLUSIONS: Plasma levels of lipid peroxidation products are higher in patients with severe PGD, in recipients of lungs from donors with smoke exposure, and in recipients exposed to higher Fio2 at reperfusion. Oxidative injury is an important mechanism of PGD and may be magnified by donor exposure to cigarette smoke and hyperoxia at reperfusion.

Net cholesterol efflux capacity of HDL enriched serum and coronary atherosclerosis in rheumatoid arthritis.

BACKGROUND/OBJECTIVES: Cardiovascular (CV) risk is increased in patients with rheumatoid arthritis (RA), but not fully explained by traditional risk factors such as LDL and HDL cholesterol concentrations. The cholesterol efflux capacity of HDL may be a better CV risk predictor than HDL concentrations. We hypothesized that HDL's cholesterol efflux capacity is impaired and inversely associated with coronary atherosclerosis in patients with RA. METHODS: We measured the net cholesterol efflux capacity of apolipoprotein B depleted serum and coronary artery calcium score in 134 patients with RA and 76 control subjects, frequency-matched for age, race and sex. The relationship between net cholesterol efflux capacity and coronary artery calcium score and other clinical variables of interest was assessed in patients with RA. RESULTS: Net cholesterol efflux capacity was similar among RA (median [IQR]: 34% removal [28, 41%]) and control subjects (35% removal [27%, 39%]) (P=0.73). In RA, increasing net cholesterol efflux capacity was not significantly associated with decreased coronary calcium score (OR=0.78 (95% CI 0.51-1.19), P=0.24, adjusted for age, race and sex, Framingham risk score and presence of diabetes). Net cholesterol efflux capacity was not significantly associated with RA disease activity score, C-reactive protein, urinary F2-isoprostanes, or degree of insulin resistance in RA. CONCLUSIONS: Net cholesterol efflux capacity is not significantly altered in patients with relatively well-controlled RA nor is it significantly associated with coronary artery calcium score.

Immune activation caused by vascular oxidation promotes fibrosis and hypertension.

Vascular oxidative injury accompanies many common conditions associated with hypertension. In the present study, we employed mouse models with excessive vascular production of ROS (tg(sm/p22phox) mice, which overexpress the NADPH oxidase subunit p22(phox) in smooth muscle, and mice with vascular-specific deletion of extracellular SOD) and have shown that these animals develop vascular collagen deposition, aortic stiffening, renal dysfunction, and hypertension with age. T cells from tg(sm/p22phox) mice produced high levels of IL-17A and IFN-γ. Crossing tg(sm/p22phox) mice with lymphocyte-deficient Rag1(-/-) mice eliminated vascular inflammation, aortic stiffening, renal dysfunction, and hypertension; however, adoptive transfer of T cells restored these processes. Isoketal-protein adducts, which are immunogenic, were increased in aortas, DCs, and macrophages of tg(sm/p22phox) mice. Autologous pulsing with tg(sm/p22phox) aortic homogenates promoted DCs of tg(sm/p22phox) mice to stimulate T cell proliferation and production of IFN-γ, IL-17A, and TNF-α. Treatment with the superoxide scavenger tempol or the isoketal scavenger 2-hydroxybenzylamine (2-HOBA) normalized blood pressure; prevented vascular inflammation, aortic stiffening, and hypertension; and prevented DC and T cell activation. Moreover, in human aortas, the aortic content of isoketal adducts correlated with fibrosis and inflammation severity. Together, these results define a pathway linking vascular oxidant stress to immune activation and aortic stiffening and provide insight into the systemic inflammation encountered in common vascular diseases.