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PurposeRheumatoid arthritis (RA) is associated with a higher risk of diabetes mellitus (DM). Our aim was to determine associations between inflammatory disease activity (including evaluation of specific cytokines and chemokines) and incident DM.MethodsParticipants were adults with physician-confirmed RA from Veteran’s Affairs Rheumatoid Arthritis Registry. Disease activity and clinical assessments occur longitudinally as part of clinical care. Thirty cytokines and chemokines were measured in banked serum obtained at the time of enrolment. Cytokine/chemokine values were log-adjusted and standardised (per SD). Incident DM was defined based on validated algorithms using diagnostic codes and medications. Multivariable Cox proportional hazard models evaluated associations between clinical factors and incident DM. Independent associations between cytokines/chemokines and incident DM were assessed adjusting for age, sex, race, smoking, body mass index (BMI) and medication use at baseline.ResultsAmong 1866 patients with RA without prevalent DM at enrolment, there were 130 incident cases over 9223 person-years of follow-up. High Disease Activity Score (DAS28)-C reactive protein (CRP), obese BMI, older age and male sex were associated with greater risk for incident DM while current smoking and methotrexate use were protective. Patients using methotrexate were at lower risk. Several cytokines/chemokines evaluated were independently associated (per 1 SD) with DM incidence including interleukin(IL)-1, IL-6 and select macrophage-derived cytokines/chemokines (HR range 1.11–1.26). These associations were independent of the DAS28-CRP.ConclusionsHigher disease activity and elevated levels of cytokines/chemokines are associated with a higher risk of incident DM in patients with RA. Future study may help to determine if targeted treatments in at-risk individuals could prevent the development of DM.

Background/objective Cytokines and chemokines (cytokines) are central to rheumatoid arthritis (RA) pathogenesis, with increasing use of multiplex immunoassays in clinical/research settings. Rheumatoid factor (RF) may interfere with assay outcomes by nonspecifically binding detection analytes. We evaluated the performance of a commercially available multiplex platform, including assessment of the impact of RF depletion. Methods Forty-five cytokines were tested using Meso Scale Discovery V-PLEX™ and samples from 40 RA and 40 osteoarthritis (OA) patients. Select samples were depleted of RF using a commercial binder. Performance was assessed using intra-assay coefficients of variation (CV), intraclass correlation coefficients (ICC), percent change following RF depletion, and disease discrimination. Values above or below quantification thresholds were imputed. Results Of the 45 cytokines analyzed, 31 yielded CVs 30%. ICCs universally exceeded 0.85 with the exception of eight analytes. RF depletion altered cytokine values by 30%) only seen for one analyte. Twenty-three cytokines differed significantly based on measurement in plasma vs. serum. Three analytes were higher in the serum of RA vs. OA (IL-10, IP-10, TNFα), and none were significantly greater in OA vs. RA. Seventeen analytes required imputation for >50% of the samples tested, primarily related to concentrations below the lower limit of quantification threshold. Conclusion The results from this commercially available multiplex assay were generally highly reproducible and interference induced by RF only meaningfully impacted the quantification of five of the analytes examined.

Background/Objective Malondialdehyde-acetaldehyde adducts (MAA) act as potent immune adjuvants and co-localize with citrullinated antigens in tissues effected by rheumatoid arthritis (RA). We sought to examine the role of MAA-adducts in promoting RA-related autoimmunity and inflammation. Methods DBA/J1 mice were immunized with human serum albumin (HSA), HSA-MAA, citrullinated HSA (HSA-Cit), or HSA-MAA-Cit with subsequent measurement of serum anti-citrullinated protein antibody (ACPA) and anti-Cit T cell responses. Cellular binding of the same antigens was examined using THP-1 monocytes and Chinese Hamster Ovary (CHO) cells transfected with specific scavenger receptors (SRs: TLR4, SR-B2, SREC-1). The effects of these antigens on THP-1 activation were then examined by quantifying plate adherence, pro-inflammatory (TNFα, IL-1β, IL-10) cytokine release, and SR (CD14, SR-B2)/co-stimulatory molecule (CD80, HLA-DR) expression. Comparisons were completed using one-way ANOVA with Tukey’s post-hoc test. Results Mice immunized with co-modified HSA produced significantly higher ACPA concentrations than all other groups whereas T cell responses to citrullinated proteins were highest following immunization with HSA-MAA. Both transfected CHO and THP-1 cells demonstrated significantly higher binding of HSA-MAA-Cit vs. HSA or HSA-Cit. THP-1 cells exposed to HSA-MAA-Cit expressed significantly higher concentrations of TNFα, IL-1β, and IL-10 vs. all other groups. Furthermore, THP-1 cells demonstrated significantly increased plate adherence and higher expression of CD14, SR-B2, and HLA-DR following incubation with HSA-MAA-Cit vs. HSA or HSA-Cit. Conclusion These studies demonstrate that MAA-adduction of citrullinated antigen greatly enhances immune and cellular responses, potentially acting as a key co-factor in RA pathogenesis.

BACKGROUND: Fracture healing in alcoholics is delayed and often associated with infections resulting in prolonged rehabilitation. It has been reported that binge drinking of alcohol increases oxidative stress and delays fracture healing in rats, which is prevented by treatment with the antioxidant n-acetyl cysteine (NAC). Oxidative stress is a significant factor in pathologies of various organs resulting from chronic alcoholism. Therefore, we hypothesize that treatment with NAC reduces oxidative stress and restores fracture healing in chronic alcoholics. METHODS: Rats (10 months old) were pair-fed the Lieber-DeCarli ethanol (EtOH) diet or control diet for 16 weeks. A closed fracture was performed and rats allowed to recover for 72 hours. Rats were divided into 4 groups—control, control + NAC, EtOH, and EtOH + NAC—and injected intraperitoneally with 200 mg/kg of NAC daily for 3 days. Serum and bone fracture callus homogenates were collected and assayed for traditional markers of inflammation, oxidative stress, and bone regeneration. RESULTS: The oxidative stress marker malondialdehyde (MDA) was increased in both serum and bone tissue in EtOH-fed animals compared to controls. NAC treatment significantly (p < 0.01) reduced MDA to near normal levels and dramatically increased the index of antioxidant efficacy (catalase/MDA ratio) (p < 0.01). Inflammatory markers tumor necrosis factor-α, interferon-γ, and interleukin-6 were significantly decreased in serum and callus following NAC treatment. NAC treatment reduced EtOH-induced bone resorption as evidenced by significant decreases in C-telopeptide of type-I-collagen levels (p < 0.05) and band-5 tartrate-resistant acid phosphatase levels in the tissue (p < 0.001). CONCLUSIONS: Oxidative stress and excessive inflammation are involved in the inhibition of fracture healing by EtOH. In this study, early short-term treatment of EtOH-fed animals with the antioxidant NAC reduced oxidative stress and normalized the innate immune response to fracture in the early phase of fracture healing, thereby restoring the normal onset of bone regeneration.

Airborne biohazards are risk factors in the development and severity of rheumatoid arthritis (RA) and RA-associated lung disease, yet the mechanisms explaining this relationship remain unclear. Lipopolysaccharide (LPS, endotoxin) is a ubiquitous inflammatory agent in numerous environmental and occupational air pollutant settings recognized to induce airway inflammation. Combining repetitive LPS inhalation exposures with the collagen induced arthritis (CIA) model, DBA1/J mice were assigned to either: sham (saline injection/saline inhalation), CIA (CIA/saline), LPS (saline/LPS 100 ng inhalation), or CIA + LPS for 5 weeks. Serum anti-citrullinated (CIT) protein antibody (ACPA) and anti-malondialdehyde-acetaldehyde (MAA) antibodies were strikingly potentiated with co-exposure (CIA + LPS). CIT- and MAA-modified lung proteins were increased with co-exposure and co-localized across treatment groups. Inhaled LPS exacerbated arthritis with CIA + LPS > LPS > CIA versus sham. Periarticular bone loss was demonstrated in CIA and CIA + LPS but not in LPS alone. LPS induced airway inflammation and neutrophil infiltrates were reduced with co-exposure (CIA + LPS). Potentially signaling transition to pro-fibrotic processes, there were increased infiltrates of activated CD11c+CD11b+ macrophages and transitioning CD11c+CD11bint monocyte-macrophage populations with CIA + LPS. Moreover, several lung remodeling proteins including fibronectin and matrix metalloproteinases as well as complement C5a were potentiated with CIA + LPS compared to other treatment groups. IL-33 concentrations in lung homogenates were enhanced with CIA + LPS with IL-33 lung staining driven by LPS. IL-33 expression was also significantly increased in lung tissues from patients with RA-associated lung disease (N = 8) versus controls (N = 7). These findings suggest that patients with RA may be more susceptible to developing interstitial lung disease following airborne biohazard exposures enriched in LPS.

Background Malondialdehyde (MDA) and acetaldehyde (AA) exist following ethanol metabolism and tobacco pyrolysis. As such, lungs of individuals with alcohol use disorders (AUD) are a target for the effects of combined alcohol and cigarette smoke metabolites. MDA and AA form a stable protein adduct, malondialdehyde-acetaldehyde (MAA) adduct, known to be immunogenic, profibrotic, and proinflammatory. MAA adduct is the dominant epitope in anti-MAA antibody formation. We hypothesized that MAA-adducted protein forms in lungs of those who both abuse alcohol and smoke cigarettes, and that this would be associated with systemically elevated anti-MAA antibodies Methods Four groups were established: AUD subjects who smoked cigarettes (+AUD/+smoke), smokers without AUD (-AUD/+smoke), AUD without smoke (+AUD/-smoke), and non-AUD/nonsmokers (-AUD/-smoke) Results We observed a significant increase in MAA adducts in lung cells of +AUD/+smoke vs. -AUD/-smoke. No significant increase in MAA adducts was observed in -AUD/+smoke or in +AUD/-smoke compared to -AUD/-smoke. Serum from +AUD/+smoke had significantly increased levels of circulating anti-MAA IgA antibodies. After one week of alcohol that MAA-adducted protein is formed in the lungs of those who smoke cigarettes and abuse alcohol, leading to a subsequent increase in serum IgA antibodies. MAA-adducted proteins could play a role in pneumonia and other diseases of the lung in the setting of AUD and smoking. This article is protected by copyright. All rights reserved.

Methotrexate (MTX) is an immunosuppressant commonly used for the treatment of autoimmune diseases. Recent observations have shown that patients treated with MTX also exhibit a reduced risk for the development of cardiovascular disease (CVD). Although MTX reduces systemic inflammation and tissue damage, the mechanisms by which MTX exerts these beneficial effects are not entirely known. We have previously demonstrated that protein adducts formed by the interaction of malondialdehyde (MDA) and acetaldehyde (AA), known as MAA-protein adducts, are present in diseased tissues of individuals with rheumatoid arthritis (RA) or CVD. In previously reported studies, MAA-adducts were shown to be highly immunogenic, supporting the concept that MAA-adducts not only serve as markers of oxidative stress but may have a direct role in the pathogenesis of inflammatory diseases. Because MAA-adducts are commonly detected in diseased tissues and are proposed to mitigate disease progression in both RA and CVD, we tested the hypothesis that MTX inhibits the generation of MAA-protein adducts by scavenging reactive oxygen species. Using a cell free system, we found that MTX reduces MAA-adduct formation by approximately 6-fold, and scavenges free radicals produced during MAA-adduct formation. Further investigation revealed that MTX directly scavenges superoxide, but not hydrogen peroxide. Additionally, using the Nrf2/ARE luciferase reporter cell line, which responds to intracellular redox changes, we observed that MTX inhibits the activation of Nrf2 in cells treated with MDA and AA. These studies define previously unrecognized mechanisms by which MTX can reduce inflammation and subsequent tissue damage, namely, scavenging free radicals, reducing oxidative stress, and inhibiting MAA-adduct formation., Graphical abstract The production of reactive oxygen species (ROS) can result in lipid peroxidation. Lipid peroxidation can, in turn, result in the formation of malondialdehyde (MDA), which can spontaneously breakdown forming acetaldehyde (AA). Malondialdehyde and acetaldehyde can interact to form the stable hybrid malondialdehyde-acetaldehyde adduct (MAA) on proteins and other macromolecules. Binding of MAA-adducted proteins to cells can lead to expression of pro-inflammatory cytokines, resulting in inflammation. MAA-adducted proteins are also themselves immunogenic and can initiate an inflammatory response. Importantly, methotrexate can scavenge free radicals. This may ameliorate the formation of lipid peroxides and the resulting formation of MDA and AA. Additionally, methotrexate can directly inhibit the formation of MAA-protein adducts. Thus, methotrexate can scavenge free radicals and inhibit the formation of MAA-adducts. These are previously undescribed mechanisms by which methotrexate may reduce inflammation and the tissue damage associated with chronic inflammatory diseases.fx1, Highlights • MTX is commonly used to treat RA and is being tested in CVD patients. • MDA and AA are produced during lipidperoxidation and can interact to form MAA-adducts. • MAA-adducts are found in atheromas and in diseased synovial tissue of RA patients. • MTX scavenges the free radical O2− and prevents the formation of MAA-adducts. • Scavenging O2− may be a mechanism by which MTX reduces inflammation and disease.

Doxycycline (DOX), a derivative of tetracycline, is a broad-spectrum antibiotic that exhibits a number of therapeutic activities in addition to its antibacterial properties. For example, DOX has been used in the management of a number of diseases characterized by chronic inflammation. One potential mechanism by which DOX inhibits the progression of these diseases is by reducing oxidative stress, thereby inhibiting subsequent lipid peroxidation and inflammatory responses. Herein, we tested the hypothesis that DOX directly scavenges reactive oxygen species (ROS) and inhibits the formation of redox-mediated malondialdehyde-acetaldehyde (MAA) protein adducts. Using a cell-free system, we demonstrated that DOX scavenged reactive oxygen species (ROS) produced during the formation of MAA-adducts and inhibits the formation of MAA-protein adducts. To determine whether DOX scavenges specific ROS, we examined the ability of DOX to directly scavenge superoxide and hydrogen peroxide. Using electron paramagnetic resonance (EPR) spectroscopy, we found that DOX directly scavenged superoxide, but not hydrogen peroxide. Additionally, we found that DOX inhibits MAA-induced activation of Nrf2, a redox-sensitive transcription factor. Together, these findings demonstrate the under-recognized direct antioxidant property of DOX that may help to explain its therapeutic potential in the treatment of conditions characterized by chronic inflammation and increased oxidative stress.

Background Although biologics represent a major advance in rheumatoid arthritis (RA), many patients fail to achieve adequate responses to these agents. We examined whether combined positivity to three well-characterized autoantibodies predicts treatment response among RA patients initiating biologics. Methods The study included biologic-naive patients initiating anti-TNF treatment, biologic-exposed patients switching to rituximab or tocilizumab, and patients (biologic naive or exposed) initiating abatacept. Rheumatoid factor (RF), anti-cyclic citrullinated peptide (CCP) antibody, and IgG antibodies to malondialdehyde-acetaldehyde (MAA) were measured using banked enrollment serum. The relationship between the number of autoantibodies positive (0–3) and treatment response (absolute improvement in 28-joint Disease Activity Score [DAS28-CRP] or improvement > 1.2) at 6 months was examined using multivariable linear and logistic regression. Results Of 1,229 patients initiating biologics, 79% were women; 89% were Caucasian. The number of baseline RA-related autoantibodies positive was associated with improved treatment response in a dose-dependent fashion. Compared to patients seronegative for all autoantibodies, adjusting for covariates, those positive for all three were more than twice (OR 2.35; 95% CI 1.57–3.51) as likely to achieve DAS28 improvement > 1.2 units. Associations of autoantibody positivity with biologic treatment response were strongest for anti-CCP antibody, persisted in analyses limited to biologic naive patients, and did not appear to differ markedly among different agents examined. Conclusion An expanded autoantibody profile appears to significantly predict RA treatment response to biologic treatment in a dose-dependent fashion. Incorporating these serologic profiles with additional biomarkers or other informative patient characteristics could provide an opportunity to personalize RA management.

BACKGROUND: Circulating uric acid (UA) is an important biomarker, not only in the detection and management of gout, but also in assessing the risk of related comorbidity. The impact of collection methods on clinical UA measurements has been the subject of limited study. After observing significant differences between UA concentrations of blood samples obtained by different collection tubes, we began examining the effects of exogenous tube components on measured UA concentrations. We aimed to: (1) demonstrate the variability in uricase-based UA measurements attributable to different collection methods and (2) identify factors influencing this variability. METHODS: Blood samples from human subjects were collected using Serum Separator Tubes (SST tubes), Acid Citrate Dextrose (ACD) tubes, and Sodium Citrate (SC) tubes. Circulating UA concentrations were measured by chemistry analyzers utilizing the uricase method. Absorbance assays were run in order to determine the effects of citric acid, sodium citrate, and dextrose on measured absorbance in the presence of leuco crystal violet dye, hydrogen peroxide, and peroxidase. Statistical analyses—including Student’s T tests and ANOVA—were used to compare results. RESULTS: UA concentrations of blood samples collected in ACD tubes were significantly lower than those collected in SST tubes (P

Precision-cut liver slices (PCLSs) provide a novel model for studies of alcoholic liver disease (ALD). This is relevant, as in vivo ethanol exposure does not appear to generate significant liver damage in ethanol-fed mice, except in the National Institute on Alcohol Abuse and Alcoholism binge model of ALD. Previous studies have shown that the two metabolites of ethanol consumption, malondialdhyde (MDA) and acetaldehyde (AA), combine to form MDA-AA (MAA) adducts, which have been correlated with the development and progression of ALD. In this study, murine PCLSs were incubated with ethanol and examined for the production of MAA adducts. PCLSs were homogenized, and homogenates were injected into C57BL/6 mice. PCLSs from control-, pair-, and ethanol-fed animals served as targets in in situ cytotoxic assays using primed T cells from mice hyperimmunized with control or ethanol-exposed PCLS homogenates. A CD45.1/CD45.2 passive-transfer model was used to determine whether T cells from the spleens of mice hyperimmunized with PCLS ethanol-exposed homogenates trafficked to the liver. PCLSs incubated with ethanol generated MAA-modified proteins in situ. Cytotoxic (CD8+) T cells from immunized mice killed naïve PCLSs from control- and pair-fed mice in vitro, a response that was blunted in PCLSs from ethanol-fed mice. Furthermore, CD45.1 CD8+ T cells from hyperimmunized mice trafficked to the liver but did not initiate liver damage. This study demonstrates that exposure to liver tissue damaged by ethanol mediates robust immune responses to well-characterized alcohol metabolites and native liver proteins in vitro. Moreover, although these proinflammatory T cells traffic to the liver, these responses appear to be dampened in vivo by locally acting pathways. NEW & NOTEWORTHY This study shows that the metabolites of ethanol and lipid breakdown produce malondialdehyde-acetaldehyde adducts in the precision-cut liver slice model system. Additionally, precision-cut liver slices exposed to ethanol and harboring malondialdehyde-acetaldehyde adducts generate liver-specific antibody and T cell responses in the spleens of naïve mice that could traffic to the liver.

Objective To compare anti-malondialdehyde–acetaldehyde (MAA) antibody concentrations between rheumatoid arthritis (RA) patients and healthy and rheumatic disease controls. Methods Anti-MAA antibody (IgA, IgM, IgG) was measured using ELISA and banked serum from patients with RA (n = 284), osteoarthritis (OA, n = 330), spondyloarthropathy (SpA, n = 50), and systemic lupus erythematosus (SLE, n = 88) as well as healthy controls (n = 82). Anti-MAA antibody concentrations and the frequency of positivity were compared across groups. Multivariable linear regression analysis limited to RA and OA patients (due to sample size and data availability) was used to identify factors associated with anti-MAA antibody concentrations. Results Although RA patients demonstrated among the highest circulating concentrations across isotypes, only IgA anti-MAA antibody was significantly higher than all other groups (p ≤ 0.02). Proportions (7% to 74%) of OA and SLE (less so for SpA) samples were positive for anti-MAA antibody, limiting the discriminatory capacity of anti-MAA antibody in RA (positive in 18% to 80%). In analyses limited to those with RA or OA, factors associated with higher anti-MAA antibody concentrations included RA case status, younger age (IgM), male sex (IgG), African American race (IgA, IgG) and current smoking (IgA). C-reactive protein levels and comorbidities were not associated with anti-MAA antibody concentrations. Conclusion With the possible exception of the IgA isotype, serum anti-MAA antibodies measured with currently available assays do not appear to adequately discriminate RA from other rheumatic conditions. With the identification of specific proteins that are MAA-modified in diseased tissues and requisite assay refinement, anti-MAA antibody holds potential promise as a biomarker in RA.

Objective Malondialdehyde-acetaldehyde (MAA) adducts are a product of oxidative stress associated with tolerance loss in several disease states. This study was undertaken to investigate the presence of MAA adducts and circulating anti-MAA antibodies in patients with rheumatoid arthritis (RA). Methods Synovial tissue from patients with RA and patients with osteoarthritis (OA) were examined for the presence of MAA-modified and citrullinated proteins. Anti-MAA antibody isotypes were measured in RA patients (n = 1,720) and healthy controls (n = 80) by enzyme-linked immunosorbent assay. Antigen-specific anti–citrullinated protein antibodies (ACPAs) were measured in RA patients using a multiplex antigen array. Anti-MAA isotype concentrations were compared in a subset of RA patients (n = 80) and matched healthy controls (n = 80). Associations of anti-MAA antibody isotypes with disease characteristics, including ACPA positivity, were examined in all RA patients. Results Expression of MAA adducts was increased in RA synovial tissue compared to OA synovial tissue, and colocalization with citrullinated proteins was found. Increased levels of anti-MAA antibody isotypes were observed in RA patients compared to controls (P < 0.001). Among RA patients, anti-MAA antibody isotypes were associated with seropositivity for ACPAs and rheumatoid factor (P < 0.001) in addition to select measures of disease activity. Higher anti-MAA antibody concentrations were associated with a greater number of positive antigen-specific ACPA analytes (expressed at high titer) (P < 0.001) and a higher ACPA score (P < 0.001), independent of other covariates. Conclusion MAA adduct formation is increased in RA and appears to result in robust antibody responses that are strongly associated with ACPAs. These results support speculation that MAA formation may be a cofactor that drives tolerance loss, resulting in the autoimmune responses characteristic of RA.

Antibodies to malondialdehyde (MDA) modified macromolecules (adducts) have been detected in the serum of patients with atherosclerosis and correlate with the progression of this disease. However, the epitope and its formation have not been characterized. Studies have shown that excess MDA can be degraded to acetaldehyde which combines with proteins to from a stable dihydropyridine adduct. To investigate, mice were immunized with (MDA) adducts in the absence of adjuvant and showed an increase in antibodies to MDA adducts and the carrier protein as the concentration of MDA was increased. In fact, a number of the commercially available antibodies to MDA modified proteins were able to be inhibited by a chemical analogue hexyl-MAA. Also, MDA/MAA adducts were detected in the serum and aortic tissue of JCR diabetic/atherosclerotic rats. These studies determined that commercially available antibodies to MDA were shown to predominantly react with the MAA adduct and are present in the JCR model of atherosclerosis in both the serum and aortic tissue. Therefore, the immune response to MDA modified proteins is most likely to the dihydropyridine structure (predominant epitope in MAA), and suggests that MAA adducts may be playing a role in the development and/or progression of atherosclerosis.

Background & Aims Aldehydes that are produced following the breakdown of ethanol (acetaldehyde) and lipid peroxidation of membranes (malondialdehyde) have been shown to bind (adduct) proteins. Additionally, these two aldehydes can combine (MAA) on non-syngeneic and syngeneic proteins to initiate numerous immune responses to the unmodified part of the protein in the absence of an adjuvant. Therefore, these studies provide a potential mechanism for the development of antigen-specific immune responses resulting in liver damage should syngeneic liver proteins be adducted with MAA.

Alcohol abuse results in liver injury, but investigations into the mechanism(s) for this injury have been hampered by the lack of appropriate in vitro culture models in which to conduct in depth and specific studies. In order to overcome these shortcomings, we have developed the use of precision-cut liver slices (PCLS) as an in vitro culture model in which to investigate how ethanol causes alcohol-induced liver injury. In these studies, it was shown that the PCLS retained excellent viability as determined by lactate dehydrogenase and adenosine triphosphate (ATP) levels over a 96-h period of incubation. More importantly, the major enzymes of ethanol detoxification; alcohol dehydrogenase, aldehyde dehydrogenase, and cytochrome P4S02E1, remained active and PCLS readily metabolized ethanol and produced acetaldehyde. Within 24 h and continuing up to 96 h the PCLS developed fatty livers and demonstrated an increase in the redox state. These PCLS secreted albumin, and albumin secretion was decreased by ethanol treatment. All of these impairments were reversed following the addition of 4-methylpyrazole, which is an inhibitor of ethanol metabolism. Therefore, this model system appears to mimic the ethanol-induced changes in the liver that have been previously reported in human and animal studies, and may be a useful model for the study of alcoholic liver disease. (C) 2008 Elsevier Inc. All rights reserved.

Malondialdehyde-acetaldehyde adducts (MAA) have been implicated in atherosclerosis. The purpose of this study was to investigate the role of MAA in atherosclerotic disease. Serum samples from controls (n = 82) and patients with; non-obstructive coronary artery disease (CAD), (n = 40), acute myocardial infarction (AMI) (n = 42), or coronary artery bypass graft (CABG) surgery due to obstructive multi-vessel CAD (n = 72), were collected and tested for antibody isotypes to MAA-modifed human serum albumin (MAA-HSA). CAD patients had elevated relative levels of IgG and IgA anti-MAA, compared to control patients (p

article Introduction: Citrullinated self-proteins are thought to be involved in the onset/progression of rheumatoid arthritis (RA). Numerous studies have been performed to look for the self-antigen that becomes citrullinated and induces RA. Importantly, these studies have been performed using citrullinated self-antigens injected into an animal model in the presence of a strong adjuvant in order to derive the response. However, to date no studies have been performed to determine if these phenotypes can be induced in the absence of an adjuvant. Methods: To investigate this possibility, mice were immunized with citrullinated or non-citrullinated mouse Type II collagen (Cit-Col or Col) in the presence or absence of Freund's Complete Adjuvant (FCA). Results: An autoimmune-like RA response was observed in mice immunized with Cit-Col in the absence of FCA; by the increase in caliper score, visual observation, and micro-CT analysis of bone erosions. Antibody and T-cell responses were increased in the Cit-Col injected mice to Cit-Col as well as antibody to Anti- Citrullinated Peptide Antigens (ACPA) as determined by a commercially available test kit. Conclusions: Therefore, the use of citrullinated mouse collagen induces an autoimmune-like RA in the absence of an adjuvant. These data also suggest that citrullinate self-proteins may be potential molecular adjuvants that assist in driving an inflammatory response, that increases the production of PAD in joint tissue, resulting in the citrullination of other self-proteins to exacerbate the disease.

The onset of alcoholic liver disease (ALD) is initiated by different cell types in the liver and a number of different factors including: products derived from ethanol-induced inflammation, ethanol metabolites, and the indirect reactions from those metabolites. Ethanol oxidation results in the production of metabolites that have been shown to bind and form protein adducts, and to increase inflammatory, fibrotic and cirrhotic responses. Lipopolysaccharide (LPS) has many deleterious effects and plays a significant role in a number of disease processes by increasing inflammatory cytokine release. In ALD, LPS is thought to be derived from a breakdown in the intestinal wall enabling LPS from resident gut bacterial cell walls to leak into the blood stream. The ability of adducts and LPS to independently stimulate the various cells of the liver provides for a two-hit mechanism by which various biological responses are induced and result in liver injury. Therefore, the purpose of this article is to evaluate the effects of a two-hit combination of ethanol metabolites and LPS on the cells of the liver to increase inflammation and fibrosis, and play a role in the development and/or progression of ALD.

Purpose Inhaled milrinone (MIL) has been used empirically to reduce pulmonary vascular resistance and improve right ventricular dysfunction without systemic adverse effects; however characterization of this process is not reported in the literature. We sought to characterize aerosolized MIL particle size, nebulization degradation profile, and quantification of drug delivery using a vibrating mesh nebulizer connected to a mechanical ventilator circuit in preparation for a human heart failure clinical trial. Methods and Materials Mass Spectrometry was performed on nebulized MIL samples and saline controls. A pharmaceutical grade impactor was used for aerodynamic particle size studies. High pressure liquid chromatography (HPLC) was used to determine final mass median aerodynamic particle size (MMAD), geometric standard deviation (GSD), and fine particle fraction (FPF). Nebulized MIL recovered at the end of an endotracheal tube (ETT) in a mechanical ventilator circuit was quantified using HPLC. All studies were performed in triplicate. Results Mass Spectrometry did not reveal MIL degradation products or formation of new drug compounds. Aerodynamic MMAD for nebulized MIL followed a Gaussian curve distribution. [ figure 1 ] MMAD = 4.22 μm with GSD = 2.29 μm and FPF = 0.62 +/− 0.7. 22.9% of nebulized MIL was recovered at the end of the ETT. Conclusions Nebulizing MIL using a vibrating mesh nebulizer attached to a mechanical ventilator circuit is feasible, results in delivery of a consistent unaltered drug product, and generates a respirable aerosol with size properties ideal for both pulmonary and systemic drug delivery.

Meeting abstracts High-fat diets and chronic alcohol consumption (CAC) can both induce a low-grade inflammation. We report that when CAC (16.6% of total calories) is administered in combination with the Lieber-DeCarli high-fat diet an additive myeloid response is induced. This increase in