Analysis of the pancreatic low molecular weight proteome in an animal model of acute pancreatitis

We used a peptidomic approach for the analysis of the low molecular weight proteome in rat pancreatic tissue extracts. The goal was to develop a method that allows identifying endogenous peptides produced in the pancreas in the course of acute pancreatitis. The workflow combines peptides enrichment by centrifugal ultrafiltration, fractionation by isoelectric focusing, and LC-MS/MS analysis without prior enzymatic digestion. The method was assessed on pancreatic extracts from 3 rats with caerulein-induced pancreatitis and 3 healthy controls. A qualitative analysis of the peptide patterns obtained from the different samples was performed to determine the main biological processes associated to the identified peptides. Comparison of peptidomic and immunoblot data for alpha-tubulin, beta-tubulin and coatomer gamma showed that the correlation between the number of identified peptides and the protein abundance was variable. Nevertheless, peptidomic analysis highlighted inflammatory and stress proteins, which peptide pattern was related to acute pancreatitis pathobiology. For these proteins, the higher number of peptides in pancreatitis samples reflected an increase in protein abundance. Moreover, for murinoglobulin-1 or carboxypeptidase B, peptide pattern could be related to protein function. These data suggest that peptidomic analysis is a complementary approach to proteomics for investigating pathobiological processes involved in acute pancreatitis

Proteomic analysis of heat shock-induced protection in acute pancreatitis

Acute pancreatitis is an inflammatory disease of the pancreas, which can result in serious morbidity or death. Acute pancreatitis severity can be reduced in experimental models by preconditioning animals with a short hyperthermia prior to disease induction. Heat shock proteins 27 and 70 are key effectors of this protective effect. In this study, we performed a comparative proteomic analysis using a combination of liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis and isobaric tagging to investigate changes in pancreatic proteins expression that were associated with thermal stress, both in healthy rats and in a model of caerulein-induced pancreatitis. In agreement with previous studies, we observed modulation of heat shock and inflammatory proteins expression in response to heat stress or pancreatitis induction. We also identified numerous other proteins, whose pancreatic level changed following pancreatitis induction, when acute pancreatitis severity was reduced by prior thermal stress, or in healthy rats in response to hyperthermia. Interestingly, we showed that the expression of various proteins associated with the secretory pathway was modified in the different experimental models, suggesting that modulation of this process is involved in the protective effect against pancreatic tissue damage

P5343 - The presence of anti-apolipoprotein A1 autoantibodies is associated with a pro-atherogenic profile in HIV-infected patients

Background: With the access to antiretroviral therapy (ART), the mortality related to the human immunodeficiency virus (HIV) has dropped, shifting the clinical challenges towards chronic disease management, including cardiovascular disease (CVD) risk assessment. Factors that potentially contribute to the physiopathology of HIV-related CVD include the HI-virus itself, adverse effects of ART, and processes such as dyslipidemia, inflammation, immune/autoimmune activation and endothelial injury. Among autoantibodies of possible cardiovascular relevance, those directed against apolipoprotein A-1 (anti-apoA-1 IgG) were shown to predict major adverse cardiovascular events and promote atherogenesis. Purpose: The aim of the study was to evaluate the prevalence of anti-apoA1 IgG in HIV-free and ART experienced and naïve HIV-infected patients as well as the association between anti-apoA1 IgG levels and, indices of viral suppression, clinical parameters (10 year Framingham Risk Score (FRS)) and inflammatory biomarkers, known to underlie atherosclerosis burden in these patients. Methods: Anti-apoA1 IgG serum levels were assessed by a homemade ELISA assay in 144 participants from a South African cohort divided in three groups: HIV-free (n=50), HIV-infected/ART experienced (n=50) and HIV-infected/ART naïve (n=44). Inflammatory biomarkers were measured. Results: HIV-infected patients displayed an increased pro-atherogenic biomarker profile compared to HIV-free subjects, but not difference in the FRS was observed between these two groups. Regarding anti-apoA1 IgG, 24% of HIV-free patients tested positive compared to 40% and 70% in HIV-infected/ART experienced and naïve groups, respectively. HIV-infected, anti-apoA1 IgG positive patients showed a significant decrease in CD4+ counts (p=0.003) and a significant increase in viremia (p=0.0130), mean heart rate (p=0.0243), albuminuria (p=0.0155), pro-inflammatory biomarkers (IFNγ, IL-10, TNFα, MIPα; all p&lt;0.05), circulating levels of intercellular adhesion molecule (ICAM-1) (p=0.0217) and vascular cell adhesion molecule (VCAM-1) (p=0.003) compared to anti-apoA1 IgG negative ones. Of note, while this profile was maintained in HIV-infected/ART experienced, these significant differences were lost in HIV-infected/ART naïve patients. No significant difference in FRS was observed between anti-apoA1 IgG positive vs negative individuals in all groups. Conclusions: HIV-infected patients presented with an increased prevalence of anti-apoA1 IgG compared to HIV-free subjects. In HIV-infected/ART experienced patients, anti-apoA1 IgG levels were associated with low CD4+ counts, levels of adhesion molecules and pro-inflammatory responses, features associated with increased cardiovascular events. ART highlighted pro-atherogenic differences between HIV-infected anti-apoA1 IgG negative and positive patients.</p

People living with HIV display increased anti-apolipoprotein A1 auto-antibodies, inflammation, and kynurenine metabolites : a case–control study

Objective This study aimed to study the relationship between auto-antibodies against apolipoprotein A1 (anti-apoA1 IgG), human immunodeficiency virus (HIV) infection, anti-retroviral therapy (ART), and the tryptophan pathways in HIV-related cardiovascular disease. Design This case–control study conducted in South Africa consisted of control volunteers ( n  = 50), people living with HIV (PLWH) on ART ( n  = 50), and untreated PLWH ( n  = 44). Cardiovascular risk scores were determined, vascular measures were performed, and an extensive biochemical characterisation (routine, metabolomic, and inflammatory systemic profiles) was performed. Methods Anti-apoA1 IgG levels were assessed by an in-house ELISA. Inflammatory biomarkers were measured with the Meso Scale Discovery® platform, and kynurenine pathway metabolites were assessed using targeted metabolomic profiling conducted by liquid chromatography-multiple reaction monitoring/mass spectrometry (LC-MRM/MS). Results Cardiovascular risk scores and vascular measures exhibited similarities across the three groups, while important differences were observed in systemic inflammatory and tryptophan pathways. Anti-apoA1 IgG seropositivity rates were 15%, 40%, and 70% in control volunteers, PLWH ART-treated, and PLWH ART-naïve, respectively. Circulating anti-apoA1 IgG levels were significantly negatively associated with CD4+ cell counts and positively associated with viremia and pro-inflammatory biomarkers (IFNγ, TNFα, MIPα, ICAM-1, VCAM-1). While circulating anti-apoA1 IgG levels were associated with increased levels of kynurenine in both control volunteers and PLWH, the kynurenine/tryptophan ratio was significantly increased in PLWH ART-treated. Conclusion HIV infection increases the humoral response against apoA1, which is associated with established HIV severity criteria and kynurenine pathway activation.</p

Antiapolipoprotein A-1 Autoantibody Positivity Is Associated with Threatened Abortion

Background . Autoantibodies against apolipoprotein A-1 (anti-ApoA-1 IgG) were demonstrated to be associated with cardiovascular outcomes in several inflammatory diseases. As balanced inflammation is critical for uncomplicated pregnancy, we aimed to investigate the prevalence of anti-ApoA-1 IgG and anti-c-terminal ApoA-1 autoantibodies (Ac-terAA1 IgG) in a cohort of pregnant women and their potential relationship with threatened abortion (TA). Methods . Between 2012 and 2014, 371 consecutive outpatient pregnant women were included in this study and followed until delivery. Anti-ApoA-1 and anti-Ac-terAA1 IgG were measured by ELISA technique on serum samples collected between the 24 th and 26 th week of pregnancy. Associations with TA were tested using linear regression analysis and C-statistics. Results . Median age was 34 with a prevalence of the Caucasian ethnicity (90.5%). TA occurred in 10 women (2.7%). C-statistics indicated that anti-ApoA-1 and anti-Ac-terAA1 IgG levels upon study inclusion were predictive of TA (0.73, 95% confidence interval [CI] 0.69-0.78, p &lt; 0.001 and 0.76, 95% CI 0.71-0.80, p = 0.01 , respectively). At the prespecified anti-ApoA-1 IgG cutoff, the negative predictive value (NPV) was 100%. For anti-Ac-terAA1 IgG, at the optimal cutoff, the NPV was 99%. Linear regression models indicated that risk associations were independent of age and the presence of autoimmune diseases for both autoantibodies ( p &lt; 0.001 ). Anti-Ac-terAA1 IgG-positive individuals were more frequently non-Caucasians ( p = 0.009 ). Conclusion . Anti-ApoA-1 and anti-Ac-terAA1 IgG are independently associated with TA during pregnancy with an appealing NPV. The causal biological mechanisms underlying this association as well as the possible clinical relevance of these findings require further investigations. </p