Mediator Profiles Research Articles

Nutritional Lipids and Mucosal Inflammation.

Inflammatory bowel disease (IBD) is characterized by chronic relapsing inflammation in the intestine. Given their role in regulation of inflammation, long-chain n-3 polyunsaturated fatty acids (PUFAs) represent a potential supplementary therapeutic approach to current drug regimens used for IBD. Mechanistically, there is ample evidence for an anti-inflammatory and pro-resolution effect of long-chain n-3 PUFAs after they incorporate into cell membrane phospholipids. They disrupt membrane rafts and when released from the membrane suppress inflammatory signaling by activating PPAR-γ and free fatty acid receptor 4; furthermore, they shift the lipid mediator profile from pro-inflammatory eicosanoids to specialized pro-resolving mediators. The allocation of long-chain n-3 PUFAs also leads to a higher microbiome diversity in the gut, increases short-chain fatty acid-producing bacteria, and improves intestinal barrier function by sealing epithelial tight junctions. In line with these mechanistic studies, most epidemiological studies support a beneficial effect of long-chain n-3 PUFAs intake on reducing the incidence of IBD. However, the results from intervention trials on the prevention of relapse in IBD patients show no or only a marginal effect of long-chain n-3 PUFAs supplementation. In light of the current literature, international recommendations are supported that adequate diet-derived n-3 PUFAs might be beneficial in maintaining remission in IBD patients.

Heterogeneity of lower airway inflammation in patients with NSAID-exacerbated respiratory disease

Nonsteroidal anti-inflammatory drug (NSAID)-exacerbated respiratory disease (N-ERD) asthma is characterized by chronic rhinosinusitis and intolerance of aspirin and other COX1 inhibitors. Clinical data point to a heterogeneity within the N-ERD phenotype. Our aim was to investigate immune mediator profiles in the lower airways of patients with N-ERD. Levels of cytokines (determined by using Luminex assay) and eicosanoids (determined by using mass spectrometry) were measured in bronchoalveolar lavage fluid (BALF) from patients with N-ERD (n= 22), patients with NSAID-tolerant asthma (n= 21), and control subjects (n= 11). mRNA expression in BALF cells was quantified by using TaqMan low-density arrays. Lower airway eosinophilia was more frequent in N-ERD (54.5%) than in NSAID-tolerant asthma (9.5% [P= .009]). The type-2 (T2) immune signature of BALF cells was more pronounced in the eosinophilic subphenotype of N-ERD. Similarly, BALF concentrations of periostin and CCL26 were significantly increased in eosinophilic N-ERD and correlated with T2 signature in BALF cells. Multiparameter analysis of BALF mediators of all patients with asthma revealed the presence of 2 immune endotypes: T2-like (with an elevated level of periostin in BALF) and non-T2/proinflammatory (with higher levels of matrix metalloproteinases and inflammatory cytokines). Patients with N-ERD were classified mostly as having the T2 endotype (68%). Changes in eicosanoid profile (eg, increased leukotriene E4 level) were limited to patients with N-ERD with airway eosinophilia. Blood eosinophilia appeared to be a useful predictor of airway T2 signature (area under the curve [AUC]= 0.83); however, surrogate biomarkers had moderate performance in distinguishing eosinophilic N-ERD (for blood eosinophils, AUC= 0.72; for periostin, AUC= 0.75). Lower airway immune profiles show considerable heterogeneity of N-ERD, with skewing toward T2 response and eosinophilic inflammation. Increased production of leukotriene E4 was restricted to a subgroup of patients with eosinophilia in the lower airway.

N-3 PUFA and inflammation: from membrane to nucleus and from bench to bedside.

Inflammation is a normal part of the immune response and should be self-limiting. Excessive or unresolved inflammation is linked to tissue damage, pathology and ill health. Prostaglandins and leukotrienes produced from the n-6 fatty acid arachidonic acid are involved in inflammation. Fatty acids may also influence inflammatory processes through mechanisms not necessarily involving lipid mediators. The n-3 fatty acids EPA and DHA possess a range of anti-inflammatory actions. Increased content of EPA and DHA in the membranes of cells involved in inflammation has effects on the physical nature of the membranes and on the formation of signalling platforms called lipid rafts. EPA and DHA interfere with arachidonic acid metabolism which yields prostaglandins and leukotrienes involved in inflammation. EPA gives rise to weak (e.g. less inflammatory) analogues and both EPA and DHA are substrates for the synthesis of specialised pro-resolving mediators. Through their effects on early signalling events in membranes and on the profile of lipid mediators produced, EPA and DHA alter both intracellular and intercellular signals. Within cells, this leads to altered patterns of gene expression and of protein production. The net result is decreased production of inflammatory cytokines, chemokines, adhesion molecules, proteases and enzymes. The anti-inflammatory and inflammation-resolving effects of EPA and DHA are relevant to both prevention and treatment of human diseases that have an inflammatory component. This has been widely studied in rheumatoid arthritis where there is good evidence that high doses of EPA + DHA reduce pain and other symptoms.

Distinct Profiles of Specialized Pro-resolving Lipid Mediators and Corresponding Receptor Gene Expression in Periodontal Inflammation.

Polyunsaturated fatty acid-derived specialized pro-resolving lipid mediators (SPMs) play an important role in modulating inflammation. The aim of the study was to compare profiles of SPMs, SPM related lipid mediators and SPM receptor gene expression in gingiva of subjects with periodontitis to healthy controls. A total of 28 subjects were included; 13 periodontally healthy and 15 periodontitis before or after non-surgical periodontal therapy. Gingival tissues were collected from two representative posterior teeth prior to and 8 weeks after scaling and root planning; only once in the healthy group. Lipid mediator-SPM metabololipidomics was performed to identify metabolites in gingiva. qRT-PCR was performed to assess relative gene expression (2−ΔΔCT) of known SPM receptors. Intergroup comparisons were made using Wilcoxon tests. Thirty-six omega-6 or omega-3 fatty acid-derived lipid mediators and seven receptor genes were identified in gingiva. Profiles of lipid mediators and receptor gene expression were significantly different between the three groups. Levels of six lipid mediators, 5-HETE, 15-HETE, 15(S)-HEPE, 4-HDHA, 7-HDHA, and 17-HDHA in periodontitis before treatment were significantly higher than in periodontitis after treatment. The expression of BLT1 in the healthy group was significantly higher than periodontitis subjects before and after treatment. The expression of GPR18 in periodontitis before treatment was significantly higher than in periodontitis after treatment while the expression of GPR32 in periodontitis before treatment was significantly lower than in periodontitis after treatment. Elevated levels of SPM biosynthetic pathway markers in periodontitis subjects before treatment indicated inflammation induced pro-resolution activity in gingiva, but receptors for these molecules were deficient in periodontitis pre-treatment suggesting that failure of resolution of inflammation contributes to excess, chronic inflammation in periodontitis.

BAL and serum multiplex lipid profiling in idiopathic pulmonary fibrosis and fibrotic hypersensitivity pneumonitis

BackgroundDifferential diagnosis between IPF and fibrotic HP (fHP) can be challenging: these two ILDs share many common features but call for different therapeutic approaches. In the present study, differential lipid mediator profiles were analysed by a new method in BAL and serum from HP and IPF patients. Materials and methods76 patients were enrolled retrospectively in the study. Median age (IQR) was 67 years (51–74); 63% were males, 30 had fHP and 46 had IPF. Serum and BAL samples were collected at initial diagnosis. For quantification of serum and BAL lipid mediators was used bead-based multiplex LEGENDPlex™ analysis (Biolegend). ResultsSerum Apo A1 levels were significantly higher in IPF than fHP patients (p = 0.314); indeed, serum levels of CCL2 and Apo C3 were lower in HP than in IPF patients (p = 0.013 and p = 0.041, respectively). BAL concentrations of Apo A1, adipsin, Apo C3 and APN were significantly lower in IPF than in fHP patients (p < 0.0001, p < 0.0001, p = 0.007 and p = 0.023, respectively). In the logistic regression, IPF was tested as dependent variable. Serum levels of Apo A1, CCL2 and Apo C3 were tested as independent variables and ROC curve analysis of model performance showed AUC 93% (p < 0.0001); on the other hand, BAL concentrations of Apo A1, adipsin, Apo C3 and APN showed AUC 81% (p < 0.0001). DiscussionLipid biomarkers evaluated in BAL in our study confirm the hypothesis that fHP and IPF have different lung fibrosis phenotypes. The former is a post-inflammatory cell-regulated ILD and the second is more related to tissue remodeling and repair.

Inflammatory macrophage memory in nonsteroidal anti-inflammatory drug–exacerbated respiratory disease

Nonsteroidal anti-inflammatory drug-exacerbated respiratory disease (N-ERD) is a chronic inflammatory condition, which is driven by an aberrant arachidonic acid metabolism. Macrophages are major producers of arachidonic acid metabolites and subject to metabolic reprogramming, but they have been neglected in N-ERD. This study sought to elucidate a potential metabolic and epigenetic macrophage reprogramming in N-ERD. Transcriptional, metabolic, and lipid mediator profiles in macrophages from patients with N-ERD and healthy controls were assessed by RNA sequencing, Seahorse assays, and LC-MS/MS. Metabolites in nasal lining fluid, sputum, and plasma from patients with N-ERD (n= 15) and healthy individuals (n= 10) were quantified by targeted metabolomics analyses. Genome-wide methylomics were deployed to define epigenetic mechanisms of macrophage reprogramming in N-ERD. This study shows that N-ERD monocytes/macrophages exhibit an overall reduction in DNA methylation, aberrant metabolic profiles, and an increased expression of chemokines, indicative of a persistent proinflammatory activation. Differentially methylated regions in N-ERD macrophages included genes involved in chemokine signaling and acylcarnitine metabolism. Acylcarnitines were increased in macrophages, sputum, nasal lining fluid, and plasma of patients with N-ERD. On inflammatory challenge, N-ERD macrophages produced increased levels of acylcarnitines, proinflammatory arachidonic acid metabolites, cytokines, and chemokines as compared to healthy macrophages. Together, these findings decipher a proinflammatory metabolic and epigenetic reprogramming of macrophages in N-ERD.

THU0085 RESOLVIN D5 MODULATES TH17/TREG CELL DIFFERENTIATION AND SUPPRESSES OSTEOCLASTOGENESIS

Background:Resolution phase of acute inflammation has been recognized not passive but active process during the last 2 decades. This active process is highly regulated by novel families of potent bioactive lipid mediators, which are coined as specialized proresolving mediators (SPMs) including resolvins. Little has been known, however, about how resolvins are involved in chronic inflammation, such as rheumatoid arthritis (RA).Objectives:To investigate whether lipid mediators (LM) are involved in the pathogenesis of RA.Methods:We investigated lipid mediator profiling in the paws of SKG arthritis mice by using lipid chromatography (LC) /mass spectrometry (MS) /MS-based LM metabololipidomics. CD4+T cells from spleens of SKG mice were cultured on anti-CD3/ CD28Abs precoated plate with IL-6/TGF-β, anti-IFNγ/IL-4 and analyzed by flow cytometry. CD4+T cells were labeled with CFSE, and cell proliferation was analyzed by flow cytometry. Mouse bone marrow cells were cultured with M-CSF and RANKL, and TRAP-positive multinucleated cells were defined as osteoclasts. Osteoclast differentiation markers were analyzed by qRT-PCR. RvD5 or normal saline was administered daily into the peritoneal cavity of arthritic SKG mice.Results:RvE3, RvD1, RvD3, RvD5 and Maresin2 were significantly elevated on the paws of arthritic SKG mice. Among the elevated SPMs, only RvD5 levels on arthritic paws were significantly correlated with arthritis disease activity (Figure 1). We demonstrated that RvD5 suppressed Th17 cell differentiation, and facilitated Treg cell differentiationin vitro. In addition, RvD5 inhibited CD4+T cell proliferation. Furthermore, RvD5 attenuated osteoclast differentiation (Figure 2) and interfered osteoclastogenesis at the molecular level. In thein vivoexperiment, incidence of arthritis tended to be lower in RvD5-treated mice than that in control group, although there was no significant difference.Conclusion:RvD5 is increased in the paws of arthritic mice, and that RvD5 suppresses Th17 cell differentiation and CD4+T cell proliferation, facilitates Treg cell differentiation, and suppresses osteoclastogenesis.

Intake of Camelina Sativa Oil and Fatty Fish Alter the Plasma Lipid Mediator Profile in Subjects with Impaired Glucose Metabolism – A Randomized Controlled Trial

n-3 and n-6 polyunsaturated fatty acids (PUFAs) and their lipid mediator metabolites are associated with inflammation. We investigated the effect of dietary intake of plant- and animal-derived n-3 PUFAs and fish protein on the circulatory concentrations of lipid mediators. Seventy-nine subjects with impaired fasting glucose who completed the controlled dietary intervention after randomization to the fatty fish (FF, n=20), lean fish (LF, n=21), Camelina sativa oil (CSO, n=18) or control group (n=20) for 12 weeks were studied. Lipid mediator profiling from fasting plasma samples before and after the intervention was performed by liquid chromatography-mass spectrometry (LC-MS/MS). The FF diet increased concentrations of 18-hydroxyeicosapentaenoic acid (18-HEPE) and 4- and 17-hydroxydocosahexaenoic acid (4-, 17-HDoHE) derived from eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), respectively. Concentrations of lipid mediators derived from α-linolenic acid (ALA) increased and arachidonic acid (AA) derived 5-iso prostaglandin F2α-VI decreased in the CSO group. There were no significant changes in lipid mediators in the LF group. The dietary intake of both plant and animal-based n-3 PUFAs increased circulatory concentrations of lipid mediators with potential anti-inflammatory properties.

Lipid Mediator Profiles Predict Response to Therapy with an Oral Frankincense Extract in Relapsing-Remitting Multiple Sclerosis

Lipid mediators (LMs) are a unique class of immunoregulatory signalling molecules and known to be affected by frankincense extracts. We performed LM profiling by metabololipidomics in plasma samples from 28 relapsing-remitting multiple sclerosis (RR-MS) patients who took a standardised frankincense extract (SFE) daily for eight months in a clinical phase IIa trial (NCT01450124) and in 28 age- and gender-matched healthy controls. Magnetic resonance imaging, immunological outcomes and serum neurofilament light chain levels were correlated to changes in the LM profiles of the RR-MS cohort. Eight out of 44 analysed LMs were significantly reduced during an eight-month treatment period by the SFE and seven of these eight significant LM derive from the 5-lipoxygenase (5-LO) pathway. Baseline levels of 12- and 15-LO products were elevated in patients who exhibited disease activity (EDA) during SFE treatment compared to no-evidence-of-disease-activity (NEDA) patients and could predict treatment response to the SFE in a prediction model at baseline. Oral treatment with an SFE significantly reduces 5-LO-derived LMs in RR-MS patients during an eight-month treatment period. Treatment response to an SFE, however, seems to be related to 12-,15-LO and cyclooxygenase product levels before SFE exposure. Further studies should confirm their biomarker potential in RR-MS and SFE treatment.

Metabolomic fingerprinting and systemic inflammatory profiling of asthma COPD overlap (ACO)

BackgroundAsthma-COPD overlap (ACO) refers to a group of poorly studied and characterised patients reporting with disease presentations of both asthma and COPD, thereby making both diagnosis and treatment challenging for the clinicians. They exhibit a higher burden in terms of both mortality and morbidity in comparison to patients with only asthma or COPD. The pathophysiology of the disease and its existence as a unique disease entity remains unclear. The present study aims to determine whether ACO has a distinct metabolic and immunological mediator profile in comparison to asthma and COPD.MethodsGlobal metabolomic profiling using two different groups of patients [discovery (D) and validation (V)] were conducted. Serum samples obtained from moderate and severe asthma [n = 34(D); n = 32(V)], moderate and severe COPD [n = 30(D); 32(V)], ACO patients [n = 35(D); 40(V)] and healthy controls [n = 33(D)] were characterized using gas chromatography mass spectrometry (GC-MS). Multiplexed analysis of 25 immunological markers (IFN-γ (interferon gamma), TNF-α (tumor necrosis factor alpha), IL-12p70 (interleukin 12p70), IL-2, IL-4, IL-5, IL-13, IL-10, IL-1α, IL-1β, TGF-β (transforming growth factor), IL-6, IL-17E, IL-21, IL-23, eotaxin, GM-CSF (granulocyte macrophage-colony stimulating factor), IFN-α (interferon alpha), IL-18, NGAL (neutrophil gelatinase-associated lipocalin), periostin, TSLP (thymic stromal lymphopoietin), MCP-1 (monocyte chemoattractant protein- 1), YKL-40 (chitinase 3 like 1) and IL-8) was also performed in the discovery cohort.ResultsEleven metabolites [serine, threonine, ethanolamine, glucose, cholesterol, 2-palmitoylglycerol, stearic acid, lactic acid, linoleic acid, D-mannose and succinic acid] were found to be significantly altered in ACO as compared with asthma and COPD. The levels and expression trends were successfully validated in a fresh cohort of subjects. Thirteen immunological mediators including TNFα, IL-1β, IL-17E, GM-CSF, IL-18, NGAL, IL-5, IL-10, MCP-1, YKL-40, IFN-γ, IL-6 and TGF-β showed distinct expression patterns in ACO. These markers and metabolites exhibited significant correlation with each other and also with lung function parameters.ConclusionsThe energy metabolites, cholesterol and fatty acids correlated significantly with the immunological mediators, suggesting existence of a possible link between the inflammatory status of these patients and impaired metabolism. The present findings could be possibly extended to better define the ACO diagnostic criteria, management and tailoring therapies exclusively for the disease.

Lipidomics Profiling of Hidradenitis Suppurativa Skin Lesions Reveals Lipoxygenase Pathway Dysregulation and Accumulation of Proinflammatory Leukotriene B4

Hidradenitis suppurativa (HS) is a chronic, recurring inflammatory dermatosis characterized by abscesses, deep-seated nodules, sinus tracts, and fibrosis in skin lesions around hair follicles of the axillary, inguinal, and anogenital regions. Whereas the exact pathogenesis remains poorly defined, clear evidence suggests that HS is a multifactorial inflammatory disease characterized by innate and adaptive immune components. Bioactive lipids are important regulators of cutaneous homeostasis, inflammation, and resolution of inflammation. Alterations in the lipid mediator profile can lead to malfunction and cutaneous inflammation. We used targeted lipidomics to analyze selected omega-3 and omega-6 polyunsaturated fatty acids in skin of patients with HS and of healthy volunteers. Lesional HS skin displayed enrichment of 5-lipoxygenase (LO)‒derived metabolites, especially leukotriene B4. In addition, 15-LO‒derived metabolites were underrepresented in HS lesions. Changes in the lipid mediator profile were accompanied by transcriptomic dysregulation of the 5-LO and 15-LO pathways. Hyperactivation of the 5-LO pathway in lesional macrophages identified these cells as potential sources of leukotriene B4, which may cause neutrophil influx and activation. Furthermore, leukotriene B4-induced mediators and pathways were elevated in HS lesions, suggesting a contribution of this proinflammatory lipid meditator to the pathophysiology of HS.

Imbalance of proresolving lipid mediators in persistent allodynia dissociated from signs of clinical arthritis.

Rheumatoid arthritis-associated pain is poorly managed, often persisting when joint inflammation is pharmacologically controlled. Comparably, in the mouse K/BxN serum-transfer model of inflammatory arthritis, hind paw nociceptive hypersensitivity occurs with ankle joint swelling (5 days after immunisation) persisting after swelling has resolved (25 days after immunisation). In this study, lipid mediator (LM) profiling of lumbar dorsal root ganglia (DRG), the site of sensory neuron cell bodies innervating the ankle joints, 5 days and 25 days after serum transfer demonstrated a shift in specialised proresolving LM profiles. Persistent nociception without joint swelling was associated with low concentrations of the specialised proresolving LM Maresin 1 (MaR1) and high macrophage numbers in DRG. MaR1 application to cultured DRG neurons inhibited both capsaicin-induced increase of intracellular calcium ions and release of calcitonin gene-related peptide in a dose-dependent manner. Furthermore, in peritoneal macrophages challenged with lipopolysaccharide, MaR1 reduced proinflammatory cytokine expression. Systemic MaR1 administration caused sustained reversal of nociceptive hypersensitivity and reduced inflammatory macrophage numbers in DRG. Unlike gabapentin, which was used as positive control, systemic MaR1 did not display acute antihyperalgesic action. Therefore, these data suggest that MaR1 effects observed after K/BxN serum transfer relate to modulation of macrophage recruitment, more likely than to direct actions on sensory neurons. Our study highlights that, in DRG, aberrant proresolution mechanisms play a key role in arthritis joint pain dissociated from joint swelling, opening novel approaches for rheumatoid arthritis pain treatment.

Flaxseed (Linum Usitatissimum L.) Supplementation in Patients Undergoing Lipoprotein Apheresis for Severe Hyperlipidemia-A Pilot Study.

Being rich in polyunsaturated fatty acids, flaxseed (Linum usitatissimum L.) is thought to be able to decrease lipid levels and dampen inflammation. In this pilot study, we aimed to determine whether flaxseed supplementation could improve the profiles of lipids and inflammatory mediators in patients with severe hyperlipidemia resistant to conventional lipid-lowering pharmacotherapy and requiring lipoprotein apheresis. To this end, six patients received, blindly—in addition to their normal lipoprotein apheresis regimen—a 10-week dietary supplementation with flaxseed (28 g/d) administered in biscuits. This was followed by a 10-week washed out-period and a 10-week supplementation phase with whole wheat placebo. Blood samples were collected at the end of each phase, before the lipoprotein apheresis session. The primary endpoint was the lipid profile and the secondary endpoints were the concentrations of inflammatory mediators and tolerability. Flaxseed supplementation was well-tolerated and resulted in a consistent and significant decrease in total cholesterol and low-density lipoprotein (LDL) levels. The median (and range) percentage decrease was 11.5% (0–18.8) and 7.3% (4.4–26.6), for cholesterol (p = 0.015) and LDL-C (p = 0.003), respectively. On the other hand, there was no significant effect of flaxseed on lipoprotein(a) (Lp(a)), C-reactive protein (CRP), and interleukin 6 (IL-6) concentrations. These observations indicate that flaxseed can produce a cholesterol- and LDL-lowering effect in patients treated with lipoprotein apheresis. Thus, flaxseed supplementation may help to control cholesterol in this patient population. The flaxseed supplementation protocol applied may be of use for further adequately-powered studies to validate and extend our findings.

Targeted lipidomics reveals extensive changes in circulating lipid mediators in patients with acutely decompensated cirrhosis

Acute-on-chronic liver failure (ACLF) is a newly described syndrome, which develops in patients with acute decompensation of cirrhosis, and is characterized by intense systemic inflammation, multiple organ failures and high short-term mortality. The profile of circulating lipid mediators, which are endogenous signaling molecules that play a major role in inflammation and immunity, is poorly characterized in ACLF. In the current study, we assessed the profile of lipid mediators by liquid chromatography coupled to tandem mass spectrometry in plasma from patients with acute decompensation of cirrhosis, with (n= 119) and without (n= 127) ACLF, and from healthy controls (n= 18). Measurements were prospectively repeated in 191 patients with acute decompensation of cirrhosis during a 28-day follow-up period. Fifty-nine lipid mediators (out of 100) were detected in plasma from cirrhotic patients, of which 16 were significantly associated with disease status. Among these, 11 lipid mediators distinguished patients at any stage from healthy controls, whereas 2 lipid mediators (LTE4 and 12-HHT, both derived from arachidonic acid) shaped a minimal plasma fingerprint that discriminated patients with ACLF from those without. Levels of LTE4 distinguished ACLF grade 3 from ACLF grades 1 and 2, followed the clinical course of the disease (increased with worsening and decreased with improvement) and positively correlated with markers of inflammation and non-apoptotic cell death. Moreover, LTE4 together with LXA5 (derived from eicosapentaenoic acid) and EKODE (derived from linoleic acid) were associated with short-term mortality. LXA5 and EKODE formed a signature associated with coagulation and liver failures. Taken together, these findings uncover specific lipid mediator profiles associated with disease severity and prognosis in patients with acute decompensation of cirrhosis. Acute-on-chronic liver failure (ACLF) is characterized by intense systemic inflammation, multiple organ failures and high short-term mortality. In the current study, we assessed the plasma lipid profile of 100 bioactive lipid mediators in healthy controls, patients with decompensated cirrhosis, and those who had developed ACLF. We identified lipid mediator signatures associated with inflammation and non-apoptotic cell death that discriminate disease severity and evolution, short-term mortality and organ failures.

Early protein expression profile in bronchoalveolar lavage fluid and clinical outcomes in primary graft dysfunction after lung transplantation.

Primary graft dysfunction (PGD) remains a major post-transplant complication and is associated with increased morbidity and mortality. Mechanisms evoking PGD are not completely clear, but inflammation plays a central role. We investigated the association between PGD and inflammatory proteins present in immediate postoperative bronchoalveolar lavage. All double-lung recipients transplanted at our institution from 2002 to 2018 were included in our study. We retrospectively selected 80 consecutive lung transplant recipients with different PGD grades (n = 20 for each PGD grades 0-1 to 2-3). In bronchoalveolar lavage performed within the first 24 h after donor aortic cross-clamping following lung transplantation, concentrations of 30 cytokines, chemokines and growth factors were assessed by enzyme-linked immunosorbent assay (ELISA) and correlated with donor and recipient demographics and outcomes. For analysis, 2 groups were defined: 'mild' PGD (grade 0-1) and 'severe' PGD (grades 2-3). Significant differences between mild and severe PGD were found in 8 biomarkers [interleukin (IL)-6, IL-10, IL-13, eotaxin, granulocyte colony-stimulating factor, interferon γ, macrophage inflammatory protein 1α, surfactant protein D (SP-D); P < 0.05]. Increased IL-10 and IL-13, but none of the other proteins, were associated with short-term outcome (longer time to extubation; P = 0.005 and P < 0.0001; increased intensive care unit stay; P = 0.012 and P < 0.0001; and hospital stay; P = 0.041 and P = 0.002). There were no significant differences in donor and recipient characteristics between the groups. Expression profiles of key inflammatory mediators in bronchoalveolar lavage fluid differed significantly between lung transplant recipients with severe versus mild PGD and correlated with clinical outcome variables. Further research should focus on the early mechanisms leading to PGD.

Activation of metabolite receptor GPR91 promotes platelet aggregation and transcellular biosynthesis of leukotriene C4

BackgroundSuccinate is a Krebs cycle intermediate whose formation is enhanced under metabolic stress, and for which a selective sensor GPR91 has been identified on various cell types including platelets. Platelet‐derived eicosanoids play pivotal roles in platelet activation/aggregation, which is key to thrombus formation and progression of atherothrombosis. ObjectivesThis study aims to decipher the molecular mechanism(s) and potential involvement of eicosanoids in succinate enhanced platelet activation/aggregation. MethodsWe used liquid chromatography‐mass spectrometry (LC‐MS)/MS‐based lipid mediator profiling to identify eicosanoids regulated by succinate. We ran light transmittance aggregometry and flow cytometry to assess platelet aggregation, P‐selectin expression, and platelet‐polymorphonuclear leukocyte (PMN) adherence. Various pharmacological tools were used to assess the contributions of GPR91 signalling and eicosanoids in platelet aggregation. ResultsSuccinate and two types of synthetic non‐metabolite GPR91 agonists—cis‐epoxysuccinate (cES) and Cmpd131—potentiated platelet aggregation, which was partially blocked by a selective GPR91 antagonist XT1. GPR91 activation increased production of 12‐hydroxy‐eicosatetraenoic acid (12‐HETE), thromboxane (TX) A2, and 12‐hydroxy‐heptadecatrienoic acid (12‐HHT) in human platelets, associated with phosphorylation of cytosolic phospholipase A2 (cPLA2), suggesting increased availability of free arachidonic acid. Blocking 12‐HETE and TXA2 synthesis, or antagonism of the TXA2 receptor, significantly reduced platelet aggregation enhanced by GPR91 signalling. Moreover, platelet‐PMN suspensions challenged with succinate exhibited enhanced transcellular biosynthesis of leukotriene C4 (LTC4), a powerful proinflammatory vascular spasmogen. ConclusionSuccinate signals through GPR91 to promote biosynthesis of eicosanoids, which contribute to platelet aggregation/activation and potentially vascular inflammation. Hence, GPR91 may be a suitable target for pharmacological intervention in atherothrombotic conditions.

16HBE Cell Lipid Mediator Responses to Mono and Co-Infections with Respiratory Pathogens.

Respiratory tract infections are a global health problem. The main causative agents of these infections are influenza A virus (IAV), Staphylococcus aureus (S. aureus), and Streptococcus pneumoniae (S. pneumoniae). Major research focuses on genetics and immune responses in these infections. Eicosanoids and other oxylipins are host-derived lipid mediators that play an important role in the activation and resolution of inflammation. In this study, we assess, for the first time, the different intracellular profiles of these bioactive lipid mediators during S. aureus LUG2012, S. pneumoniae TIGR4, IAV, and corresponding viral and bacterial co-infections of 16HBE cells. We observed a multitude of altered lipid mediators. Changes in the amount of 5-hydroxyeicosatetraenoic acid (5-HETE) were prominent for all bacterial infections. The infection with S. pneumoniae showed the strongest impact on bioactive lipid production and led to alterations in the amount of PPARγ ligands and precursors of pro-resolving lipid mediators.

Dysregulated Antibody, Natural Killer Cell and Immune Mediator Profiles in Autoimmune Thyroid Diseases.

The pathogenesis of autoimmune thyroid diseases (AITD) is poorly understood and the association between different immune features and the germline variants involved in AITD are yet unclear. We previously observed systemic depletion of IgG core fucosylation and antennary α1,2 fucosylation in peripheral blood mononuclear cells in AITD, correlated with anti-thyroid peroxidase antibody (TPOAb) levels. Fucose depletion is known to potentiate strong antibody-mediated NK cell activation and enhanced target antigen-expressing cell killing. In autoimmunity, this may translate to autoantibody-mediated immune cell recruitment and attack of self-antigen expressing normal tissues. Hence, we investigated the crosstalk between immune cell traits, secreted proteins, genetic variants and the glycosylation patterns of serum IgG, in a multi-omic and cross-sectional study of 622 individuals from the TwinsUK cohort, 172 of whom were diagnosed with AITD. We observed associations between two genetic variants (rs505922 and rs687621), AITD status, the secretion of Desmoglein-2 protein, and the profile of two IgG N-glycan traits in AITD, but further studies need to be performed to better understand their crosstalk in AITD. On the other side, enhanced afucosylated IgG was positively associated with activatory CD335- CD314+ CD158b+ NK cell subsets. Increased levels of the apoptosis and inflammation markers Caspase-2 and Interleukin-1α positively associated with AITD. Two genetic variants associated with AITD, rs1521 and rs3094228, were also associated with altered expression of the thyrocyte-expressed ligands known to recognize the NK cell immunoreceptors CD314 and CD158b. Our analyses reveal a combination of heightened Fc-active IgG antibodies, effector cells, cytokines and apoptotic signals in AITD, and AITD genetic variants associated with altered expression of thyrocyte-expressed ligands to NK cell immunoreceptors. Together, TPOAb responses, dysregulated immune features, germline variants associated with immunoactivity profiles, are consistent with a positive autoreactive antibody-dependent NK cell-mediated immune response likely drawn to the thyroid gland in AITD.

Dynamic changes of angiopoietins and endothelial nitric oxide supply during fluid resuscitation for major gyn-oncological surgery: a prospective observation

BackgroundDespite goal-directed hemodynamic therapy, vascular function may deteriorate during surgery for advanced abdominal tumor masses. Fluid administration has been shown to be associated with distinct changes in serum levels of functional proteins. We sought to determine how serum total protein and angiopoietin (ANG) levels change during major abdominal tumor surgery. In addition, ex vivo endothelial nitric oxide synthase (eNOS) activation as well as NO bioavailability in vivo were assessed.Methods30 patients scheduled for laparotomy for late-stage ovarian or uterine cancer were prospectively included. Advanced hemodynamic monitoring as well as protocol-driven goal-directed fluid optimization were performed. Total serum protein, ANG-1, -2, and soluble TIE2 were determined pre-, intra-, and postoperatively. Phosphorylation of eNOS was assessed in microvascular endothelial cells after incubation with patient serum, and microvascular reactivity was determined in vivo by near-infrared spectroscopy and arterial vascular occlusion.ResultsCardiac output as well as preload gradually decreased during surgery and were associated with a median total fluid intake of 12.8 (9.7–15.4) mL/kg*h and a postoperative fluid balance of 6710 (4113–9271) mL. Total serum protein decreased significantly from baseline (66.5 (56.4–73.3) mg/mL) by almost half intraoperatively (42.7 (36.8–51.5) mg/mL, p < 0.0001) and remained at low level. While ANG-1 showed no significant dilutional change (baseline: 12.7 (11.9–13.9) ng/mL, postop.: 11.6 (10.8 –13.5) ng/mL, p = 0.06), serum levels of ANG-2 were even increased postoperatively (baseline: 2.2 (1.6–2.6) ng/mL vs. postop.: 3.4 (2.3–3.8) ng/mL, p < 0.0001), resulting in a significant shift in ANG-2 to ANG-1 ratio. Ex vivo phosphorylation of eNOS was decreased depending on increased ANG-2 levels and ANG-2/1 ratio (Spearman r = − 0.37, p = 0.007). In vivo, increased ANG-2 levels were associated with impaired capillary recruitment and NO bioavailability (Spearman r = − 0.83, p = 0.01).ConclusionsFluid resuscitation-associated changes in serum vascular mediator profile during abdominal tumor surgery were accompanied by impaired eNOS activity ex vivo as well as reduced NO bioavailability in vivo. Our results may explain disturbed microvascular function in major surgery despite goal-directed hemodynamic optimization.

Prenatal dietary supplements influence the infant airway microbiota in a randomized factorial clinical trial

Maternal dietary interventions during pregnancy with fish oil and high dose vitamin D have been shown to reduce the incidence of asthma and wheeze in offspring, potentially through microbial effects in pregnancy or early childhood. Here we analyze the bacterial compositions in longitudinal samples from 695 pregnant women and their children according to intervention group in a nested, factorial, double-blind, placebo-controlled, randomized trial of n-3 long-chain fatty acids and vitamin D supplementation. The dietary interventions affect the infant airways, but not the infant fecal or maternal vaginal microbiota. Changes in overall beta diversity are observed, which in turn associates with a change in immune mediator profile. In addition, airway microbial maturation and the relative abundance of specific bacterial genera are altered. Furthermore, mediation analysis reveals the changed airway microbiota to be a minor and non-significant mediator of the protective effect of the dietary interventions on risk of asthma. Our results demonstrate the potential of prenatal dietary supplements as manipulators of the early airway bacterial colonization.