Iatrogenic air embolism: pathoanatomy, thromboinflammation, endotheliopathy, and therapies.

Iatrogenic vascular air embolism is a relatively infrequent event but is associated with significant morbidity and mortality. These emboli can arise in many clinical settings such as neurosurgery, cardiac surgery, and liver transplantation, but more recently, endoscopy, hemodialysis, thoracentesis, tissue biopsy, angiography, and central and peripheral venous access and removal have overtaken surgery and trauma as significant causes of vascular air embolism. The true incidence may be greater since many of these air emboli are asymptomatic and frequently go undiagnosed or unreported. Due to the rarity of vascular air embolism and because of the many manifestations, diagnoses can be difficult and require immediate therapeutic intervention. An iatrogenic air embolism can result in both venous and arterial emboli whose anatomic locations dictate the clinical course. Most clinically significant iatrogenic air emboli are caused by arterial obstruction of small vessels because the pulmonary gas exchange filters the more frequent, smaller volume bubbles that gain access to the venous circulation. However, there is a subset of patients with venous air emboli caused by larger volumes of air who present with more protean manifestations. There have been significant gains in the understanding of the interactions of fluid dynamics, hemostasis, and inflammation caused by air emboli due to in vitro and in vivo studies on flow dynamics of bubbles in small vessels. Intensive research regarding the thromboinflammatory changes at the level of the endothelium has been described recently. The obstruction of vessels by air emboli causes immediate pathoanatomic and immunologic and thromboinflammatory responses at the level of the endothelium. In this review, we describe those immunologic and thromboinflammatory responses at the level of the endothelium as well as evaluate traditional and novel forms of therapy for this rare and often unrecognized clinical condition.

Systemic inflammation early after kidney transplantation is associated with long-term graft loss: a cohort study.

Background: Early graft loss following kidney transplantation is mainly a result of acute rejection or surgical complications, while long-term kidney allograft loss is more complex. We examined the association between systemic inflammation early after kidney transplantation and long-term graft loss, as well as correlations between systemic inflammation scores and inflammatory findings in biopsies 6 weeks and 1 year after kidney transplantation. Methods: We measured 21 inflammatory biomarkers 10 weeks after transplantation in 699 patients who were transplanted between 2009 and 2012 at Oslo University Hospital, Rikshospitalet, Norway. Low-grade inflammation was assessed with predefined inflammation scores based on specific biomarkers: one overall inflammation score and five pathway-specific scores. Surveillance or indication biopsies were performed in all patients 6 weeks after transplantation. The scores were tested in Cox regression models. Results: Median follow-up time was 9.1 years (interquartile range 7.6-10.7 years). During the study period, there were 84 (12.2%) death-censored graft losses. The overall inflammation score was associated with long-term kidney graft loss both when assessed as a continuous variable (hazard ratio 1.03, 95% CI 1.01-1.06, P = 0.005) and as a categorical variable (4th quartile: hazard ratio 3.19, 95% CI 1.43-7.10, P = 0.005). In the pathway-specific analyses, fibrogenesis activity and vascular inflammation stood out. The vascular inflammation score was associated with inflammation in biopsies 6 weeks and 1 year after transplantation, while the fibrinogenesis score was associated with interstitial fibrosis and tubular atrophy. Conclusion: In conclusion, a systemic inflammatory environment early after kidney transplantation was associated with biopsy-confirmed kidney graft pathology and long-term kidney graft loss. The systemic vascular inflammation score correlated with inflammatory findings in biopsies 6 weeks and 1 year after transplantation.

In vitro evaluation of iron oxide nanoparticle-induced thromboinflammatory response using a combined human whole blood and endothelial cell model.

Iron oxide nanoparticles (IONPs) are widely used in diagnostic and therapeutic settings. Upon systemic administration, however, they are rapidly recognized by components of innate immunity, which limit their therapeutic capacity and can potentially lead to adverse side effects. IONPs were previously found to induce the inflammatory response in human whole blood, including activation of the complement system and increased secretion of cytokines. Here, we investigated the thromboinflammatory response of 10-30 nm IONPs in lepirudin anticoagulated whole blood in interplay with endothelial cells and evaluated the therapeutic effect of applying complement inhibitors to limit adverse effects related to thromboinflammation. We found that IONPs induced complement activation, primarily at the C3-level, in whole blood incubated for up to four hours at 37°C with and without human microvascular endothelial cells. Furthermore, IONPs mediated a strong thromboinflammatory response, as seen by the significantly increased release of 21 of the 27 analyzed cytokines (p<0.05). IONPs also significantly increased cell-activation markers of endothelial cells [ICAM-1 (p<0.0001), P/E-selectin (p<0.05)], monocytes, and granulocytes [CD11b (p<0.001)], and platelets [CD62P (p<0.05), CD63 (p<0.05), NAP-2 (p<0.01), PF4 (p<0.05)], and showed cytotoxic effects, as seen by increased LDH (p<0.001) and heme (p<0.0001) levels. We found that inflammation and endothelial cell activation were partly complement-dependent and inhibition of complement at the level of C3 by compstatin Cp40 significantly attenuated expression of ICAM-1 (p<0.01) and selectins (p<0.05). We show that complement activation plays an important role in the IONPs-induced thromboinflammatory response and that complement inhibition is promising in improving IONPs biocompatibility.

H2S-Enriched Flush out Does Not Increase Donor Organ Quality in a Porcine Kidney Perfusion Model.

Kidney extraction time has a detrimental effect on post-transplantation outcome. This study aims to improve the flush-out and potentially decrease ischemic injury by the addition of hydrogen sulphide (H2S) to the flush medium. Porcine kidneys (n = 22) were extracted during organ recovery surgery. Pigs underwent brain death induction or a Sham operation, resulting in four groups: donation after brain death (DBD) control, DBD H2S, non-DBD control, and non-DBD H2S. Directly after the abdominal flush, kidneys were extracted and flushed with or without H2S and stored for 13 h via static cold storage (SCS) +/- H2S before reperfusion on normothermic machine perfusion. Pro-inflammatory cytokines IL-1b and IL-8 were significantly lower in H2S treated DBD kidneys during NMP (p = 0.03). The non-DBD kidneys show superiority in renal function (creatinine clearance and FENa) compared to the DBD control group (p = 0.03 and p = 0.004). No differences were seen in perfusion parameters, injury markers and histological appearance. We found an overall trend of better renal function in the non-DBD kidneys compared to the DBD kidneys. The addition of H2S during the flush out and SCS resulted in a reduction in pro-inflammatory cytokines without affecting renal function or injury markers.

The neoepitope of the complement C5b-9 Membrane Attack Complex is formed by proximity of adjacent ancillary regions of C9.

The Membrane Attack Complex (MAC) is responsible for forming large ß-barrel channels in the membranes of pathogens, such as gram-negative bacteria. Off-target MAC assembly on endogenous tissue is associated with inflammatory diseases and cancer. Accordingly, a human C5b-9 specific antibody, aE11, has been developed that detects a neoepitope exposed in C9 when it is incorporated into the C5b-9 complex, but not present in the plasma native C9. For nearly four decades aE11 has been routinely used to study complement, MAC-related inflammation, and pathophysiology. However, the identity of C9 neoepitope remains unknown. Here, we determined the cryo-EM structure of aE11 in complex with polyC9 at 3.2 Å resolution. The aE11 binding site is formed by two separate surfaces of the oligomeric C9 periphery and is therefore a discontinuous quaternary epitope. These surfaces are contributed by portions of the adjacent TSP1, LDLRA, and MACPF domains of two neighbouring C9 protomers. By substituting key antibody interacting residues to the murine orthologue, we validated the unusual binding modality of aE11. Furthermore, aE11 can recognise a partial epitope in purified monomeric C9 in vitro, albeit weakly. Taken together, our results reveal the structural basis for MAC recognition by aE11.

Synthetic Oligodeoxynucleotide CpG Motifs Activate Human Complement through Their Backbone Structure and Induce Complement-Dependent Cytokine Release.

Bacterial and mitochondrial DNA, sharing an evolutionary origin, act as danger-associated molecular patterns in infectious and sterile inflammation. They both contain immunomodulatory CpG motifs. Interactions between CpG motifs and the complement system are sparsely described, and mechanisms of complement activation by CpG remain unclear. Lepirudin-anticoagulated human whole blood and plasma were incubated with increasing concentrations of three classes of synthetic CpGs: CpG-A, -B, and -C oligodeoxynucleotides and their GpC sequence controls. Complement activation products were analyzed by immunoassays. Cytokine levels were determined via 27-plex beads-based immunoassay, and CpG interactions with individual complement proteins were evaluated using magnetic beads coated with CpG-B. In whole blood and plasma, CpG-B and CpG-C (p < 0.05 for both), but not CpG-A (p > 0.8 for all), led to time- and dose-dependent increase of soluble C5b-9, the alternative complement convertase C3bBbP, and the C3 cleavage product C3bc. GpC-A, -B, and -C changed soluble fluid-phase C5b-9, C3bBbP, and C3bc to the same extent as CpG-A, -B, and -C, indicating a DNA backbone-dependent effect. Dose-dependent CpG-B binding was found to C1q (r = 0.83; p = 0.006) and factor H (r = 0.93; p < 0.001). The stimulatory complement effect was partly preserved in C2-deficient plasma and completely preserved in MASP-2-deficient serum. CpG-B increased levels of IL-1ß, IL-2, IL-6, IL-8, MCP-1, and TNF in whole blood, which were completely abolished by inhibition of C5 and C5aR1 (p < 0.05 for all). In conclusion, synthetic analogs of bacterial and mitochondrial DNA activate the complement system via the DNA backbone. We suggest that CpG-B interacts directly with classical and alternative pathway components, resulting in complement-C5aR1-dependent cytokine release.

Venous Air Embolism Activates Complement C3 Without Corresponding C5 Activation and Trigger Thromboinflammation in Pigs.

Introduction: Air embolism may complicate invasive medical procedures. Bubbles trigger complement C3-mediated cytokine release, coagulation, and platelet activation in vitro in human whole blood. Since these findings have not been verified in vivo, we aimed to examine the effects of air embolism in pigs on thromboinflammation. Methods: Forty-five landrace pigs, average 17 kg (range 8.5-30), underwent intravenous air infusion for 300 or 360 minutes (n=29) or served as sham (n=14). Fourteen pigs were excluded due to e.g. infections or persistent foramen ovale. Blood was analyzed for white blood cells (WBC), complement activation (C3a and terminal C5b-9 complement complex [TCC]), cytokines, and hemostatic parameters including thrombin-antithrombin (TAT) using immunoassays and rotational thromboelastometry (ROTEM). Lung tissue was analyzed for complement and cytokines using qPCR and immunoassays. Results are presented as medians with interquartile range. Results: In 24 pigs receiving air infusion, WBC increased from 17×109/L (10-24) to 28 (16-42) (p<0.001). C3a increased from 21 ng/mL (15-46) to 67 (39-84) (p<0.001), whereas TCC increased only modestly (p=0.02). TAT increased from 35 µg/mL (28-42) to 51 (38-89) (p=0.002). ROTEM changed during first 120 minutes: Clotting time decreased from 613 seconds (531-677) to 538 (399-620) (p=0.006), clot formation time decreased from 161 seconds (122-195) to 124 (83-162) (p=0.02) and α-angle increased from 62 degrees (57-68) to 68 (62-74) (p=0.02). In lungs from pigs receiving air compared to sham animals, C3a was 34 ng/mL (14-50) versus 4.1 (2.4-5.7) (p<0.001), whereas TCC was 0.3 CAU/mL (0.2-0.3) versus 0.2 (0.1-0.2) (p=0.02). Lung cytokines in pigs receiving air compared to sham animals were: IL-1ß 302 pg/mL (190-437) versus 107 (66-120), IL-6 644 pg/mL (358-1094) versus 25 (23-30), IL-8 203 pg/mL (81-377) versus 21 (20-35), and TNF 113 pg/mL (96-147) versus 16 (13-22) (all p<0.001). Cytokine mRNA in lung tissue from pigs receiving air compared to sham animals increased 12-fold for IL-1ß, 121-fold for IL-6, and 17-fold for IL-8 (all p<0.001). Conclusion: Venous air embolism in pigs activated C3 without a corresponding C5 activation and triggered thromboinflammation, consistent with a C3-dependent mechanism. C3-inhibition might represent a therapeutic approach to attenuate this response.

Mitochondrial C5aR1 activity in macrophages controls IL-1ß production underlying sterile inflammation.

While serum-circulating complement destroys invading pathogens, intracellularly active complement, termed the "complosome," functions as a vital orchestrator of cell-metabolic events underlying T cell effector responses. Whether intracellular complement is also nonredundant for the activity of myeloid immune cells is currently unknown. Here, we show that monocytes and macrophages constitutively express complement component (C) 5 and generate autocrine C5a via formation of an intracellular C5 convertase. Cholesterol crystal sensing by macrophages induced C5aR1 signaling on mitochondrial membranes, which shifted ATP production via reverse electron chain flux toward reactive oxygen species generation and anaerobic glycolysis to favor IL-1ß production, both at the transcriptional level and processing of pro­IL-1ß. Consequently, atherosclerosis-prone mice lacking macrophage-specific C5ar1 had ameliorated cardiovascular disease on a high-cholesterol diet. Conversely, inflammatory gene signatures and IL-1ß produced by cells in unstable atherosclerotic plaques of patients were normalized by a specific cell-permeable C5aR1 antagonist. Deficiency of the macrophage cell-autonomous C5 system also protected mice from crystal nephropathy mediated by folic acid. These data demonstrate the unexpected intracellular formation of a C5 convertase and identify C5aR1 as a direct modulator of mitochondrial function and inflammatory output from myeloid cells. Together, these findings suggest that the complosome is a contributor to the biologic processes underlying sterile inflammation and indicate that targeting this system could be beneficial in macrophage-dependent diseases, such as atherosclerosis.

Elevated Terminal C5b-9 Complement Complex 10 Weeks Post Kidney Transplantation Was Associated With Reduced Long-Term Patient and Kidney Graft Survival.

Background: The major reason for graft loss is chronic tissue damage, as interstitial fibrosis and tubular atrophy (IF/TA), where complement activation may serve as a mediator. The association of complement activation in a stable phase early after kidney transplantation with long-term outcomes is unexplored. Methods: We examined plasma terminal C5b-9 complement complex (TCC) 10 weeks posttransplant in 900 patients receiving a kidney between 2007 and 2012. Clinical outcomes were assessed after a median observation time of 9.3 years [interquartile range (IQR) 7.5-10.6]. Results: Elevated TCC plasma values (≥0.7 CAU/ml) were present in 138 patients (15.3%) and associated with a lower 10-year patient survival rate (65.7% vs. 75.5%, P < 0.003). Similarly, 10-year graft survival was lower with elevated TCC; 56.9% vs. 67.3% (P < 0.002). Graft survival was also lower when censored for death; 81.5% vs. 87.3% (P = 0.04). In multivariable Cox analyses, impaired patient survival was significantly associated with elevated TCC [hazard ratio (HR) 1.40 (1.02-1.91), P = 0.04] along with male sex, recipient and donor age, smoking, diabetes, and overall survival more than 1 year in renal replacement therapy prior to engraftment. Likewise, elevated TCC was independently associated with graft loss [HR 1.40 (1.06-1.85), P = 0.02] along with the same covariates. Finally, elevated TCC was in addition independently associated with death-censored graft loss [HR 1.69 (1.06-2.71), P = 0.03] as were also HLA-DR mismatches and higher immunological risk. Conclusions: Early complement activation, assessed by plasma TCC, was associated with impaired long-term patient and graft survival.

Open chest and pericardium facilitate transpulmonary passage of venous air emboli.

BACKGROUND: Transpulmonary passage of air emboli can lead to fatal brain- and myocardial infarctions. We studied whether pigs with open chest and pericardium had a greater transpulmonary passage of venous air emboli than pigs with closed thorax. METHODS: We allocated pigs with verified closed foramen ovale to venous air infusion with either open chest with sternotomy and opening of the pleura and pericardium (n = 8) or closed thorax (n = 16). All pigs received a five-hour intravenous infusion of ambient air, starting at 4-6 mL/kg/h and increased by 2 mL/kg/h each hour. We assessed transpulmonary air passage by transesophageal M-mode echocardiography and present the results as median with inter-quartile range (IQR). RESULTS: Transpulmonary air passage occurred in all pigs with open chest and pericardium and in nine pigs with closed thorax (56%). Compared to pigs with closed thorax, pigs with open chest and pericardium had a shorter to air passage (10 minutes (5-16) vs. 120 minutes (44-212), P < .0001), a smaller volume of infused air at the time of transpulmonary passage (12 mL (10-23) vs.170 mL (107-494), P < .0001), shorter time to death (122 minutes (48-185) vs 263 minutes (248-300, P = .0005) and a smaller volume of infused air at the time of death (264 mL (53-466) vs 727 mL (564-968), P = .001). In pigs with open chest and, infused air and time to death correlated strongly (r = 0.95, P = .001). CONCLUSION: Open chest and pericardium facilitated the transpulmonary passage of intravenously infused air in pigs.

CD14 inhibition improves survival and attenuates thrombo-inflammation and cardiopulmonary dysfunction in a baboon model of Escherichia coli sepsis.

BACKGROUND: During sepsis, gram-negative bacteria induce robust inflammation primarily via lipopolysacharride (LPS) signaling through TLR4, a process that involves the glycosylphosphatidylinositol (GPI)-anchored receptor CD14 transferring LPS to the Toll-like receptor 4/myeloid differentiation factor 2 (TLR4/MD-2) complex. Sepsis also triggers the onset of disseminated intravascular coagulation and consumptive coagulopathy. OBJECTIVES: We investigated the effect of CD14 blockade on sepsis-induced coagulopathy, inflammation, organ dysfunction, and mortality. METHODS: We used a baboon model of lethal Escherichia (E) coli sepsis to study two experimental groups (n = 5): (a) E coli challenge; (b) E coli challenge plus anti-CD14 (23G4) inhibitory antibody administered as an intravenous bolus 30 minutes before the E coli. RESULTS: Following anti-CD14 treatment, two animals reached the 7-day end-point survivor criteria, while three animals had a significantly prolonged survival as compared to the non-treated animals that developed multiple organ failure and died within 30 hours. Anti-CD14 reduced the activation of coagulation through inhibition of tissue factor-dependent pathway, especially in the survivors, and enhanced the fibrinolysis due to strong inhibition of plasminogen activator inhibitor 1. The treatment prevented the robust complement activation induced by E coli, as shown by significantly decreased C3b, C5a, and sC5b-9. Vital signs, organ function biomarkers, bacteria clearance, and leukocyte and fibrinogen consumption were all improved at varying levels. Anti-CD14 reduced neutrophil activation, cell death, LPS levels, and pro-inflammatory cytokines (tumor necrosis factor, interleukin (IL)-6, IL-1ß, IL-8, interferon gamma, monocyte chemoattractant protein-1), more significantly in the survivors than non-surviving animals. CONCLUSIONS: Our results highlight the crosstalk between coagulation/fibrinolysis, inflammation, and complement systems and suggest a protective role of anti-CD14 treatment in E coli sepsis.

Gut Microbiota-Dependent Trimethylamine N-Oxide Associates With Inflammation in Common Variable Immunodeficiency.

A substantial proportion of patients with common variable immunodeficiency (CVID) have inflammatory and autoimmune complications of unknown etiology. We have previously shown that systemic inflammation in CVID correlates with their gut microbial dysbiosis. The gut microbiota dependent metabolite trimethylamine N-oxide (TMAO) has been linked to several metabolic and inflammatory disorders, but has hitherto not been investigated in relation to CVID. We hypothesized that TMAO is involved in systemic inflammation in CVID. To explore this, we measured plasma concentrations of TMAO, inflammatory markers, and lipopolysaccharide (LPS) in 104 CVID patients and 30 controls. Gut microbiota profiles and the bacterial genes CutC and CntA, which encode enzymes that can convert dietary metabolites to trimethylamine in the colon, were examined in fecal samples from 40 CVID patients and 86 controls. Furthermore, a food frequency questionnaire and the effect of oral antibiotic rifaximin on plasma TMAO concentrations were explored in these 40 patients. We found CVID patients to have higher plasma concentrations of TMAO than controls (TMAO 5.0 [2.9-8.6] vs. 3.2 [2.2-6.3], p = 0.022, median with IQR). The TMAO concentration correlated positively with tumor necrosis factor (p = 0.008, rho = 0.26), interleukin-12 (p = 0.012, rho = 0.25) and LPS (p = 0.034, rho = 0.21). Dietary intake of meat (p = 0.678), fish (p = 0.715), egg (p = 0.138), dairy products (p = 0.284), and fiber (p = 0.767) did not significantly impact on the TMAO concentrations in plasma, nor did a 2-week course of the oral antibiotic rifaximin (p = 0.975). However, plasma TMAO concentrations correlated positively with gut microbial abundance of Gammaproteobacteria (p = 0.021, rho = 0.36). Bacterial gene CntA was present in significantly more CVID samples (75%) than controls (53%), p = 0.020, potentially related to the increased abundance of Gammaproteobacteria in these samples. The current study demonstrates that elevated TMAO concentrations are associated with systemic inflammation and increased gut microbial abundance of Gammaproteobacteria in CVID patients, suggesting that TMAO could be a link between gut microbial dysbiosis and systemic inflammation. Gut microbiota composition could thus be a potential therapeutic target to reduce systemic inflammation in CVID.

Cholesterol crystals use complement to increase NLRP3 signaling pathways in coronary and carotid atherosclerosis.

BACKGROUND: During atherogenesis, cholesterol precipitates into cholesterol crystals (CC) in the vessel wall, which trigger plaque inflammation by activating the NACHT, LRR and PYD domains-containing protein 3 (NLRP3) inflammasome. We investigated the relationship between CC, complement and NLRP3 in patients with cardiovascular disease. METHODS: We analysed plasma, peripheral blood mononuclear cells (PBMC) and carotid plaques from patients with advanced atherosclerosis applying ELISAs, multiplex cytokine assay, qPCR, immunohistochemistry, and gene profiling. FINDINGS: Transcripts of interleukin (IL)-1beta(ß) and NLRP3 were increased and correlated in PBMC from patients with acute coronary syndrome (ACS). Priming of these cells with complement factor 5a (C5a) and tumour necrosis factor (TNF) before incubation with CC resulted in increased IL-1ß protein when compared to healthy controls. As opposed to healthy controls, systemic complement was significantly increased in patients with stable angina pectoris or ACS. In carotid plaques, complement C1q and C5b-9 complex accumulated around CC-clefts, and complement receptors C5aR1, C5aR2 and C3aR1 were higher in carotid plaques compared to control arteries. Priming human carotid plaques with C5a followed by CC incubation resulted in pronounced release of IL-1ß, IL-18 and IL-1α. Additionally, mRNA profiling demonstrated that C5a and TNF priming followed by CC incubation upregulated plaque expression of NLRP3 inflammasome components. INTERPRETATION: We demonstrate that CC are important local- and systemic complement activators, and we reveal that the interaction between CC and complement could exert its effect by activating the NLRP3 inflammasome, thus promoting the progression of atherosclerosis.

Systemic Inflammation Persists the First Year after Mild Traumatic Brain Injury: Results from the Prospective Trondheim Mild Traumatic Brain Injury Study.

Innate immune activation has been attributed a key role in traumatic brain injury (TBI) and successive morbidity. In mild TBI (mTBI), however, the extent and persistence of innate immune activation are unknown. We determined plasma cytokine level changes over 12 months after an mTBI in hospitalized and non-hospitalized patients compared with community controls; and examined their associations to injury-related and demographic variables at admission. Prospectively, 207 patients presenting to the emergency department (ED) or general practitioner with clinically confirmed mTBI and 82 matched community controls were included. Plasma samples were obtained at admission, after 2 weeks, 3 months, and 12 months. Cytokine levels were analysed with a 27-plex beads-based immunoassay. Brain magnetic resonance imaging (MRI) was performed on all participants. Twelve cytokines were reliably detected. Plasma levels of interferon gamma (IFN-γ), interleukin 8 (IL-8), eotaxin, macrophage inflammatory protein-1-beta (MIP-1ß), monocyte chemoattractant protein 1 (MCP-1), IL-17A, IL-9, tumor necrosis factor (TNF), and basic fibroblast growth factor (FGF-basic) were significantly increased at all time-points in patients compared with controls, whereas IFN-γ-inducing protein 10 (IP-10), platelet-derived growth factor (PDGF), and IL-1ra were not. IL-17A and FGF-basic showed significant increases in patients from admission to follow-up at 3 months, and remained increased at 12 months compared with admission. Interestingly, MRI findings were negatively associated with four cytokines: eotaxin, MIP-1ß, IL-9, and IP-10, whereas age was positively associated with nine cytokines: IL-8, eotaxin, MIP-1ß, MCP-1, IL-17A, IL-9, TNF, FGF-basic, and IL-1ra. TNF was also increased in those with presence of other injuries. In conclusion, mTBI activated the innate immune system consistently and this is the first study to show that several inflammatory cytokines remain increased for up to 1 year post-injury.

Comparison of cytokine changes in three different lipoprotein apheresis systems in an ex vivo whole blood model.

INTRODUCTION: Even if proprotein convertase subtilisin/kexin type 9 inhibitors have replaced lipoprotein apheresis in many patients, lipoprotein apheresis still is an important option in homozygous familial hypercholesterolemia, progressive atherosclerosis or when removal of lipoprotein(a) is indicated. Additional possible favorable effects beyond lipid lowering could include changes in the concentration of cytokines and improvement of hemorheology. METHODS: We evaluated how whole blood adsorption, dextran sulfate plasma adsorption, and double filtration plasmapheresis lipoprotein apheresis systems affected cytokine concentrations, using a human whole blood ex vivo model differentiating the effect of the lipoprotein apheresis and plasma separation columns and describing temporal changes. RESULTS: Compared to the control bag, the whole blood adsorption system reduced Interferon-γ (IFN-γ), IL-8, IL-1ra, eotaxin, tumor necrosis factor (TNF), monocyte chemoattractant protein 1 (MCP-1), platelet derived growth factor (PDGF)-BB, regulated on activation T cell expressed and secreted (RANTES), macrophage inflammatory protein-1ß (MIP-1ß), and IP-10 (P < .05). The dextran sulfate plasma adsorption system reduced IFN-γ, IL-8, IL-1ra, eotaxin, TNF, MCP-1, PDGF-BB, MIP-1ß, and IP-10 (P < .05). Vascular endothelial growth factor (VEGF) and granulocyte macrophage colony stimulating factor (GM-CSF) were increased in the whole blood and dextran sulfate plasma adsorption systems (P < .05). The double filtration plasmapheresis system reduced IFN-γ, IL-1ra, TNF, MIP-1ß, and IP-10 (P < .05), while MCP-1,VEGF, GM-CSF, and RANTES were increased (P < .05). The plasma separation column increased concentration of RANTES, and was a barrier to reduction of eotaxin. Temporal patterns of concentration change indicated first pass increase of PDGF-BB and first pass reduction of IP-10. CONCLUSION: There were marked differences in how the three systems affected total and temporal cytokine concentration changes in this in vitro model, as well as compared to former in vivo studies.

Impaired HDL Function Amplifies Systemic Inflammation in Common Variable Immunodeficiency.

Common variable immunodeficiency (CVID) is the most common symptomatic primary immunodeficiency, characterized by inadequate antibody responses and recurrent bacterial infections. Paradoxically, a majority of CVID patients have non-infectious inflammatory and autoimmune complications, associated with systemic immune activation. Our aim was to explore if HDL, known to have anti-inflammatory properties, had impaired function in CVID patients and thereby contributed to their inflammatory phenotype. We found reduced HDL cholesterol levels in plasma of CVID patients compared to healthy controls, particularly in patients with inflammatory and autoimmune complications, correlating negatively with inflammatory markers CRP and sCD25. Reverse cholesterol transport capacity testing showed reduced serum acceptance capacity for cholesterol in CVID patients with inflammatory and autoimmune complications. They also had reduced cholesterol efflux capacity from macrophages to serum and decreased expression of ATP-binding cassette transporter ABCA1. Human HDL suppressed TLR2-induced TNF release less in blood mononuclear cells from CVID patients, associated with decreased expression of transcriptional factor ATF3. Our data suggest a link between impaired HDL function and systemic inflammation in CVID patients, particularly in those with autoimmune and inflammatory complications. This identifies HDL as a novel therapeutic target in CVID as well as other more common conditions characterized by sterile inflammation or autoimmunity.

Iron oxide nanoparticles enhance Toll-like receptor-induced cytokines in a particle size- and actin-dependent manner in human blood.

Aim: To assess the effects of different-sized iron oxide nanoparticles (IONPs) on inflammatory responses in human whole blood. Materials & methods: Human whole blood with and without 10 and 30 nm IONPs was incubated with Toll-like receptor (TLR) ligands. Cytokine levels, complement activation, reactive oxygen species and viability were determined. Results: The 10 nm IONPs enhanced the TLR2/6, TLR4 and partly TLR8-mediated cytokine production, whereas the 30 nm IONPs partly enhanced TLR2/6 and decreased TLR8-mediated cytokine production. Particle-mediated enhancement of TLR4-induced cytokines could not be explained by complement activation, but was dependent on TLR4/MD2 and CD14, as well as actin polymerization. Conclusion: The IONPs differentially affected the TLR ligand-induced cytokines, which has important implications for biomedical applications of IONPs.

Iron oxide nanoparticles induce cytokine secretion in a complement-dependent manner in a human whole blood model.

Iron oxide nanoparticles (IONPs) are promising nanomaterials for biomedical applications. However, their inflammatory potential has not been fully established. Here, we used a lepirudin anti-coagulated human whole blood model to evaluate the potential of 10 nm IONPs to activate the complement system and induce cytokine production. Reactive oxygen species and cell death were also assessed. The IONPs activated complement, as measured by C3a, C5a and sC5b-9, and induced the production of pro-inflammatory cytokines in a particle-dose dependent manner, with the strongest response at 10 µg/mL IONPs. Complement inhibitors at C3 (compstatin analog Cp40) and C5 (eculizumab) levels completely inhibited complement activation and secretion of inflammatory mediators induced by the IONPs. Additionally, blockade of complement receptors C3aR and C5aR1 significantly reduced the levels of various cytokines, indicating that the particle-induced secretion of inflammatory mediators is mainly C5a and C3a mediated. The IONPs did not induce cell death or reactive oxygen species, which further suggests that complement activation alone was responsible for most of the particle-induced cytokines. These data suggest that the lepirudin anti-coagulated human whole blood model is a valuable ex vivo system to study the inflammatory potential of IONPs. We conclude that IONPs induce complement-mediated cytokine secretion in human whole blood.

Combined inhibition of C5 and CD14 efficiently attenuated the inflammatory response in a porcine model of meningococcal sepsis.

BACKGROUND: Fulminant meningococcal sepsis, characterized by overwhelming innate immune activation, mostly affects young people and causes high mortality. This study aimed to investigate the effect of targeting two key molecules of innate immunity, complement component C5, and co-receptor CD14 in the Toll-like receptor system, on the inflammatory response in meningococcal sepsis. METHODS: Meningococcal sepsis was simulated by continuous intravenous infusion of an escalating dose of heat-inactivated Neisseria meningitidis administered over 3 h. The piglets were randomized, blinded to the investigators, to a positive control group (n = 12) receiving saline and to an interventional group (n = 12) receiving a recombinant anti-CD14 monoclonal antibody together with the C5 inhibitor coversin. RESULTS: A substantial increase in plasma complement activation in the untreated group was completely abolished in the treatment group (p = 0.006). The following inflammatory mediators were substantially reduced in plasma in the treatment group: Interferon-γ by 75% (p = 0.0001), tumor necrosis factor by 50% (p = 0.01), Interleukin (IL)-8 by 50% (p = 0.03), IL-10 by 40% (p = 0.04), IL-12p40 by 50% (p = 0.03), and granulocyte CD11b (CR3) expression by 20% (p = 0.01). CONCLUSION: Inhibition of C5 and CD14 may be beneficial in attenuating the detrimental effects of complement activation and modulating the cytokine storm in patients with fulminant meningococcal sepsis.

Plasma Levels of Marine n-3 Fatty Acids Are Inversely Correlated With Proinflammatory Markers sTNFR1 and IL-6 in Renal Transplant Recipients.

OBJECTIVE: Marine n-3 polyunsaturated fatty acids (PUFAs) exert potential anti-inflammatory effects and might improve long-term outcomes after renal transplantation. We assessed associations between plasma phospholipid levels of marine n-3 PUFAs and plasma inflammatory biomarkers 10 weeks after renal transplantation. DESIGN: Cross-sectional single-center study. SUBJECTS: A study population of 861 renal transplant recipients transplanted at Oslo University Hospital between 2007 and 2011. METHODS AND MAIN OUTCOME MEASURE: Plasma phospholipid fatty acids were determined by gas chromatography. Marine n-3 PUFA levels were defined as the sum of eicosapentaenoic acid, docosahexaenoic acid, and docosapentaenoic acid levels in weight percentage of total plasma phospholipid fatty acids. Plasma inflammatory biomarkers were measured by enzyme immunoassays. We used multivariable linear regression analysis to assess associations between levels of marine n-3 PUFAs and inflammatory biomarkers in plasma. RESULTS: Plasma marine n-3 PUFA levels were inversely associated with plasma levels of proinflammatory biomarkers soluble tumor necrosis factor receptor 1 (standardized regression coefficient -0.11, P < .001) and interleukin-6 (standardized regression coefficient -0.09, P = .01). In contrast, there was no association between plasma levels of marine n-3 PUFAs and the anti-inflammatory mediator interleukin-10. CONCLUSIONS: In this renal transplant cohort, inverse associations between plasma levels of marine n-3 PUFAs and markers of inflammation were demonstrated.