Publications by authors named "Tom Eirik Mollnes"

185 Publications

Investigating novel biomarkers of immune activation and modulation in the context of sedentary behaviour: a multicentre prospective ischemic stroke cohort study.

BMC Neurol 2021 Aug 16;21(1):318. Epub 2021 Aug 16.

Department of Geriatric Medicine, Oslo University Hospital, Ullevaal, Oslo, Norway.

Background: Sedentary behaviour is associated with disease, but the molecular mechanisms are not understood. Valid biomarkers with predictive and explanatory properties are required. Therefore, we have investigated traditional and novel biomarkers of inflammation and immune modulation and their association to objectively measured sedentary behaviour in an ischemic stroke population.

Methods: Patients admitted to hospital with acute ischemic stroke were included in the multicentre Norwegian Cognitive Impairment After Stroke (Nor-COAST) study (n = 815). For this sub-study (n = 257), sedentary behaviour was registered 3 months after stroke using position transition data from the body-worn sensor, ActivPal®. Blood samples were analysed for high sensitive C-reactive protein (hsCRP), the cytokines interleukin-6 (IL-6) and 10 (IL-10), neopterin, tryptophan (Trp), kynurenine (kyn), kynurenic acid (KA), and three B6 vitamers, pyridoxal 5'-phosphate (PLP), pyridoxal (PL), and pyridoxic acid (PA). The kynurenine/tryptophan ratio (KTR) and the pyridoxic acid ratio index (PAr = PA: PL + PLP) were calculated.

Results: Of the 815 patients included in the main study, 700 attended the three-month follow-up, and 257 fulfilled the inclusion criteria for this study. Sedentary time was significantly associated with levels of hsCRP, IL-6, neopterin, PAr-index, and KA adjusted for age, sex, waist circumference, and creatinine. In a fully adjusted model including all the significant biomarkers except hsCRP (because of missing values), sedentary time was independently positively associated with the PAr-index and negatively with KA. We did not find an association between sedentary behaviour, IL-10, and KTR.

Conclusions: The PAr-index is known to capture several modes of inflammation and has previously shown predictive abilities for future stroke. This novel result indicates that the PAr-index could be a useful biomarker in future studies on sedentary behaviour and disease progression. KA is an important modulator of inflammation, and this finding opens new and exciting pathways to understand the hazards of sedentary behaviour.

Trial Registration: The study was registered at Clinicaltrials.gov ( NCT02650531 ). First posted 08/01/2016.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s12883-021-02343-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8365944PMC
August 2021

A Conformational Change of Complement C5 Is Required for Thrombin-Mediated Cleavage, Revealed by a Novel Ex Vivo Human Whole Blood Model Preserving Full Thrombin Activity.

J Immunol 2021 09 11;207(6):1641-1651. Epub 2021 Aug 11.

Department of Immunology, University of Oslo and Oslo University Hospital Rikshospitalet, Oslo, Norway;

Thrombin activation of C5 connects thrombosis to inflammation. Complement research in whole blood ex vivo necessitates anticoagulation, which potentially interferes with the inflammatory modulation by thrombin. We challenged the concept of thrombin as an activator of native C5 by analyzing complement activation and C5 cleavage in human whole blood anticoagulated with Gly-Pro-Arg-Pro (GPRP), a peptide targeting fibrin polymerization downstream of thrombin, allowing complete endogenous thrombin generation. GPRP dose-dependently inhibited coagulation but allowed for platelet activation in accordance with thrombin generation. Spontaneous and bacterial-induced complement activation by and , analyzed at the level of C3 and C5, were similar in blood anticoagulated with GPRP and the thrombin inhibitor lepirudin. In the GPRP model, endogenous thrombin, even at supra-physiologic concentrations, did not cleave native C5, despite efficiently cleaving commercially sourced purified C5 protein, both in buffer and when added to C5-deficient serum. In normal serum, only exogenously added, commercially sourced C5 was cleaved, whereas the native plasma C5 remained intact. Crucially, affinity-purified C5, eluted under mild conditions using an MgCl solution, was not cleaved by thrombin. Acidification of plasma to pH ≤ 6.8 by hydrochloric or lactic acid induced a C5 antigenic change, nonreversible by pH neutralization, that permitted cleavage by thrombin. Circular dichroism on purified C5 confirmed the structural change during acidification. Thus, we propose that pH-induced conformational change allows thrombin-mediated cleavage of C5 and that, contrary to previous reports, thrombin does not cleave plasma C5 in its native form, suggesting that thrombin cleavage of C5 may be restricted to certain pathophysiological conditions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4049/jimmunol.2001471DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8428748PMC
September 2021

Elevated plasma concentration of complement factor C5 is associated with risk of future venous thromboembolism.

Blood 2021 Aug 2. Epub 2021 Aug 2.

Division of Internal Medicine, University Hospital of North Norway, Tromsø, Norway.

The role of complement in the pathogenesis of venous thromboembolism (VTE) is unclear. We aimed to (i) investigate whether plasma complement component C5 levels are influenced by genetic variants or chronic inflammation, and (ii) investigate the association between plasma C5 and risk of future VTE in a nested case-control study with 415 VTE patients and 848 age- and sex-matched controls derived from the Tromsø study. Plasma C5 levels were measured at inclusion. Odds ratios (ORs) with 95% confidence intervals (95% CI) for provoked and unprovoked VTE across tertiles of C5 concentrations were estimated using logistic regression. C-reactive protein (CRP) was adjusted for as a proxy for general inflammation. Whole exome sequencing and protein quantitative trait loci analyses were performed to assess genetic influence on C5 concentrations. There was no association between genome-wide or C5-related gene variants and C5 levels. The association between plasma C5 levels and VTE risk displayed a threshold effect, where subjects with C5 levels above the lowest tertile had increased VTE risk. Subjects in tertile 3 (highest C5 levels) had an age and sex-adjusted OR of 1.45 (95% CI 1.07-1.96) compared to tertile 1 (lowest). This was more pronounced for unprovoked VTE (OR 1.70, 95% CI 1.11-2.60). Adjustments for body mass index and CRP had minor impact on risk estimates. The ORs increased substantially with shorter time between blood sampling and VTE event. In conclusion, plasma C5 was associated with risk of future VTE. C5 levels were not genetically regulated and only slightly influenced by chronic inflammation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1182/blood.2021010822DOI Listing
August 2021

Vitamin C, Hydrocortisone and The Combination Thereof Significantly Inhibited Two of Nine Inflammatory Markers Induced by Escherichia Coli but not by Staphylococcus Aureus - When Incubated in Human Whole Blood.

Shock 2021 Jul 8. Epub 2021 Jul 8.

Department of Immunology, Oslo University Hospital and University of Oslo, Oslo, Norway Research Laboratory, Nordland Hospital, Bodø and Faculty of Health Sciences, K. G. Jebsen Center, University of Tromsø, Norway Division of Emergencies and Critical Care, Oslo University Hospital, Oslo, Norway Institute of Hygiene and Medical Microbiology, Medical University of Innsbruck, Innsbruck, Austria Center of Molecular Inflammation Research, Norwegian University of Science and Technology, Trondheim, Norway.

Abstract: Vitamin C combined with hydrocortisone is increasingly being used to treat septic patients, even though this treatment regimen is based on questionable evidence. When used, a marked effect on key players of innate immunity would be expected, as sepsis is featured by a dysregulated immune response.Here, we explored the effect of vitamin C and hydrocortisone alone and combined, in an ex vivo human whole-blood model of Escherichia coli- or Staphylococcus aureus-induced inflammation. Inflammatory markers for activation of complement (TCC), granulocytes (myeloperoxidase), platelets (β-thromboglobulin), cytokines (TNF, IL-1β, IL6 and IL-8) and leukocytes (CD11b and oxidative burst) were quantified, by ELISA, multiplex technology and flow cytometry.In E. coli- and S. aureus-stimulated whole blood, a broad dose-titration of vitamin C and hydrocortisone alone did not lead to dose-response effects for the central innate immune mediators TCC and IL-6. Hence, the clinically relevant doses were used further. Compared to the untreated control sample, 2 of the 9 biomarkers induced by Escherichia coli were reduced by hydrocortisone and/or vitamin C. TNF was reduced by hydrocortisone alone (19%, p  = 0.01) and by the combination (31%, p  = 0.01). The oxidative burst of monocytes and granulocytes was reduced for both drugs alone and their combination, (ranging 8-19%, p < 0.05). Using Staphylococcus aureus, neither of the drugs, alone nor in combination, had any effects on the 9 biomarkers.In conclusion, despite the limitation of the ex vivo model, the effect of vitamin C and hydrocortisone on bacteria-induced inflammatory response in human whole blood is limited and following the clinical data.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/SHK.0000000000001834DOI Listing
July 2021

Complement activation is associated with poor outcome after out-of-hospital cardiac arrest.

Resuscitation 2021 Sep 11;166:129-136. Epub 2021 Jun 11.

Dept. of Immunology, University of Oslo and Oslo University Hospital, Oslo, Norway; Dept. of Anaesthesiology, Division of Emergencies and Critical Care, Oslo University Hospital, Oslo, Norway; Institute of Clinical Medicine, University of Oslo, Oslo, Norway; Dept. of Research and Development, Division of Emergencies and Critical Care, Oslo University Hospital, Oslo, Norway. Electronic address:

Background: Cardiopulmonary resuscitation after cardiac arrest initiates a whole-body ischemia-reperfusion injury, which may activate the innate immune system, including the complement system. We hypothesized that complement activation and subsequent release of soluble endothelial activation markers were associated with cerebral outcome including death.

Methods: Outcome was assessed at six months and defined by cerebral performance category scale (1-2; good outcome, 3-5; poor outcome including death) in 232 resuscitated out-of-hospital cardiac arrest patients. Plasma samples obtained at admission and day three were analysed for complement activation products C3bc, the soluble terminal complement complex (sC5b-9), and soluble CD14. Endothelial cell activation was measured by soluble markers syndecan-1, sE-selectin, thrombomodulin, and vascular cell adhesion molecule.

Results: Forty-nine percent of the patients had good outcome. C3bc and sC5b-9 were significantly higher at admission compared to day three (p < 0.001 for both) and in patients with poor compared to good outcome (p = 0.03 and p < 0.001, respectively). Unadjusted, higher sC5b-9 at admission was associated with poor outcome (odds ratio 1.08 (95% CI 1.01-1.14), p = 0.024). Adjusted, sC5b-9 was still associated with outcome, but the association became non-significant when time to return-of-spontaneous-circulation above 25 min was included as a covariate. Endothelial cell activation markers increased from admission to day three, but only sE-selectin and thrombomodulin were significantly higher in patients with poor versus good outcome (p = 0.004 and p = 0.03, respectively) and correlated to sCD14 and sC5b-9/C3bc, respectively.

Conclusion: Complement system activation, reflected by sC5b-9 at admission, leading to subsequent endothelial cell activation, was associated with poor outcome in out-of-hospital cardiac arrest patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.resuscitation.2021.05.038DOI Listing
September 2021

Patient Derived Colonoids as Drug Testing Platforms-Critical Importance of Oxygen Concentration.

Front Pharmacol 2021 13;12:679741. Epub 2021 May 13.

Department of Clinical and Molecular Medicine (IKOM), Faculty of Medicine and Health Sciences, NTNU- Norwegian University of Science and Technology, Trondheim, Norway.

Treatment of inflammatory bowel disease (IBD) is challenging, with a series of available drugs each helping only a fraction of patients. Patients may face time-consuming drug trials while the disease is active, thus there is an unmet need for biomarkers and assays to predict drug effect. It is well known that the intestinal epithelium is an important factor in disease pathogenesis, exhibiting physical, biochemical and immunologic driven barrier dysfunctions. One promising test system to study effects of existing or emerging IBD treatments targeting intestinal epithelial cells (IECs) is intestinal organoids ("mini-guts"). However, the fact that healthy intestinal epithelium is in a physiologically hypoxic state has largely been neglected, and studies with intestinal organoids are mainly performed at oxygen concentration of 20%. We hypothesized that lowering the incubator oxygen level from 20% to 2% would recapitulate better the physiological environment of colonic epithelial cells and enhance the translational value of intestinal organoids as a drug testing platform. In the present study we examine the effects of the key IBD cytokines and drug targets TNF/IL17 on human colonic organoids (colonoids) under atmospheric (20%) or reduced (2%) O. We show that colonoids derived from both healthy controls and IBD-patients are viable and responsive to IBD-relevant cytokines at 2% oxygen. Because chemokine release is one of the important immunoregulatory traits of the epithelium that may be fine-tuned by IBD-drugs, we also examined chemokine expression and release at different oxygen concentrations. We show that chemokine responses to TNF/IL17 in organoids display similarities to inflamed epithelium in IBD-patients. However, inflammation-associated genes induced by TNF/IL17 were attenuated at low oxygen concentration. We detected substantial oxygen-dependent differences in gene expression in untreated as well as TNF/IL17 treated colonoids in all donors. Further, for some of the IBD-relevant cytokines differences between colonoids from healthy controls and IBD patients were more pronounced in 2% O than 20% O. Our results strongly indicate that an oxygen concentration similar to the epithelial cell environment is of essence in experimental pharmacology.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fphar.2021.679741DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8156408PMC
May 2021

Increased Local Inflammatory Response to MOC31PE Immunotoxin After Cytoreductive Surgery and Hyperthermic Intraperitoneal Chemotherapy.

Ann Surg Oncol 2021 Sep 21;28(9):5252-5262. Epub 2021 May 21.

Department of Gastroenterological Surgery, Oslo University Hospital The Radium Hospital, Oslo, Norway.

Background: Despite extensive cytoreductive surgery and hyperthermic intraperitoneal chemotherapy (CRS-HIPEC), most patients with resectable peritoneal metastases from colorectal cancer experience disease relapse. MOC31PE immunotoxin is being explored as a novel treatment option for these patients. MOC31PE targets the cancer-associated epithelial cell adhesion molecule, and kills cancer cells by distinct mechanisms, simultaneously causing immune activation by induction of immunogenic cell death (ICD).

Methods: Systemic and local cytokine responses were analyzed in serum and intraperitoneal fluid samples collected the first three postoperative days from clinically comparable patients undergoing CRS-HIPEC with (n = 12) or without (n = 26) intraperitoneal instillation of MOC31PE. A broad panel of 27 pro- and antiinflammatory interleukins, chemokines, interferons, and growth factors was analyzed using multiplex technology.

Results: The time course and magnitude of the systemic and local postoperative cytokine response after CRS-HIPEC were highly compartmentalized, with modest systemic responses contrasting substantial intraperitoneal responses. Administration of MOC31PE resulted in changes that were broader and of higher magnitude compared with CRS-HIPEC alone. Significantly increased levels of innate proinflammatory cytokines, such as interleukin (IL)-6, IL-1β, and tumor necrosis factor (TNF) as well as an interesting time response curve for the strong T-cell stimulator interferon (IFN)-γ and its associated chemokine interferon gamma-induced protein/chemokine (C-X-C motif) ligand 10 (IP-10) were detected, all associated with ICD.

Conclusions: Our study revealed a predominately local rather than systemic inflammatory response to CRS-HIPEC, which was strongly enhanced by MOC31PE treatment. The MOC31PE-induced intraperitoneal inflammatory reaction could contribute to improve remnant cancer cell killing, but the mechanisms remain to be elucidated in future studies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1245/s10434-021-10022-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8349350PMC
September 2021

Innate Immune Invisible Ultrasmall Gold Nanoparticles-Framework for Synthesis and Evaluation.

ACS Appl Mater Interfaces 2021 May 12;13(20):23410-23422. Epub 2021 May 12.

Department of Chemical Engineering and Biotechnology, University of Cambridge, Cambridge CB3 0AS, U.K.

Nanomedicine is seen as a potential central player in the delivery of personalized medicine. Biocompatibility issues of nanoparticles have largely been resolved over the past decade. Despite their tremendous progress, less than 1% of applied nanosystems can hit their intended target location, such as a solid tumor, and this remains an obstacle to their full ability and potential with a high translational value. Therefore, achieving immune-tolerable, blood-compatible, and biofriendly nanoparticles remains an unmet need. The translational success of nanoformulations from bench to bedside involves a thorough assessment of their design, compatibility beyond cytotoxicity such as immune toxicity, blood compatibility, and immune-mediated destruction/rejection/clearance profile. Here, we report a one-pot process-engineered synthesis of ultrasmall gold nanoparticles (uGNPs) suitable for better body and renal clearance delivery of their payloads. We have obtained uGNP sizes of as low as 3 nm and have engineered the synthesis to allow them to be accurately sized (almost nanometer by nanometer). The synthesized uGNPs are biocompatible and can easily be functionalized to carry drugs, peptides, antibodies, and other therapeutic molecules. We have performed cell viability assays, immunotoxicity assays, inflammatory cytokine analysis, a complement activation study, and blood coagulation studies with the uGNPs to confirm their safety. These can help to set up a long-term safety-benefit framework of experimentation to reveal whether any designed nanoparticles are immune-tolerable and can be used as payload carriers for next-generation vaccines, chemotherapeutic drugs, and theranostic agents with better body clearance ability and deep tissue penetration.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1021/acsami.1c02834DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8289183PMC
May 2021

Tuning gelatin-based hydrogel towards bioadhesive ocular tissue engineering applications.

Bioact Mater 2021 Nov 17;6(11):3947-3961. Epub 2021 Apr 17.

Massachusetts Eye and Ear and Schepens Eye Research Institute, Department of Ophthalmology, Harvard Medical School, Boston, MA, USA.

Gelatin based adhesives have been used in the last decades in different biomedical applications due to the excellent biocompatibility, easy processability, transparency, non-toxicity, and reasonable mechanical properties to mimic the extracellular matrix (ECM). Gelatin adhesives can be easily tuned to gain different viscoelastic and mechanical properties that facilitate its ocular application. We herein grafted glycidyl methacrylate on the gelatin backbone with a simple chemical modification of the precursor, utilizing epoxide ring-opening reactions and visible light-crosslinking. This chemical modification allows the obtaining of an elastic protein-based hydrogel (GELGYM) with excellent biomimetic properties, approaching those of the native tissue. GELGYM can be modulated to be stretched up to 4 times its initial length and withstand high tensile stresses up to 1.95 MPa with compressive strains as high as 80% compared to Gelatin-methacryloyl (GeIMA), the most studied derivative of gelatin used as a bioadhesive. GELGYM is also highly biocompatible and supports cellular adhesion, proliferation, and migration in both 2 and 3-dimensional cell-cultures. These characteristics along with its super adhesion to biological tissues such as cornea, aorta, heart, muscle, kidney, liver, and spleen suggest widespread applications of this hydrogel in many biomedical areas such as transplantation, tissue adhesive, wound dressing, bioprinting, and drug and cell delivery.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bioactmat.2021.03.042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8080056PMC
November 2021

Quantification of Porcine Complement Activation Fragment C3a by a Neoepitope-Based Enzyme-Linked Immunosorbent Assay.

Methods Mol Biol 2021 ;2227:51-59

Department of Immunology, University of Oslo and Oslo University Hospital Rikshospitalet, Oslo, Norway.

Enzyme-linked immunosorbent assay (ELISA) enables fast and simple quantification of analytes in the pico- to nanogram range in complex samples. Here, we describe an ELISA for the detection of porcine C3a as a marker for complement activation. Antibody specificity is critical for a robust assay. This assay is based on a pair of antibodies specific for the porcine C3a molecule and thus does not react with native C3.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/978-1-0716-1016-9_5DOI Listing
June 2021

Increased complement activation 3 to 6 h after trauma is a predictor of prolonged mechanical ventilation and multiple organ dysfunction syndrome: a prospective observational study.

Mol Med 2021 04 8;27(1):35. Epub 2021 Apr 8.

Department of Immunology, Oslo University Hospital and University of Oslo, Oslo, Norway.

Background: Complement activation is a central mechanism in systemic inflammation and remote organ dysfunction following major trauma. Data on temporal changes of complement activation early after injury is largely missing. We aimed to describe in detail the kinetics of complement activation in individual trauma patients from admission to 10 days after injury, and the association with trauma characteristics and outcome.

Methods: In a prospective cohort of 136 trauma patients, plasma samples obtained with high time resolution (admission, 2, 4, 6, 8 h, and thereafter daily) were assessed for terminal complement complex (TCC). We studied individual TCC concentration curves and calculated a summary measure to obtain the accumulated TCC response 3 to 6 h after injury (TCC-AUC). Correlation analyses and multivariable linear regression analyses were used to explore associations between individual patients' admission TCC, TCC-AUC, daily TCC during the intensive care unit stay, trauma characteristics, and predefined outcome measures.

Results: TCC concentration curves showed great variability in temporal shapes between individuals. However, the highest values were generally seen within the first 6 h after injury, before they subsided and remained elevated throughout the intensive care unit stay. Both admission TCC and TCC-AUC correlated positively with New Injury Severity Score (Spearman's rho, p-value 0.31, 0.0003 and 0.21, 0.02) and negatively with admission Base Excess (- 0.21, 0.02 and - 0.30, 0.001). Multivariable analyses confirmed that deranged physiology was an important predictor of complement activation. For patients without major head injury, admission TCC and TCC-AUC were negatively associated with ventilator-free days. TCC-AUC outperformed admission TCC as a predictor of Sequential Organ Failure Assessment score at day 0 and 4.

Conclusions: Complement activation 3 to 6 h after injury was a better predictor of prolonged mechanical ventilation and multiple organ dysfunction syndrome than admission TCC. Our data suggest that the greatest surge of complement activation is found within the first 6 h after injury, and we argue that this time period should be in focus in the design of future experimental studies and clinical trials using complement inhibitors.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s10020-021-00286-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8028580PMC
April 2021

Combined blockade of complement C5 and TLR co-receptor CD14 synergistically inhibits pig-to-human corneal xenograft induced innate inflammatory responses.

Acta Biomater 2021 06 27;127:169-179. Epub 2021 Mar 27.

Department of Immunology, Oslo University Hospital and University of Oslo, Oslo, Norway; Centre of Molecular Inflammation Research, and Department of Clinical and Molecular Medicine, Norwegian University of Science and Technology, Trondheim, Norway; Research Laboratory, Nordland Hospital, Bodo, Norway; Faculty of Health Sciences, K. G. Jebsen Thrombosis Research and Expertise Center, University of Tromsø, Tromso, Norway.

Inadequate supplies of donor corneas have evoked an escalating interest in corneal xenotransplantation. However, innate immune responses contribute significantly to the mechanism of xenograft rejection. We hypothesized that complement component C5 and TLR co-receptor CD14 inhibition would inhibit porcine cornea induced innate immune responses. Therefore, we measured cytokine release in human blood, induced by three forms of corneal xenografts with or without inhibitors. Native porcine cornea (NPC) induced interleukins (IL-1β, IL-2, IL-6, IL-8, IL-1ra), chemokines (MCP-1, MIP-1α, MIP-1β) and other cytokines (TNF, G-CSF, INF-γ, FGF-basic). Decellularized (DPC) and gamma-irradiated cornea (g-DPC) elevated the release of those cytokines. C5-blockade by eculizumab inhibited all the cytokines except G-CSF when induced by NPC. However, C5-blockade failed to reduce DPC and g-DPC induced cytokines. Blockade of CD14 inhibited DPC-induced cytokines except for IL-8, MCP-1, MIP-1α, and G-CSF, while it inhibited all of them when induced by g-DPC. Combined blockade of C5 and CD14 inhibited the maximum number of cytokines regardless of the xenograft type. Finally, by using the TLR4 specific inhibitor Eritoran, we showed that TLR4 activation was the basis for the CD14 effect. Thus, blockade of C5, when combined with TLR4 inhibition, may have therapeutic potential in pig-to-human corneal xenotransplantation. STATEMENT OF SIGNIFICANCE: Bio-engineered corneal xenografts are on the verge of becoming a viable alternative to allogenic human-donor-cornea, but the host's innate immune response is still a critical barrier for graft acceptance. By overruling this barrier, limited graft availability would no longer be an issue for treating corneal diseases. We showed that the xenograft induced inflammation is initiated by the complement system and toll-like receptor activation. Intriguingly, the inflammatory response was efficiently blocked by simultaneously targeting bottleneck molecules in the complement system (C5) and the TLR co-receptor CD14 with pharmaceutical inhibitors. We postulate that a combination of C5 and CD14 inhibition could have a great therapeutic potential to overcome the immunologic barrier in pig-to-human corneal xenotransplantation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.actbio.2021.03.047DOI Listing
June 2021

Rapid systemic surge of IL-33 after severe human trauma: a prospective observational study.

Mol Med 2021 03 26;27(1):29. Epub 2021 Mar 26.

Department of Anaesthesiology, Division of Emergencies and Critical Care, Oslo University Hospital Ullevål, Oslo, Norway.

Background: Alarmins are considered proximal mediators of the immune response after tissue injury. Understanding their biology could pave the way for development of new therapeutic targets and biomarkers in human disease, including multiple trauma. In this study we explored high-resolution concentration kinetics of the alarmin interleukin-33 (IL-33) early after human trauma.

Methods: Plasma samples were serially collected from 136 trauma patients immediately after hospital admission, 2, 4, 6, and 8 h thereafter, and every morning in the ICU. Levels of IL-33 and its decoy receptor sST2 were measured by immunoassays.

Results: We observed a rapid and transient surge of IL-33 in a subset of critically injured patients. These patients had more widespread tissue injuries and a greater degree of early coagulopathy. IL-33 half-life (t) was 1.4 h (95% CI 1.2-1.6). sST2 displayed a distinctly different pattern with low initial levels but massive increase at later time points.

Conclusions: We describe for the first time early high-resolution IL-33 concentration kinetics in individual patients after trauma and correlate systemic IL-33 release to clinical data. These findings provide insight into a potentially important axis of danger signaling in humans.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s10020-021-00288-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8004436PMC
March 2021

The lytic polysaccharide monooxygenase CbpD promotes Pseudomonas aeruginosa virulence in systemic infection.

Nat Commun 2021 02 23;12(1):1230. Epub 2021 Feb 23.

Faculty of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences (NMBU), Ås, Norway.

The recently discovered lytic polysaccharide monooxygenases (LPMOs), which cleave polysaccharides by oxidation, have been associated with bacterial virulence, but supporting functional data is scarce. Here we show that CbpD, the LPMO of Pseudomonas aeruginosa, is a chitin-oxidizing virulence factor that promotes survival of the bacterium in human blood. The catalytic activity of CbpD was promoted by azurin and pyocyanin, two redox-active virulence factors also secreted by P. aeruginosa. Homology modeling, molecular dynamics simulations, and small angle X-ray scattering indicated that CbpD is a monomeric tri-modular enzyme with flexible linkers. Deletion of cbpD rendered P. aeruginosa unable to establish a lethal systemic infection, associated with enhanced bacterial clearance in vivo. CbpD-dependent survival of the wild-type bacterium was not attributable to dampening of pro-inflammatory responses by CbpD ex vivo or in vivo. Rather, we found that CbpD attenuates the terminal complement cascade in human serum. Studies with an active site mutant of CbpD indicated that catalytic activity is crucial for virulence function. Finally, profiling of the bacterial and splenic proteomes showed that the lack of this single enzyme resulted in substantial re-organization of the bacterial and host proteomes. LPMOs similar to CbpD occur in other pathogens and may have similar immune evasive functions.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-021-21473-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7902821PMC
February 2021

Hemorrhagic Shock Induces a Rapid Transcriptomic Shift of the Immune Balance in Leukocytes after Experimental Multiple Injury.

Mediators Inflamm 2021 27;2021:6654318. Epub 2021 Jan 27.

Institute of Clinical and Experimental Trauma Immunology, University Hospital Ulm, Ulm, Germany.

The immune response following trauma represents a major driving force of organ dysfunction and poor outcome. Therefore, we investigated the influence of an additional hemorrhagic shock (HS) on the early posttraumatic immune dysbalance in the whole population of blood leukocytes. A well-established murine polytrauma (PT) model with or without an additional pressure-controlled HS (mean arterial pressure of 30 mmHg (±5 mmHg) for 60 mins, afterwards fluid resuscitation with balanced electrolyte solution four times the volume of blood drawn) was used. C57BL/6 mice were randomized into a control, PT, and PT + HS group with three animals in each group. Four hours after trauma, corresponding to three hours after induction of hemorrhage, RNA was isolated from all peripheral blood leukocytes, and a microarray analysis was performed. Enrichment analysis was conducted on selected genes strongly modulated by the HS. After additional HS in PT mice, the gene expression of pathways related to the innate immunity, such as IL-6 production, neutrophil chemotaxis, cell adhesion, and toll-like receptor signaling was upregulated, whereas pathways of the adaptive immune system, such as B- and T-cell activation as well as the MHC class II protein complex, were downregulated. These results demonstrate that an additional HS plays an important role in the immune dysregulation early after PT by shifting the balance to increased innate and reduced adaptive immune responses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1155/2021/6654318DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7857921PMC
September 2021

The allosteric modulation of complement C5 by knob domain peptides.

Elife 2021 02 11;10. Epub 2021 Feb 11.

Department of Biology and Biochemistry, University of Bath, Bath, United Kingdom.

Bovines have evolved a subset of antibodies with ultra-long heavy chain complementarity determining regions that harbour cysteine-rich knob domains. To produce high-affinity peptides, we previously isolated autonomous 3-6 kDa knob domains from bovine antibodies. Here, we show that binding of four knob domain peptides elicits a range of effects on the clinically validated drug target complement C5. Allosteric mechanisms predominated, with one peptide selectively inhibiting C5 cleavage by the alternative pathway C5 convertase, revealing a targetable mechanistic difference between the classical and alternative pathway C5 convertases. Taking a hybrid biophysical approach, we present C5-knob domain co-crystal structures and, by solution methods, observed allosteric effects propagating >50 Å from the binding sites. This study expands the therapeutic scope of C5, presents new inhibitors, and introduces knob domains as new, low molecular weight antibody fragments, with therapeutic potential.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.7554/eLife.63586DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7972453PMC
February 2021

Sputter Deposition of Titanium on Poly(Methyl Methacrylate) Enhances Corneal Biocompatibility.

Transl Vis Sci Technol 2020 12 23;9(13):41. Epub 2020 Dec 23.

Disruptive Technology Laboratory and Schepens Eye Research Institute, Massachusetts Eye and Ear; Department of Ophthalmology, Harvard Medical School, Boston, MA, USA.

Purpose: To evaluate titanium (Ti) sputtering of the poly(methyl methacrylate) (PMMA) stem of the Boston Keratoprosthesis (BK) as a method to enhance interfacial adhesion between the PMMA and the recipient corneal tissue.

Methods: PMMA specimens were plasma treated with Ar/O and coated with Ti using a DC magnetron sputtering instrument. The topography and hydrophilicity of the surfaces were characterized using atomic force microscopy and a water contact angle instrument, respectively. Scratch hardness and adhesion of the Ti film were measured using a mechanical tester. Biocompatibility assessments were performed using cultured human corneal fibroblasts and whole blood ex vivo. The optical quality of the Ti sputtered BK was evaluated using a custom-made optical bench.

Results: By contact angle studies, the Ti coating improved PMMA hydrophilicity to match that of medical-grade Ti (Ti-6Al-4V-ELI). Ti sputtering of contact surfaces resulted in a plate-like morphology with increased surface roughness, without impacting the transparency of the BK optical component. Scratch testing indicated that the mechanical behavior of the Ti coating was similar to that of casted Ti, and the coating was stable in pull-off adhesion testing. Sputtered Ti film was highly biocompatible based on tests of cell viability, adhesion, proliferation, differentiation, collagen deposition, and keratocan expression, the properties of which exceeded those of uncoated PMMA and did not induce increased complement activation.

Conclusions: Titanium coating of the BK stem generated a mechanically and biologically favorable interface, which may help to enhance corneal stromal adhesion and biocompatibility.

Translational Relevance: Improving the biocompatibility of the BK PMMA stem may improve long-term outcomes of implantation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1167/tvst.9.13.41DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7774111PMC
December 2020

Increased interleukin-6 and macrophage chemoattractant protein-1 are associated with respiratory failure in COVID-19.

Sci Rep 2020 12 10;10(1):21697. Epub 2020 Dec 10.

Department of Infectious Diseases, Oslo University Hospital, Oslo, Norway.

In SARS-CoV-2 infection there is an urgent need to identify patients that will progress to severe COVID-19 and may benefit from targeted treatment. In this study we analyzed plasma cytokines in COVID-19 patients and investigated their association with respiratory failure (RF) and treatment in Intensive Care Unit (ICU). Hospitalized patients (n = 34) with confirmed COVID-19 were recruited into a prospective cohort study. Clinical data and blood samples were collected at inclusion and after 2-5 and 7-10 days. RF was defined as PaO2/FiO2 ratio (P/F) < 40 kPa. Plasma cytokines were analyzed by a Human Cytokine 27-plex assay. COVID-19 patients with RF and/or treated in ICU showed overall increased systemic cytokine levels. Plasma IL-6, IL-8, G-CSF, MCP-1, MIP-1α levels were negatively correlated with P/F, whereas combinations of IL-6, IP-10, IL-1ra and MCP-1 showed the best association with RF in ROC analysis (AUC 0.79-0.80, p < 0.05). During hospitalization the decline was most significant for IP-10 (p < 0.001). Elevated levels of pro-inflammatory cytokines were present in patients with severe COVID-19. IL-6 and MCP-1 were inversely correlated with P/F with the largest AUC in ROC analyses and should be further explored as biomarkers to identify patients at risk for severe RF and as targets for improved treatment strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-020-78710-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7729930PMC
December 2020

Avoiding ambient air in test tubes during incubations of human whole-blood minimizes complement background activation.

J Immunol Methods 2020 12 6;487:112876. Epub 2020 Oct 6.

Department of Anesthesia and Intensive Care Medicine, Surgical Clinic, Nordland Hospital, Bodø, Norway; Institute of Clinical Medicine, University of Tromsø, Tromsø, Norway; Faculty of Nursing and Health Sciences, Nord University, Bodø, Norway; Faculty of Medicine, Institute of Clinical Medicine, University of Oslo, Norway.

Background: In vitro, the complement system can be studied in test tubes incubated with anticoagulated human whole-blood. Background activation of complement may mask inflammatory signals. Air bubbles are known to activate complement. We examined if removing ambient air from test tubes before incubation reduced background complement activation.

Methods: Blood from twelve donors was anticoagulated with the thrombin inhibitor lepirudin and incubated with either no air, ambient air or air bubbles in polypropylene tubes at 37 °C for 180 min on a roller mixer. After incubation, EDTA was added, plasma isolated and analyzed for seven complement activation products using ELISA. Results are presented as means with 95% confidence intervals.

Results: Blood incubated without air had significantly lower complement activation compared to blood incubated with ambient air; C4d 273 (192-364) vs. 379 (263-494) ng/mL (p = 0.002), C4bc 8.2 (4.1-13) vs. 12 (3.2-21) CAU/mL (p = 0.01), C3a 1351 (873-1838) vs. 2944 (2315-3572) ng/mL (p = 0.0005), C3bc 31 (17-46) vs. 68 (52-84) CAU/mL (p = 0.002), C3bBbP 134 (97-171) vs. 427 (358-506) CAU/mL (p < 0.0001), C5a 3.5 (1.9-5 0.2) vs. 15 (1.8-27)) ng/mL (p = 0.003), TCC 4.6 (2.8-6.3) vs. 9.9 (7.3-12) CAU/mL (p = 0.006). At the end of the experiment blood incubated with air bubbles had a higher complement activation than blood incubated with ambient air with an average 26 fold increase (range 1.6-59) from baseline of all activation products; C4d 551 (337-766) ng/mL, C4bc 21 (5.0-36) CAU/mL, C3a 3983 (3518-4448) ng/mL, C4bc 103 (86-121) CAU/mL, C3bBbP 626 (543-708) CAU/mL, C5a 10 (2.8-18) ng/mL and TCC 10 (6.0-14) CAU/mL.

Conclusion: Avoiding air in test tubes during whole-blood experiments reduced background complement activation substantially and represents an important improvement to the lepirudin whole-blood model. This could also apply to other in vitro models.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.jim.2020.112876DOI Listing
December 2020

A Complement C3-Specific Nanobody for Modulation of the Alternative Cascade Identifies the C-Terminal Domain of C3b as Functional in C5 Convertase Activity.

J Immunol 2020 10 16;205(8):2287-2300. Epub 2020 Sep 16.

Department of Molecular Biology and Genetics, Aarhus University, DK-8000 Aarhus, Denmark;

The complement system is an intricate cascade of the innate immune system and plays a key role in microbial defense, inflammation, organ development, and tissue regeneration. There is increasing interest in developing complement regulatory and inhibitory agents to treat complement dysfunction. In this study, we describe the nanobody hC3Nb3, which is specific for the C-terminal C345c domain of human and mouse complement component C3/C3b/C3c and potently inhibits C3 cleavage by the alternative pathway. A high-resolution structure of the hC3Nb3-C345c complex explains how the nanobody blocks proconvertase assembly. Surprisingly, although the nanobody does not affect classical pathway-mediated C3 cleavage, hC3Nb3 inhibits classical pathway-driven hemolysis, suggesting that the C-terminal domain of C3b has an important function in classical pathway C5 convertase activity. The hC3Nb3 nanobody binds C3 with low nanomolar affinity in an SDS-resistant complex, and the nanobody is demonstrated to be a powerful reagent for C3 detection in immunohistochemistry and flow cytometry. Overall, the hC3Nb3 nanobody represents a potent inhibitor of both the alternative pathway and the terminal pathway, with possible applications in complement research, diagnostics, and therapeutics.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4049/jimmunol.2000752DOI Listing
October 2020

Soluble collectin-12 mediates C3-independent docking of properdin that activates the alternative pathway of complement.

Elife 2020 09 10;9. Epub 2020 Sep 10.

The Laboratory of Molecular Medicine, Department of Clinical Immunology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.

Properdin stabilizes the alternative C3 convertase (C3bBb), whereas its role as pattern-recognition molecule mediating complement activation is disputed for decades. Previously, we have found that soluble collectin-12 (sCL-12) synergizes complement alternative pathway (AP) activation. However, whether this observation is C3 dependent is unknown. By application of the C3-inhibitor Cp40, we found that properdin in normal human serum bound to solely in a C3b-dependent manner. Cp40 also prevented properdin binding when properdin-depleted serum reconstituted with purified properdin was applied, in analogy with the findings achieved by C3-depleted serum. However, when opsonized with sCL-12, properdin bound in a C3-independent manner exclusively via its tetrameric structure and directed in situ C3bBb assembly. In conclusion, a prerequisite for properdin binding and in situ C3bBb assembly was the initial docking of sCL-12. This implies a new important function of properdin in host defense bridging pattern recognition and specific AP activation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.7554/eLife.60908DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7511233PMC
September 2020

Anti-inflammatory effects of non-statin low-density lipoprotein cholesterol-lowering drugs: an unused potential?

Scand Cardiovasc J 2020 Oct 5;54(5):274-279. Epub 2020 Jun 5.

Coronary Care Unit, Division of Internal Medicine, Nordland Hospital, Bodø, Norway.

. Inflammatory responses are closely knit with low-density lipoprotein (LDL)-cholesterol in driving atherosclerosis. Even if LDL-cholesterol is causative to atherosclerotic diseases and LDL-cholesterol lowering reduces hard clinical endpoints, there is a residual risk for clinical events, possibly driven by inflammatory processes, in accordance with its role in autoimmune diseases. . As LDL-cholesterol treatment targets are reduced, the use of non-statin lipid-lowering drugs will probably increase. Atherosclerotic plaques evolve through lipid infiltration and modification in the intima, furthermore infiltration of cells including monocytes, macrophages, T-lymphocytes and neutrophils initiating inflammatory signaling. Here we briefly review inflammation in atherosclerosis and the effects of the non-statin lipid-lowering drugs on inflammation. The review is limited to the most common non-statin lipid lowering drugs, i.e. proprotein convertase subtilisin-kexin type 9 (PCSK9) inhibitors, bile acid sequestrants (BAS) and cholesterol absorption inhibitors. . PCSK9 inhibition is mostly studied together with statins and is associated with a reduction of pro-inflammatory cytokines. Furthermore, PCSK9 inhibitors seem to have an effect on monocyte migration trough CCR2. They also have an interaction with sirtuins, possibly offering a therapeutic target. BAS have several interesting effects on inflammation, including reduction of pro-inflammatory cytokines and a reduction of the number of infiltrating macrophages, however there are relatively few reports considering that these drugs have been on the market for decades. Ezetimibe also has effects on inflammation including reduction of pro-inflammatory cytokines and adhesion molecules, however these effects are usually accomplished in tandem with statins. . This topic adds an interesting piece to the puzzle of atherosclerosis, indicating that PCSK9 inhibition, BAS and ezetimibe all affect thromboinflammation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/14017431.2020.1775878DOI Listing
October 2020

Intestinal inflammatory profile shows increase in a diversity of biomarkers in irritable bowel syndrome.

Scand J Gastroenterol 2020 May 24;55(5):537-542. Epub 2020 Apr 24.

Research Group of Gastroenterology and Nutrition, Institute of Clinical Medicine, Norwegian Arctic University, Tromsø, Norway.

It has been proposed that irritable bowel syndrome (IBS) is a low-grade mucosal inflammatory disease. To characterize the intestinal inflammatory profile in IBS patients with or without fructose intolerance. Patients referred to colonoscopy with IBS complaints were screened for participation. IBS patients diagnosed according to the Rome II criteria and with no organic gastrointestinal disease were included in the study. One subgroup was patients included in a fructose-reduced diet study for 2 months with effects based on VAS symptom scores. Healthy controls were subjects under investigation of colorectal cancer screening with no IBS or other gastrointestinal diseases. All patients included had normal histology from rectum. Mucosal cytokines, chemokines and growth factors were measured by multiplex technology. Of 27 inflammatory markers tested in the mucosal tissue, 13 were significantly increased and none was significantly decreased in IBS as compared to controls. Significantly increased were the proinflammatory cytokines tumor necrosis factor, the typical TH1 markers IFNγ, IL-1β, IL-2 and RANTES, the typical TH2 markers IL-5 and IL-9, the TH17 marker IL-17, TNF, the pleiotropic IL-15, and the growth factors bFGF and GM-CSF. In IBS patients with fructose intolerance only IL-5 was significantly increased compared to patients without fructose intolerance. A dysregulated mucosal inflammatory profile with an increased level of TH1, TH2 and TH17 markers, and growth factors were observed in bowel mucosa in of IBS patients when compared to healthy controls.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/00365521.2020.1754455DOI Listing
May 2020

Complement Activation in 22q11.2 Deletion Syndrome.

J Clin Immunol 2020 04 9;40(3):515-523. Epub 2020 Mar 9.

Center for Rare Diseases, Division of Pediatric and Adolescent Medicine, Oslo University Hospital, Oslo, Norway.

The 22q11.2 deletion syndrome (22q11.2 del), also known as DiGeorge syndrome, is a genetic disorder with an estimated incidence of 1:3000 to 1:6000 births. These patients may suffer from affection of many organ systems with cardiac malformations, immunodeficiency, hypoparathyroidism, autoimmunity, palate anomalies, and psychiatric disorders being the most frequent. The importance of the complement system in 22q11.2 del has not been investigated. The objective of this study was to evaluate the complement system in relation to clinical and immunological parameters in patients. A national cohort of patients (n = 69) with a proven heterozygous deletion of chromosome 22q11.2 and a group of age and sex matched controls (n = 56) were studied. Functional capacity of the classical, lectin, and alternative pathways of the complement system as well as complement activation products C3bc and terminal complement complex (TCC) were accessed and correlated to clinical features. All patients in our study had normal complement activation in both classical and alternative pathways. The frequency of mannose-binding lectin deficiency was comparable to the normal population. The patients had significantly raised plasma levels of C3bc and a slight, but not significant, increase in TCC compared with controls. This increase was associated with the presence of psychiatric disorders in patients. The present study shows no complement deficiencies in 22q11.2 deletion syndrome. On the contrary, there are signs of increased complement activation in these patients. Complement activation is particularly associated with the presence of psychiatric disorders.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10875-020-00766-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7142058PMC
April 2020

Intestinal Epithelial Cells Express Immunomodulatory ISG15 During Active Ulcerative Colitis and Crohn's Disease.

J Crohns Colitis 2020 Jul;14(7):920-934

Department of Clinical and Molecular Medicine, NTNU-Norwegian University of Science and Technology, Trondheim, Norway.

Background And Aims: Intestinal epithelial cells [IECs] secrete cytokines that recruit immune cells to the mucosa and regulate immune responses that drive inflammation in inflammatory bowel disease [IBD]. However, experiments in patient-derived IEC models are still scarce. Here, we aimed to investigate how innate immunity and IEC-specific pattern recognition receptor [PRR] signalling can be involved in an enhanced type I interferon [IFN] gene signature observed in colon epithelium of patients with active IBD, with a special focus on secreted ubiquitin-like protein ISG15.

Methods: Gene and protein expression in whole mucosa biopsies and in microdissected human colonic epithelial lining, in HT29 human intestinal epithelial cells and primary 3D colonoids treated with PRR-ligands and cytokines, were detected by transcriptomics, in situ hybridisation, immunohistochemistry, western blots, and enzyme-linked immunosorbent assay [ELISA]. Effects of IEC-secreted cytokines were examined in human peripheral blood mononuclear cells [PBMCs] by multiplex chemokine profiling and ELISA.

Results: The type I IFN gene signature in human mucosal biopsies was mimicked in Toll-like receptor TLR3 and to some extent tumour necrosis factor [TNF]-treated human IECs. In intestinal biopsies, ISG15 expression correlated with expression of the newly identified receptor for extracellular ISG15, LFA-1 integrin. ISG15 was expressed and secreted from HT29 cells and primary 3D colonoids through both JAK1-pSTAT-IRF9-dependent and independent pathways. In experiments using PBMCs, we show that ISG15 releases IBD-relevant proinflammatory cytokines such as CXCL1, CXCL5, CXCL8, CCL20, IL1, IL6, TNF, and IFNγ.

Conclusions: ISG15 is secreted from primary IECs upon extracellular stimulation, and mucosal ISG15 emerges as an intriguing candidate for immunotherapy in IBD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/ecco-jcc/jjaa022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7392169PMC
July 2020

Complement component C3 and the TLR co-receptor CD14 are not involved in angiotensin II induced cardiac remodelling.

Biochem Biophys Res Commun 2020 03 16;523(4):867-873. Epub 2020 Jan 16.

Research Institute of Internal Medicine, Oslo University Hospital, Rikshospitalet, Oslo, Norway; Institute of Clinical Medicine, University of Oslo, Oslo, Norway; Center for Heart Failure Research, University of Oslo, Oslo, Norway; K.G. Jebsen Inflammation Research Center, University of Oslo, Oslo, Norway.

Inflammation is centrally involved in the development of cardiac hypertrophy and the processes of remodelling. The complement system and Toll-like receptor (TLR) family, two upstream arms of the innate immune system, have previously been reported to be involved in cardiac remodelling. However, the role of complement component 3 (C3), TLR co-receptor CD14 and the synergy between them have not been addressed during pressure overload-induced cardiac remodelling. Here, we examined angiotensin II-induced cardiac hypertrophy and remodelling for 7 days in male C57Bl/6 J mice deficient in C3, CD14, or both (C3CD14), and WT controls. Angiotensin II infusion induced a mild concentric hypertrophic phenotype in WT mice with increased left ventricle weight, wall thicknesses and reduced ventricular internal diameter, associated with increased cardiac fibrosis. However, there were no differences between WT mice and mice deficient for C3, CD14 or C3CD14, as systolic blood pressure, cardiac function and structure and levels of fibrosis were comparable between WT mice and the three other genotypes. C5a did not change in angiotensin II treated mice, whereas Mac2 levels were increased in angiotensin II treated mice, but did not differ between genotypes. The inflammatory IL-6 response was comparable between WT and C3 deficient mice, however, it was decreased in CD14 and C3CD14 deficient mice. We conclude that deficiency in C3, CD14 or C3CD14 had no effect on cardiac remodelling following angiotensin II-induced pressure overload. This suggests that C3 and CD14 are not involved in angiotensin II-induced adverse cardiac remodelling.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.bbrc.2020.01.018DOI Listing
March 2020

Analysis of cytokines.

Tidsskr Nor Laegeforen 2020 01 6;140(1). Epub 2020 Jan 6.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.4045/tidsskr.18.0961DOI Listing
January 2020

Endotoxin Removal in Septic Shock with the Alteco LPS Adsorber Was Safe But Showed no Benefit Compared to Placebo in the Double-Blind Randomized Controlled Trial-the Asset Study.

Shock 2020 08;54(2):224-231

Department of Surgical Sciences/Anaesthesiology and Intensive Care Medicine, Uppsala University, Uppsala, Sweden.

Purpose: Lipopolysaccharides (LPS) are presumed to contribute to the inflammatory response in sepsis. We investigated if extracorporeal Alteco LPS Adsorber for LPS removal in early gram-negative septic shock was feasible and safe. Also, effects on endotoxin level, inflammatory response, and organ function were assessed.

Methods: A pilot, double-blinded, randomized, Phase IIa, feasibility clinical investigation was undertaken in six Scandinavian intensive care units aiming to allocate 32 septic shock patients with abdominal or urogenital focus on LPS Adsorber therapy or a Sham Adsorber, therapy without active LPS binding. The study treatment was initiated within 12 h of inclusion and given for 6 h daily on first 2 days. LPS was measured in all patients.

Results: The investigation was terminated after 527 days with eight patients included in the LPS Adsorber group and seven in the Sham group. Twenty-one adverse effects, judged not to be related to the device, were reported in three patients in the LPS Adsorber group and two in the Sham group. Two patients in the Sham group and no patients in the LPS Adsorber group died within 28 days. Plasma LPS levels were low without groups differences during or after adsorber therapy. The changes in inflammatory markers and organ function were similar in the groups.

Conclusions: In a small cohort of patients with presumed gram-negative septic shock, levels of circulating endotoxin were low and no adverse effects within 28 days after LPS adsorber-treatment were observed. No benefit compared with a sham device was seen when using a LPS adsorber in addition to standard care.

Trial Registration: Clinicaltrials.gov NCT02335723. Registered: November 28, 2014.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/SHK.0000000000001503DOI Listing
August 2020

Selective and marked decrease of complement receptor C5aR2 in human thoracic aortic aneurysms: a dysregulation with potential inflammatory effects.

Open Heart 2019;6(2):e001098. Epub 2019 Nov 10.

Institute of Clinical Medicine, University of Oslo Faculty of Medicine, Oslo, Norway.

Objective: The aetiology of thoracic aortic aneurysm (TAA) is largely unknown, but inflammation is likely to play a central role in the pathogenesis. In this present study, we aim to investigate the complement receptors in TAA.

Methods: Aortic tissue and blood from 31 patients with non-syndromic TAA undergoing thoracic aortic repair surgery were collected. Aortic tissue and blood from 36 patients with atherosclerosis undergoing coronary artery bypass surgery or aortic valve replacement were collected and served as control material. The expression of the complement anaphylatoxin receptors C3aR1, C5aR1 and C5aR2 in aortic tissue were examined by quantitative RT-PCR and C5aR2 protein by immunohistochemistry. Colocalisation of C5aR2 to different cell types was analysed by immunofluorescence. Complement activation products C3bc and sC5b-9 were measured in plasma.

Results: Compared with controls, TAA patients had substantial (73%) downregulated gene expression of C5aR2 as seen both at the mRNA (p=0.005) level and protein (p=0.03) level. In contrast, there were no differences in the expression of C3aR1 and C5aR1 between the two groups. Immunofluorescence examination showed that C5aR2 was colocalised to macrophages and T cells in the aortic media. There were no differences in the degree of systemic complement activation between the two groups.

Conclusion: Our findings suggest downregulation of the C5aR2, regarded to act mainly anti-inflammatory, in electively operated TAA as compared with non-aneurysmatic aortas of patients with aortic stenosis and/or coronary artery disease. This may tip the balance towards a relative increase in the inflammatory responses induced by C5aR1 and thus enhance the inflammatory processes in TAA.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1136/openhrt-2019-001098DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6861114PMC
February 2021

NHDL, a recombinant V/V hybrid antibody control for IgG2/4 antibodies.

MAbs 2020 Jan-Dec;12(1):1686319

Research Laboratory, Nordland Hospital Trust, Bodø, Norway.

The mechanism of action of recombinant IgG2/4 antibodies involves blocking of their target without the induction of effector functions. Examples are eculizumab (Soliris®), which is used clinically to block complement factor C5, as well as anti-human CD14 (r18D11) and anti-porcine CD14 (rMIL2) produced in our laboratory. So far, no proper IgG2/4 control antibody has been available for controlled validation of IgG2/4 antibody functions. Here, we describe the design of a recombinant control antibody (NHDL), which was generated by combining the variable light (V) and heavy (V) chains from two unrelated specificities. NHDL was readily expressed and purified as a stable IgG2/4 antibody, and showed no detectable specificity toward any putative antigen present in human or porcine blood. The approach of artificial V/V combination may be adopted for the design of other recombinant control antibodies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1080/19420862.2019.1686319DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6927768PMC
January 2021
-->