Publications by authors named "Sofie Struyf"

128 Publications

The Role of Post-Translational Modifications of Chemokines by CD26 in Cancer.

Cancers (Basel) 2021 Aug 24;13(17). Epub 2021 Aug 24.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, KU Leuven, B-3000 Leuven, Belgium.

Chemokines are a large family of small chemotactic cytokines that fulfill a central function in cancer. Both tumor-promoting and -impeding roles have been ascribed to chemokines, which they exert in a direct or indirect manner. An important post-translational modification that regulates chemokine activity is the NH-terminal truncation by peptidases. CD26 is a dipeptidyl peptidase (DPPIV), which typically clips a NH-terminal dipeptide from the chemokine. With a certain degree of selectivity in terms of chemokine substrate, CD26 only recognizes chemokines with a penultimate proline or alanine. Chemokines can be protected against CD26 recognition by specific amino acid residues within the chemokine structure, by oligomerization or by binding to cellular glycosaminoglycans (GAGs). Upon truncation, the binding affinity for receptors and GAGs is altered, which influences chemokine function. The consequences of CD26-mediated clipping vary, as unchanged, enhanced, and reduced activities are reported. In tumors, CD26 most likely has the most profound effect on CXCL12 and the interferon (IFN)-inducible CXCR3 ligands, which are converted into receptor antagonists upon truncation. Depending on the tumor type, expression of CD26 is upregulated or downregulated and often results in the preferential generation of the chemokine isoform most favorable for tumor progression. Considering the tight relationship between chemokine sequence and chemokine binding specificity, molecules with the appropriate characteristics can be chemically engineered to provide innovative therapeutic strategies in a cancer setting.
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http://dx.doi.org/10.3390/cancers13174247DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8428238PMC
August 2021

The Antimicrobial Activity of Peripheral Blood Neutrophils Is Altered in Patients with Primary Ciliary Dyskinesia.

Int J Mol Sci 2021 Jun 8;22(12). Epub 2021 Jun 8.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, 3000 Leuven, Belgium.

The airways of patients with primary ciliary dyskinesia (PCD) contain persistently elevated neutrophil numbers and CXCL8 levels. Despite their abundance, neutrophils fail to clear the airways from bacterial infections. We investigated whether neutrophil functions are altered in patients with PCD. Neutrophils from patients and healthy controls (HC) were isolated from peripheral blood and exposed to various bacterial stimuli or cytokines. Neutrophils from patients with PCD were less responsive to low levels of fMLF in three different chemotaxis assays ( < 0.05), but expression of the fMLF receptors was unaltered. PCD neutrophils showed normal phagocytic function and expression of adhesion molecules. However, PCD neutrophils produced less reactive oxygen species upon stimulation with bacterial products or cytokines compared to HC neutrophils ( < 0.05). Finally, the capacity to release DNA, as observed during neutrophil extracellular trap formation, seemed to be reduced in patients with PCD compared to HC ( = 0.066). These results suggest that peripheral blood neutrophils from patients with PCD, in contrast to those of patients with cystic fibrosis or COPD, do not show features of over-activation, neither on baseline nor after stimulation. If these findings extend to lung-resident neutrophils, the reduced neutrophil activity could possibly contribute to the recurrent respiratory infections in patients with PCD.
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http://dx.doi.org/10.3390/ijms22126172DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8230338PMC
June 2021

Kinetics of peripheral blood neutrophils in severe coronavirus disease 2019.

Clin Transl Immunology 2021 29;10(4):e1271. Epub 2021 Apr 29.

Laboratory of Molecular Immunology Department of Microbiology, Immunology and Transplantation Rega Institute, KU Leuven Leuven Belgium.

Objectives: Emerging evidence of dysregulation of the myeloid cell compartment urges investigations on neutrophil characteristics in coronavirus disease 2019 (COVID-19). We isolated neutrophils from the blood of COVID-19 patients receiving general ward care and from patients hospitalised at intensive care units (ICUs) to explore the kinetics of circulating neutrophils and factors important for neutrophil migration and activation.

Methods: Multicolour flow cytometry was exploited for the analysis of neutrophil differentiation and activation markers. Multiplex and ELISA technologies were used for the quantification of protease, protease inhibitor, chemokine and cytokine concentrations in plasma. Neutrophil polarisation responses were evaluated microscopically. Gelatinolytic and metalloproteinase activity in plasma was determined using a fluorogenic substrate. Co-culturing healthy donor neutrophils with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) allowed us to investigate viral replication in neutrophils.

Results: Upon ICU admission, patients displayed high plasma concentrations of granulocyte-colony-stimulating factor (G-CSF) and the chemokine CXCL8, accompanied by emergency myelopoiesis as illustrated by high levels of circulating CD10, immature neutrophils with reduced CXCR2 and C5aR expression. Neutrophil elastase and non-metalloproteinase-derived gelatinolytic activity were increased in plasma from ICU patients. Significantly higher levels of circulating tissue inhibitor of metalloproteinase 1 (TIMP-1) in patients at ICU admission yielded decreased total MMP proteolytic activity in blood. COVID-19 neutrophils were hyper-responsive to CXCL8 and CXCL12 in shape change assays. Finally, SARS-CoV-2 failed to replicate inside human neutrophils.

Conclusion: Our study provides detailed insights into the kinetics of neutrophil phenotype and function in severe COVID-19 patients, and supports the concept of an increased neutrophil activation state in the circulation.
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http://dx.doi.org/10.1002/cti2.1271DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8082714PMC
April 2021

From ELISA to Immunosorbent Tandem Mass Spectrometry Proteoform Analysis: The Example of CXCL8/Interleukin-8.

Front Immunol 2021 11;12:644725. Epub 2021 Mar 11.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute, Katholieke Universiteit Leuven, Leuven, Belgium.

With ELISAs one detects the ensemble of immunoreactive molecules in biological samples. For biomolecules undergoing proteolysis for activation, potentiation or inhibition, other techniques are necessary to study biology. Here we develop methodology that combines immunosorbent sample preparation and nano-scale liquid chromatography-tandem mass spectrometry (nano-LC-MS/MS) for proteoform analysis (ISTAMPA) and apply this to the aglycosyl chemokine CXCL8. CXCL8, the most powerful human chemokine with neutrophil chemotactic and -activating properties, occurs in different NH-terminal proteoforms due to its susceptibility to site-specific proteolytic modification. Specific proteoforms display up to 30-fold enhanced activity. The immunosorbent ion trap top-down mass spectrometry-based approach for proteoform analysis allows for simultaneous detection and quantification of full-length CXCL8(1-77), elongated CXCL8(-2-77) and all naturally occurring truncated CXCL8 forms in biological samples. For the first time we demonstrate site-specific proteolytic activation of CXCL8 in synovial fluids from patients with chronic joint inflammation and address the importance of sample collection and processing.
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http://dx.doi.org/10.3389/fimmu.2021.644725DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7991300PMC
March 2021

Phenotypical and Functional Characterization of Neutrophils in Two Pyrin-Associated Auto-inflammatory Diseases.

J Clin Immunol 2021 Jul 5;41(5):1072-1084. Epub 2021 Mar 5.

Laboratory of Immunobiology, Rega Institute, KU Leuven, Leuven, Belgium.

Purpose: Familial Mediterranean Fever (FMF) and Pyrin-Associated Autoinflammation with Neutrophilic Dermatosis (PAAND) are clinically distinct autoinflammatory disorders caused by mutations in the pyrin-encoding gene MEFV. We investigated the transcriptional, phenotypical, and functional characteristics of patient neutrophils to explore their potential role in FMF and PAAND pathophysiology.

Methods: RNA sequencing was performed to discover transcriptional aberrancies. The phenotypical features, degranulation properties, and phagocytic capacity of neutrophils were assessed by flow cytometry. Production of reactive oxygen species (ROS), myeloperoxidase (MPO) release, and chemotactic responses were investigated via chemiluminescence, ELISA, and Boyden chamber assays, respectively.

Results: Neutrophils from PAAND and FMF patients showed a partially overlapping, activated gene expression profile with increased expression of S100A8, S100A9, S100A12, IL-4R, CD48, F5, MMP9, and NFKB. Increased MMP9 and S100A8/A9 expression levels were accompanied by high plasma concentrations of the encoded proteins. Phenotypical analysis revealed that neutrophils from FMF patients exhibited an immature character with downregulation of chemoattractant receptors CXCR2, C5aR, and BLTR1 and increased expression of Toll-like receptor 4 (TLR4) and TLR9. PAAND neutrophils displayed an increased random, but reduced CXCL8-induced migration. A tendency for enhanced random migration was observed for FMF neutrophils. PAAND neutrophils showed a moderately but significantly enhanced phagocytic activity as opposed to neutrophils from FMF patients. Neutrophils from both patient groups showed increased MPO release and ROS production.

Conclusions: Neutrophils from patients with FMF and PAAND, carrying different mutations in the MEFV gene, share a pro-inflammatory phenotype yet demonstrate diverse features, underscoring the distinction between both diseases.
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http://dx.doi.org/10.1007/s10875-021-01008-4DOI Listing
July 2021

Evaluation of Proteoforms of the Transmembrane Chemokines CXCL16 and CX3CL1, Their Receptors, and Their Processing Metalloproteinases ADAM10 and ADAM17 in Proliferative Diabetic Retinopathy.

Front Immunol 2020 20;11:601639. Epub 2021 Jan 20.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, Leuven, Belgium.

The transmembrane chemokine pathways CXCL16/CXCR6 and CX3CL1/CX3CR1 are strongly implicated in inflammation and angiogenesis. We investigated the involvement of these chemokine pathways and their processing metalloproteinases ADAM10 and ADAM17 in the pathophysiology of proliferative diabetic retinopathy (PDR). Vitreous samples from 32 PDR and 24 non-diabetic patients, epiretinal membranes from 18 patients with PDR, rat retinas, human retinal Müller glial cells and human retinal microvascular endothelial cells (HRMECs) were studied by enzyme-linked immunosorbent assay, immunohistochemistry and Western blot analysis. angiogenesis assays were performed and the adherence of leukocytes to CXCL16-stimulated HRMECs was assessed. CXCL16, CX3CL1, ADAM10, ADAM17 and vascular endothelial growth factor (VEGF) levels were significantly increased in vitreous samples from PDR patients. The levels of CXCL16 were 417-fold higher than those of CX3CL1 in PDR vitreous samples. Significant positive correlations were found between the levels of VEGF and the levels of CXCL16, CX3CL1, ADAM10 and ADAM17. Significant positive correlations were detected between the numbers of blood vessels expressing CD31, reflecting the angiogenic activity of PDR epiretinal membranes, and the numbers of blood vessels and stromal cells expressing CXCL16, CXCR6, ADAM10 and ADAM17. CXCL16 induced upregulation of phospho-ERK1/2, p65 subunit of NF-κB and VEGF in cultured Müller cells and tumor necrosis factor-α induced upregulation of soluble CXCL16 and ADAM17 in Müller cells. Treatment of HRMECs with CXCL16 resulted in increased expression of intercellular adhesion molecule-1 (ICAM-1) and increased leukocyte adhesion to HRMECs. CXCL16 induced HRMEC proliferation, formation of sprouts from HRMEC spheroids and phosphorylation of ERK1/2. Intravitreal administration of CXCL16 in normal rats induced significant upregulation of the p65 subunit of NF-κB, VEGF and ICAM-1 in the retina. Our findings suggest that the chemokine axis CXCL16/CXCR6 and the processing metalloproteinases ADAM10 and ADAM17 might serve a role in the initiation and progression of PDR.
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http://dx.doi.org/10.3389/fimmu.2020.601639DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7854927PMC
June 2021

Studying Neutrophil Function in vitro: Cell Models and Environmental Factors.

J Inflamm Res 2021 20;14:141-162. Epub 2021 Jan 20.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, Leuven 3000, Belgium.

Neutrophils are the most abundant immune cell type in the blood and constitute the first line of defense against invading pathogens. Despite their important role in many diseases, they are challenging to study due to their short life span and the inability to cryopreserve or expand them in vitro. Thus, research into neutrophils has to rely on cells freshly isolated from peripheral blood of human donors, introducing donor-dependent variation in the experimental data. To counteract these problems, researchers tried to develop adequate cell models, such as cell lines. For those functional studies that cannot rely on cell models, a standardization of protocols regarding neutrophil purification and culturing could be a solution. In this review, we provide an overview of the most commonly used models for neutrophil function (HL-60, PLB-985, NB4, Kasumi-1 and induced pluripotent stem cells). In addition, we describe the effects of glucose concentration, pH, oxygen tension and temperature on neutrophil function.
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http://dx.doi.org/10.2147/JIR.S284941DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7829132PMC
January 2021

The turning away of serum amyloid A biological activities and receptor usage.

Immunology 2021 Jun 4;163(2):115-127. Epub 2021 Jan 4.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, Leuven, Belgium.

Serum amyloid A (SAA) is an acute-phase protein (APP) to which multiple immunological functions have been attributed. Regardless, the true biological role of SAA remains poorly understood. SAA is remarkably conserved in mammalian evolution, thereby suggesting an important biological function. Since its discovery in the 1970s, the majority of researchers have investigated SAA using recombinant forms made available through bacterial expression. Nevertheless, recent studies indicate that these recombinant forms of SAA are unreliable. Indeed, commercial SAA variants have been shown to be contaminated with bacterial products including lipopolysaccharides and lipoproteins. As such, biological activities and receptor usage (TLR2, TLR4) revealed through the use of commercial SAA variants may not reflect the inherent nature of this APP. Within this review, we discuss the biological effects of SAA that have been demonstrated through more solid experimental approaches. SAA takes part in the innate immune response via the recruitment of leucocytes and executes, through pathogen recognition, antimicrobial activity. Knockout animal models implicate SAA in a range of functions, such as regulation of T-cell-mediated responses and monopoiesis. Moreover, through its structural motifs, not only does SAA function as an extracellular matrix protein, but it also binds extracellular matrix proteins. Finally, we here also provide an overview of definite SAA receptor-mediated functions and highlight those that are yet to be validated. The role of FPR2 in SAA-mediated leucocyte recruitment has been confirmed; nevertheless, SAA has been linked to a range of other receptors including CD36, SR-BI/II, RAGE and P2RX7.
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http://dx.doi.org/10.1111/imm.13295DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8114209PMC
June 2021

Citrullination as a novel posttranslational modification of matrix metalloproteinases.

Matrix Biol 2021 01 4;95:68-83. Epub 2020 Nov 4.

Rega Institute for Medical Research, Laboratory of Immunobiology, Department of Microbiology, Immunology and Transplantation, KU Leuven, Herestraat 49 box 1044, Leuven 3000, Belgium. Electronic address:

Matrix metalloproteinases (MMPs) are enzymes with critical roles in biology and pathology. Glycosylation, nitrosylation and proteolysis are known posttranslational modifications (PTMs) regulating intrinsically the activities of MMPs. We discovered MMP citrullination by peptidyl arginine deiminases (PADs) as a new PTM. Upon hypercitrullination, MMP-9 acquired a higher affinity for gelatin than control MMP-9. Furthermore, hypercitrullinated proMMP-9 was more efficiently activated by MMP-3 compared to control MMP-9. JNJ0966, a specific therapeutic inhibitor of MMP-9 activation, inhibited the activation of hypercitrullinated proMMP-9 by MMP-3 significantly less in comparison with control proMMP-9. The presence of citrullinated/homocitrullinated MMP-9 was detected in vivo in neutrophil-rich sputum samples of cystic fibrosis patients. In addition to citrullination of MMP-9, we report efficient citrullination of MMP-1 and lower citrullination levels of MMP-3 and MMP-13 by PAD2 in vitro. In conclusion, citrullination of MMPs is a new PTM worthy of additional biochemical and biological studies.
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http://dx.doi.org/10.1016/j.matbio.2020.10.005DOI Listing
January 2021

CXCL14 Preferentially Synergizes With Homeostatic Chemokine Receptor Systems.

Front Immunol 2020 5;11:561404. Epub 2020 Oct 5.

Division of Infection and Immunity, Cardiff University School of Medicine, Cardiff, United Kingdom.

Reflecting their importance in immunity, the activity of chemokines is regulated on several levels, including tissue and context-specific expression and availability of their cognate receptor on target cells. Chemokine synergism, affecting both chemokine and chemokine receptor function, has emerged as an additional control mechanism. We previously demonstrated that CXCL14 is a positive allosteric modulator of CXCR4 in its ability to synergize with CXCL12 in diverse cellular responses. Here, we have extended our study to additional homeostatic, as well as a selection of inflammatory chemokine systems. We report that CXCL14 strongly synergizes with low (sub-active) concentrations of CXCL13 and CCL19/CCL21 in chemotaxis with immune cells expressing the corresponding receptors CXCR5 and CCR7, respectively. CXCL14 by itself was inactive, not only on cells expressing CXCR5 or CCR7 but also on cells expressing any other known conventional or atypical chemokine receptor, as assessed by chemotaxis and/or β-arrestin recruitment assays. Furthermore, synergistic migration responses between CXCL14 and inflammatory chemokines CXCL10/CXCL11 and CCL5, targeting CXCR3 and CCR5, respectively, were marginal and occasional synergistic Ca flux responses were observed. CXCL14 bound to 300-19 cells and interfered with CCL19 binding to CCR7-expressing cells, suggesting that these cellular interactions contributed to the reported CXCL14-mediated synergistic activities. We propose a model whereby tissue-expressed CXCL14 contributes to cell localization under steady-state conditions at sites with prominent expression of homeostatic chemokines.
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http://dx.doi.org/10.3389/fimmu.2020.561404DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7570948PMC
May 2021

Efficacy of B Cell Depletion Therapy with Rituximab in Refractory Chronic Recurrent Uveitis Associated with Vogt-Koyanagi-Harada Disease.

Ocul Immunol Inflamm 2020 Sep 29:1-8. Epub 2020 Sep 29.

Rega Institute for Medical Research, Department of Microbiology and Immunology, University of Leuven, KU Leuven, Leuven, Belgium.

Purpose: To evaluate the efficacy of B cell depletion therapy with the chimeric mouse/human anti-CD20 monoclonal antibody rituximab for refractory chronic recurrent granulomatous uveitis associated with Vogt-Koyanagi-Harada (VKH) disease.

Methods: Retrospective study of 9 patients (18 eyes) who failed to respond to conventional combination immunosuppressive therapy.

Results: All the patients received 3 rituximab infusions. The follow-up period after initiation of rituximab therapy ranged from 9 to 36 months (mean ±SD, 19.2 ± 10.1). All patients achieved remission and visual acuity significantly improved ( < .001). Rituximab provided corticosteroid-sparing effect along with control of inflammation. No rituximab-related complications were observed.

Conclusions: Rituximab is effective for the treatment of refractory chronic recurrent granulomatous uveitis associated with VKH disease.
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http://dx.doi.org/10.1080/09273948.2020.1820531DOI Listing
September 2020

Biological Characterization of Commercial Recombinantly Expressed Immunomodulating Proteins Contaminated with Bacterial Products in the Year 2020: The SAA3 Case.

Mediators Inflamm 2020 6;2020:6087109. Epub 2020 Jul 6.

KU Leuven, Department of Microbiology and Immunology, Rega Institute, Laboratory of Molecular Immunology, Herestraat 49, Box 1042, 3000 Leuven, Belgium.

The serum amyloid A (SAA) gene family is highly conserved and encodes acute phase proteins that are upregulated in response to inflammatory triggers. Over the years, a considerable amount of literature has been published attributing a wide range of biological effects to SAAs such as leukocyte recruitment, cytokine and chemokine expression and induction of matrix metalloproteinases. Furthermore, SAAs have also been linked to protumorigenic, proatherogenic and anti-inflammatory effects. Here, we investigated the biological effects conveyed by murine SAA3 (mu rSAA3) recombinantly expressed in . We observed the upregulation of a number of chemokines including CCL2, CCL3, CXCL1, CXCL2, CXCL6 or CXCL8 following stimulation of monocytic, fibroblastoid and peritoneal cells with mu rSAA3. Furthermore, this SAA variant displayed potent recruitment of neutrophils through the activation of TLR4. However, a major problem associated with proteins derived from recombinant expression in bacteria is potential contamination with various bacterial products, such as lipopolysaccharide, lipoproteins and formylated peptides. This is of particular relevance in the case of SAA as there currently exists a discrepancy in biological activity between SAA derived from recombinant expression and that of an endogenous source, i.e. inflammatory plasma. Therefore, we subjected commercial recombinant mu rSAA3 to purification to homogeneity via reversed-phase high-performance liquid chromatography (RP-HPLC) and re-assessed its biological potential. RP-HPLC-purified mu rSAA3 did not induce chemokines and lacked neutrophil chemotactic activity, but retained the capacity to synergize with CXCL8 in the activation of neutrophils. In conclusion, experimental results obtained when using proteins recombinantly expressed in bacteria should always be interpreted with care.
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http://dx.doi.org/10.1155/2020/6087109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7362292PMC
July 2021

Neutrophils: Underestimated Players in the Pathogenesis of Multiple Sclerosis (MS).

Int J Mol Sci 2020 Jun 26;21(12). Epub 2020 Jun 26.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, KU Leuven, Herestraat 49-Box 1042, 3000 Leuven, Belgium.

Neutrophils are the most abundant circulating and first-responding innate myeloid cells and have so far been underestimated in the context of multiple sclerosis (MS). MS is the most frequent, immune-mediated, inflammatory disease of the central nervous system. MS is treatable but not curable and its cause(s) and pathogenesis remain elusive. The involvement of neutrophils in MS pathogenesis has been suggested by the use of preclinical animal disease models, as well as on the basis of patient sample analysis. In this review, we provide an overview of the possible mechanisms and functions by which neutrophils may contribute to the development and pathology of MS. Neutrophils display a broad variety of effector functions enabling disease pathogenesis, including (1) the release of inflammatory mediators and enzymes, such as interleukin-1β, myeloperoxidase and various proteinases, (2) destruction and phagocytosis of myelin (as debris), (3) release of neutrophil extracellular traps, (4) production of reactive oxygen species, (5) breakdown of the blood-brain barrier and (6) generation and presentation of autoantigens. An important question relates to the issue of whether neutrophils exhibit a predominantly proinflammatory function or are also implicated in the resolution of chronic inflammatory responses in MS.
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http://dx.doi.org/10.3390/ijms21124558DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7349048PMC
June 2020

Serum Amyloid A1 (SAA1) Revisited: Restricted Leukocyte-Activating Properties of Homogeneous SAA1.

Front Immunol 2020 14;11:843. Epub 2020 May 14.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, KU Leuven, Leuven, Belgium.

Infection, sterile injury, and chronic inflammation trigger the acute phase response in order to re-establish homeostasis. This response includes production of positive acute phase proteins in the liver, such as members of the serum amyloid A (SAA) family. In humans the major acute phase SAAs comprise a group of closely related variants of SAA1 and SAA2. SAA1 was proven to be chemotactic for several leukocyte subtypes through activation of the G protein-coupled receptor FPRL1/FPR2. Several other biological activities of SAA1, such as cytokine induction, reported to be mediated via TLRs, have been debated recently. Especially commercial SAA1, recombinantly produced in , was found to be contaminated with bacterial products confounding biological assays performed with this rSAA1. We purified rSAA1 by RP-HPLC to homogeneity, removing contaminants such as lipopolysaccharides, lipoproteins and formylated peptides, and re-assessed several biological activities attributed to SAA1 (chemotaxis, cytokine induction, MMP-9 release, ROS generation, and macrophage differentiation). The homogeneous rSAA1 (hrSAA1) lacked most cell-activating properties, but its leukocyte-recruiting capacity and it's synergy with other leukocyte attractants remained preserved. Furthermore, hrSAA1 maintained the ability to promote monocyte survival. This indicates that pure hrSAA1 retains its potential to activate FPR2, whereas TLR-mediated effects seem to be related to traces of bacterial TLR ligands in the -produced human rSAA1.
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http://dx.doi.org/10.3389/fimmu.2020.00843DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7240019PMC
March 2021

Induction of Chemokines by Hepatitis C Virus Proteins: Synergy of the Core Protein with Interleukin-1β and Interferon-γ in Liver Bystander Cells.

J Interferon Cytokine Res 2020 04 7;40(4):195-206. Epub 2020 Feb 7.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, Leuven, Belgium.

Chronic hepatitis C virus (HCV) infection accounts for a large proportion of hepatic fibrosis and carcinoma cases observed worldwide. Mechanisms involved in HCV-induced hepatic injury have yet to be fully elucidated. Of particular interest is the capacity of HCV to regulate inflammatory responses. Here, we reveal modulation of cytokine activity by the HCV proteins non-structural protein 3 (NS3), glycoprotein E2, and core protein for their ability to induce chemokine expression in various liver bystander cells. Chemokines sustain chronic liver inflammation and relay multiple fibrogenic effects. CCL2, CCL3, CCL20, CXCL8, and CXCL10 were differentially expressed after treatment of monocytes, fibroblasts, or liver sinusoidal microvascular endothelial cells (LSECs) with HCV proteins. In comparison to NS3 and glycoprotein E2, core protein was a stronger inducer of chemokines in liver bystander cells. Interferon-γ (IFN-γ) and interleukin-1β (IL-1β) synergized with core protein to induce CCL2, CCL20, CXCL8, or CXCL10 in fibroblasts or LSECs. These findings reveal new mechanisms of hepatic injury caused by HCV.
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http://dx.doi.org/10.1089/jir.2019.0115DOI Listing
April 2020

The Proinflammatory and Proangiogenic Macrophage Migration Inhibitory Factor Is a Potential Regulator in Proliferative Diabetic Retinopathy.

Front Immunol 2019 4;10:2752. Epub 2019 Dec 4.

Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven, Leuven, Belgium.

The macrophage migration inhibitory factor (MIF)/CD74 signaling pathway is strongly implicated in inflammation and angiogenesis. We investigated the expression of MIF and its receptor CD74 in proliferative diabetic retinopathy (PDR) to reveal a possible role of this pathway in the pathogenesis of PDR. Levels of MIF, soluble (s)CD74, soluble intercellular adhesion molecule-1 (sICAM-1) and vascular endothelial growth factor (VEGF) were significantly increased in the vitreous from patients with PDR compared to nondiabetic control samples. We detected significant positive correlations between the levels of MIF and the levels of sICAM-1 ( = 0.43; = 0.001) and VEGF ( = 0.7; < 0.001). Through immunohistochemical analysis of PDR epiretinal membranes, significant positive correlations were also found between microvessel density (CD31 expression) and the numbers of blood vessels expressing MIF ( = 0.56; = 0.045) and stromal cells expressing MIF ( = 0.79; = 0.001) and CD74 ( = 0.59; = 0.045). Similar to VEGF, MIF was induced in Müller cells cultured under hypoxic conditions and MIF induced phosphorylation of ERK1/2 and VEGF production in Müller cells. Intravitreal administration of MIF in normal rats induced increased retinal vascular permeability and significant upregulation of phospho-ERK1/2, NF-κB, ICAM-1 and vascular cell adhesion molecule-1 expression in the retina. MIF induced migration and proliferation of human retinal microvascular endothelial cells. These results suggest that MIF/CD74 signaling is involved in PDR angiogenesis.
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http://dx.doi.org/10.3389/fimmu.2019.02752DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6904364PMC
October 2020

Soluble cytokine receptor levels in aqueous humour of patients with specific autoimmune uveitic entities: sCD30 is a biomarker of granulomatous uveitis.

Eye (Lond) 2020 09 5;34(9):1614-1623. Epub 2019 Dec 5.

Department of Microbiology and Immunology, Rega Institute for Medical Research, University of Leuven, KU Leuven, Leuven, Belgium.

Purpose: Soluble cytokine receptors are potential biomarkers for immune activation and have a promising potential as immunotherapeutic agents. We investigated the levels of soluble cytokine receptors in aqueous humour (AH) samples from patients with specific autoimmune uveitic entities.

Methods: Patients with active uveitis associated with Behçet's disease (BD) (n = 13), sarcoidosis (n = 8), HLA-B27-related inflammation (n = 12), Vogt-Koyanagi-Harada (VKH) disease (n = 12) and control subjects (n = 9) were included. AH samples were analyzed with the use of multiplex assays for the proinflammatory cytokine tumour necrosis factor (TNF)-α and the soluble cytokine receptors sCD30, sCD163, sgp130, sIL-6 receptor-α (sIL-6R), sTNFRI and sTNFRII.

Results: TNF-α and soluble cytokine receptor AH levels were significantly higher in uveitis patients (n = 45) compared with controls (n = 9). When nongranulomatous uveitis (BD and HLA-B27-associated uveitis) was compared with granulomatous uveitis (sarcoidosis and VKH disease), the levels of sCD30 and sTNFRI/TNF-α and sTNFRII/TNF-α ratios were significantly enhanced in granulomatous uveitis. Finally, when comparing the profile in the specific uveitis entities, sCD30 levels were highest in patients with VKH disease. sgp130, sCD163, sIL-6R, sTNFRI and sTNFRII did not differ significantly between the four different clinical uveitic subgroups.

Conclusions: Soluble cytokine receptors are significantly upregulated in autoimmune uveitis. CD30 T cells might contribute to the inflammatory process in granulomatous uveitis, particularly in VKH disease. Granulomatous uveitis is also characterized by significantly higher sTNFRs/TNF-α ratios than nongranulomatous uveitis.
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http://dx.doi.org/10.1038/s41433-019-0693-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7608430PMC
September 2020

Cytokines and serum amyloid A in the pathogenesis of hepatitis C virus infection.

Cytokine Growth Factor Rev 2019 12 28;50:29-42. Epub 2019 Oct 28.

Laboratory of Molecular Immunology, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, University of Leuven, Leuven, 3000, Belgium. Electronic address:

Expression of the acute phase protein serum amyloid A (SAA) is dependent on the release of the pro-inflammatory cytokines IL-1, IL-6 and TNF-α during infection and inflammation. Hepatitis C virus (HCV) upregulates SAA-inducing cytokines. In line with this, a segment of chronically infected individuals display increased circulating levels of SAA. SAA has even been proposed to be a potential biomarker to evaluate treatment efficiency and the course of disease. SAA possesses antiviral activity against HCV via direct interaction with the viral particle, but might also divert infectivity through its function as an apolipoprotein. On the other hand, SAA shares inflammatory and angiogenic activity with chemotactic cytokines by activating the G protein-coupled receptor, formyl peptide receptor 2. These latter properties might promote chronic inflammation and hepatic injury. Indeed, up to 80 % of infected individuals develop chronic disease because they cannot completely clear the infection, due to diversion of the immune response. In this review, we summarize the interconnection between SAA and cytokines in the context of HCV infection and highlight the dual role SAA could play in this disease. Nevertheless, more research is needed to establish whether the balance between those opposing activities can be tilted in favor of the host defense.
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http://dx.doi.org/10.1016/j.cytogfr.2019.10.006DOI Listing
December 2019

Local Cytokine Expression Profiling in Patients with Specific Autoimmune Uveitic Entities.

Ocul Immunol Inflamm 2020 Apr 4;28(3):453-462. Epub 2019 Jun 4.

Laboratory of Immunobiology, Rega Institute for Medical Research and Department of Microbiology and Immunology, University of Leuven, KU Leuven, Leuven, Belgium.

: To evaluate expression of cytokines GM-CSF, IL-11, IL-12p40, IL-12p70, IL-27p28, IL-35, APRIL, BAFF, TWEAK, and LIGHT in uveitis.: Aqueous humor samples from patients with active uveitis associated with Behçet's disease (BD), sarcoidosis, HLA-B27-related inflammation, and Vogt-Koyanagi-Harada (VKH) disease and control patients were assayed with a multiplex assay.: Comparing all patients to controls, GM-CSF, IL-11, IL-12p40, APRIL, and BAFF were significantly increased, whereas LIGHT was significantly decreased. IL-11 and BAFF were the most strongly upregulated, being elevated 19.7-fold and 14.1-fold, respectively, compared with controls. IL-11 was significantly highest in HLA-B27 uveitis. GM-CSF, IL-11, and IL-12p40 were significantly higher in nongranulomatous uveitis (BD and HLA-B27) than in granulomatous uveitis (sarcoidosis and VKH), whereas APRIL and TWEAK were significantly higher in granulomatous uveitis.: IL-11-driven immune responses might be more potent in nongranulomatous uveitis, particularly in HLA-B27 uveitis. BAFF and APRIL might contribute to B cell-driven autoimmune response in uveitis.
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http://dx.doi.org/10.1080/09273948.2019.1604974DOI Listing
April 2020

Human DOCK2 Deficiency: Report of a Novel Mutation and Evidence for Neutrophil Dysfunction.

J Clin Immunol 2019 04 5;39(3):298-308. Epub 2019 Mar 5.

Laboratory for Inborn Errors of Immunity, Department of Immunology and Microbiology, KU Leuven, Leuven, EU, Belgium.

DOCK2 is a guanine-nucleotide-exchange factor for Rac proteins. Activated Rac serves various cellular functions including the reorganization of the actin cytoskeleton in lymphocytes and neutrophils and production of reactive oxygen species in neutrophils. Since 2015, six unrelated patients with combined immunodeficiency and early-onset severe viral infections caused by bi-allelic loss-of-function mutations in DOCK2 have been described. Until now, the function of phagocytes, specifically neutrophils, has not been assessed in human DOCK2 deficiency. Here, we describe a new kindred with four affected siblings harboring a homozygous splice-site mutation (c.2704-2 A > C) in DOCK2. The mutation results in alternative splicing and a complete loss of DOCK2 protein expression. The patients presented with leaky severe combined immunodeficiency or Omenn syndrome. The novel mutation affects EBV-B cell migration and results in NK cell dysfunction similar to previous observations. Moreover, both cytoskeletal rearrangement and reactive oxygen species production are partially impaired in DOCK2-deficient neutrophils.
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http://dx.doi.org/10.1007/s10875-019-00603-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6647034PMC
April 2019

Expression of interleukin (IL)-10 family cytokines in aqueous humour of patients with specific endogenous uveitic entities: elevated levels of IL-19 in human leucocyte antigen-B27-associated uveitis.

Acta Ophthalmol 2019 Aug 13;97(5):e780-e784. Epub 2019 Feb 13.

Rega Institute for Medical Research, Department of Microbiology and Immunology, University of Leuven, KU Leuven, Leuven, Belgium.

Purpose: Evidence exists that the interleukin (IL)-10 family of cytokines is involved in autoimmune diseases. The aim of this study was to analyse the levels of the IL-10 family cytokines IL-10, IL-19, IL-20, IL-22, IL-26, IL-28A and IL-29 in aqueous humour (AH) samples from patients with specific uveitic entities. In addition, we correlated their levels with the levels of the proinflammatory cytokines tumour necrosis factor-α (TNF-α) and IL-1β.

Methods: Aqueous humour (AH) samples from patients with active uveitis associated with Behçet's disease (BD; n = 13), sarcoidosis (n = 8), human leucocyte antigen (HLA)-B27-related inflammation (n = 12), Vogt-Koyanagi-Harada (VKH) disease (n = 12) and control subjects (n = 9) were assayed with the use of a multiplex assay.

Results: Of all the IL-10 family cytokines studied, only IL-19 levels were significantly higher in AH samples of patients (n = 45) than in controls (p = 0.022). When comparing the four individual disease groups to controls, IL-19 levels were only significantly higher in HLA-B27-associated uveitis (p < 0.001). IL-19 levels were significantly higher in patients with HLA-B27-associated uveitis than in patients with BD, sarcoidosis and VKH disease (p < 0.001; p = 0.002; p < 0.001, respectively). Significant correlations were found between AH levels of IL-19 and AH levels of TNF-α, (r = 0.3; p = 0.03) and IL-1β (r = 0.56; p < 0.001).

Conclusions: Among the IL-10 family cytokines analysed, IL-19 demonstrated the highest expression in endogenous uveitis, particularly in HLA-B27-associated uveitis. IL-19 thus might assist in the regulation of inflammation in HLA-B27-associated uveitis.
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http://dx.doi.org/10.1111/aos.14039DOI Listing
August 2019

Pathological roles of the homeostatic chemokine CXCL12.

Cytokine Growth Factor Rev 2018 12 23;44:51-68. Epub 2018 Oct 23.

KU Leuven, University of Leuven, Department of Microbiology and Immunology, Rega Institute for Medical Research, Laboratory of Molecular Immunology, B-3000 Leuven, Belgium. Electronic address:

CXCL12 is a CXC chemokine that traditionally has been classified as a homeostatic chemokine. It contributes to physiological processes such as embryogenesis, hematopoiesis and angiogenesis. In contrast to these homeostatic functions, increased expression of CXCL12 in general, or of a specific CXCL12 splicing variant has been demonstrated in various pathologies. In addition to this increased or differential transcription of CXCL12, also upregulation of its receptors CXC chemokine receptor 4 (CXCR4) and atypical chemokine receptor 3 (ACKR3) contributes to the onset or progression of diseases. Moreover, posttranslational modification of CXCL12 during disease progression, through interaction with locally produced molecules or enzymes, also affects CXCL12 activity, adding further complexity. As CXCL12, CXCR4 and ACKR3 are broadly expressed, the number of pathologies wherein CXCL12 is involved is growing. In this review, the role of the CXCL12/CXCR4/ACKR3 axis will be discussed for the most prevalent pathologies. Administration of CXCL12-neutralizing antibodies or small-molecule antagonists of CXCR4 or ACKR3 delays disease onset or prevents disease progression in cancer, viral infections, inflammatory bowel diseases, rheumatoid arthritis and osteoarthritis, asthma and acute lung injury, amyotrophic lateral sclerosis and WHIM syndrome. On the other hand, CXCL12 has protective properties in Alzheimer's disease and multiple sclerosis, has a beneficial role in wound healing and has crucial homeostatic properties in general.
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http://dx.doi.org/10.1016/j.cytogfr.2018.10.004DOI Listing
December 2018

Chemokine-Induced Macrophage Polarization in Inflammatory Conditions.

Front Immunol 2018 7;9:1930. Epub 2018 Sep 7.

Laboratory of Molecular Immunology, Department of Microbiology and Immunology, REGA Institute KU Leuven, Leuven, Belgium.

Macrophages represent a heterogeneous cell population and are known to display a remarkable plasticity. In response to distinct micro-environmental stimuli, e.g., tumor stroma vs. infected tissue, they polarize into different cell subtypes. Originally, two subpopulations were defined: classically activated macrophages or M1, and alternatively activated macrophages or M2. Nowadays, the M1/M2 classification is considered as an oversimplified approach that does not adequately cover the total spectrum of macrophage phenotypes observed . Especially in pathological circumstances, macrophages behave as plastic cells modifying their expression and transcription profile along a continuous spectrum with M1 and M2 phenotypes as extremes. Here, we focus on the effect of chemokines on macrophage differentiation and polarization in physiological and pathological conditions. In particular, we discuss chemokine-induced macrophage polarization in inflammatory diseases, including obesity, cancer, and atherosclerosis.
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http://dx.doi.org/10.3389/fimmu.2018.01930DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6137099PMC
September 2019

The CC chemokines CCL8, CCL13 and CCL20 are local inflammatory biomarkers of HLA-B27-associated uveitis.

Acta Ophthalmol 2019 Feb 21;97(1):e122-e128. Epub 2018 Sep 21.

Rega Institute for Medical Research, Department of Microbiology and Immunology, University of Leuven, KU Leuven, Leuven, Belgium.

Purpose: To determine the concentrations of the CC chemokines CCL2, CCL7, CCL8, CCL11, CCL13, CCL20, CCL24 and CCL26 in aqueous humour (AH) samples from patients with specific uveitic entities.

Methods: Aqueous humour samples from patients with active uveitis associated with Behçet's disease (BD) (n = 13), sarcoidosis (n = 8), HLA-B27-related inflammation (n = 12), Vogt-Koyanagi-Harada (VKH) disease (n = 12) and control patients (n = 9) were assayed with the use of a multiplex assay.

Results: When considering all uveitis patients as one group, all chemokine levels except CCL2 were significantly increased compared to controls. CCL8, CCL13 and CCL20 were the most strongly upregulated, 48-fold, 118-fold and 173-fold, respectively, above control AH levels. CCL8 and CCL13 levels were significantly higher in HLA-B27-associated uveitis than in sarcoidosis and VKH disease. CCL20 levels were significantly higher in HLA-B27-associated uveitis than in BD, sarcoidosis and VKH disease. In addition, CCL20 levels were significantly higher in BD than in VKH disease. In HLA-B27-associated uveitis, CCL8, CCL13 and CCL20 were upregulated 111-fold, 255-fold and 465-fold, respectively, compared with controls. CCL8, CCL13 and CCL20 levels were significantly higher in nongranulomatous uveitis (BD and HLA-B27-associated uveitis) than in granulomatous uveitis (sarcoidosis and VKH disease).

Conclusion: Immune responses mediated by CCL8, CCL13 and CCL20 appear to be more potent in nongranulomatous uveitis, particularly in HLA-B27-associated uveitis.
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http://dx.doi.org/10.1111/aos.13835DOI Listing
February 2019

Peroxynitrite Exposure of CXCL12 Impairs Monocyte, Lymphocyte and Endothelial Cell Chemotaxis, Lymphocyte Extravasation and Anti-HIV-1 Activity.

Front Immunol 2018 28;9:1933. Epub 2018 Aug 28.

Laboratory of Molecular Immunology, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven, University of Leuven, Leuven, Belgium.

CXCL12 is a chemotactic cytokine that attracts many different cell types for homeostasis and during inflammation. Under stress conditions, macrophages and granulocytes produce factors such as peroxynitrite as a consequence of their oxidative response. After short incubations of CXCL12 with peroxynitrite, the gradual nitration of Tyr7, Tyr61, or both Tyr7 and Tyr61 was demonstrated with the use of mass spectrometry, whereas longer incubations caused CXCL12 degradation. Native CXCL12 and the nitrated forms, [3-NT]CXCL12 and [3-NT]CXCL12, were chemically synthesized to evaluate the effects of Tyr nitration on the biological activity of CXCL12. All CXCL12 forms had a similar binding affinity for heparin, the G protein-coupled chemokine receptor CXCR4 and the atypical chemokine receptor ACKR3. However, nitration significantly enhanced the affinity of CXCL12 for chondroitin sulfate. Internalization of CXCR4 and β-arrestin 2 recruitment to CXCR4 was significantly reduced for [3-NT]CXCL12 compared to CXCL12, whereas β-arrestin 2 recruitment to ACKR3 was similar for all CXCL12 variants. [3-NT]CXCL12 was weaker in calcium signaling assays and in i chemotaxis assays with monocytes, lymphocytes and endothelial cells. Surprisingly, nitration of Tyr61, but not Tyr7, partially protected CXCL12 against cleavage by the specific serine protease CD26. , the effects were more pronounced compared to native CXCL12. Nitration of any Tyr residue drastically lowered lymphocyte extravasation to joints compared to native CXCL12. Finally, the anti-HIV-1 activity of [3-NT]CXCL12 and [3-NT]CXCL12 was reduced, whereas CXCL12 and [3-NT]CXCL12 were equally potent. In conclusion, nitration of CXCL12 occurs readily upon contact with peroxynitrite and specifically nitration of Tyr7 fully reduces its and biological activities.
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http://dx.doi.org/10.3389/fimmu.2018.01933DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6127631PMC
September 2019

Anti-inflammatory effects of the GAG-binding CXCL9(74-103) peptide in dinitrofluorobenzene-induced contact hypersensitivity in mice.

Clin Exp Allergy 2018 10 14;48(10):1333-1344. Epub 2018 Aug 14.

Laboratory of Molecular Immunology, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven, Leuven, Belgium.

Background: To recruit leucocytes to an inflammatory site, chemokine binding to glycosaminoglycans (GAGs) is critical. Therefore, strategies to interfere with this interaction, aiming at the production of anti-inflammatory agents, were developed. These include production of modified chemokines without affinity for G protein-coupled receptors but with enhanced affinity for GAGs. Such modified chemokines compete with functional chemokines for GAG binding, prevent chemokine immobilization and presentation, and inhibit leucocyte migration. In addition to modified chemokines, a GAG-binding peptide consisting of the 30 COOH-terminal residues of CXCL9, that is CXCL9(74-103), inhibited CXCL8- and monosodium urate crystal-induced neutrophil migration.

Objective: We wanted to explore whether interference with chemokine-GAG interactions by CXCL9(74-103) reduces inflammation in neutrophil-dependent dinitrofluorobenzene-induced contact hypersensitivity.

Methods: For this study, we evaluated several inflammatory parameters, including ear swelling and the levels of chemokines, cytokines, proteases and neutrophils in the ears of dinitrofluorobenzene-induced mice treated with CXCL9(74-103) or buffer.

Results: One intravenous injection of CXCL9(74-103), just before painting with dinitrofluorobenzene on the ear, did not affect protein levels of the major murine neutrophil attractant, that is CXCL6, in this contact hypersensitivity model. However, IL-6, CXCL1, CCL2 and matrix metalloproteinase-9 (MMP-9) protein concentrations and peroxidase activity in challenged ears were reduced. In addition, intravenous injection of the CXCL9-derived peptide led to a reduced ear swelling response, indicating that the locally produced chemokines were hindered to attract leucocytes. The inhibiting potential of CXCL9(74-103) was explained by its competition for GAG binding with CXCL1, CXCL6 and CCL3 and inhibition of transendothelial migration of neutrophils to CXCL6.

Conclusions And Clinical Relevance: The CXCL9(74-103) peptide inhibited dinitrofluorobenzene-induced infiltration of neutrophils and neutrophil-dependent inflammation in ears. Therefore, CXCL9(74-103) may be a lead molecule for the development of therapeutic peptides or peptide derivatives that compete with functional chemokines for GAG binding.
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http://dx.doi.org/10.1111/cea.13227DOI Listing
October 2018

Matrix Metalloproteinase-9-Generated COOH-, but Not NH-Terminal Fragments of Serum Amyloid A1 Retain Potentiating Activity in Neutrophil Migration to CXCL8, With Loss of Direct Chemotactic and Cytokine-Inducing Capacity.

Front Immunol 2018 4;9:1081. Epub 2018 Jun 4.

Laboratory of Molecular Immunology, Department of Microbiology and Immunology, Rega Institute for Medical Research, University of Leuven, Leuven, Belgium.

Serum amyloid A1 (SAA1) is a prototypic acute phase protein, induced to extremely high levels by physical insults, including inflammation and infection. Human SAA and its NH-terminal part have been studied extensively in the context of amyloidosis. By contrast, little is known about COOH-terminal fragments of SAA. Intact SAA1 chemoattracts leukocytes the G protein-coupled receptor formyl peptide receptor like 1/formyl peptide receptor 2 (FPR2). In addition to direct leukocyte activation, SAA1 induces chemokine production by signaling through toll-like receptor 2. We recently discovered that these induced chemokines synergize with intact SAA1 to chemoattract leukocytes and . Gelatinase B or matrix metalloproteinase-9 (MMP-9) is also induced by SAA1 during infection and inflammation and processes many substrates in the immune system. We demonstrate here that MMP-9 rapidly cleaves SAA1 at a known consensus sequence that is also present in gelatins. Processing of SAA1 by MMP-9 at an accessible loop between two alpha helices yielded predominantly three COOH-terminal fragments: SAA1(52-104), SAA1(57-104), and SAA1(58-104), with a relative molecular mass of 5,884.4, 5,327.3, and 5,256.3, respectively. To investigate the effect of proteolytic processing on the biological activity of SAA1, we chemically synthesized the COOH-terminal SAA fragments SAA1(52-104) and SAA1(58-104) and the complementary NH-terminal peptide SAA1(1-51). In contrast to intact SAA1, the synthesized SAA1 peptides did not induce interleukin-8/CXCL8 in monocytes or fibroblasts. Moreover, these fragments possessed no direct chemotactic activity for neutrophils, as observed for intact SAA1. However, comparable to intact SAA1, SAA1(58-104) cooperated with CXCL8 in neutrophil activation and migration, whereas SAA1(1-51) lacked this potentiating activity. This cooperative interaction between the COOH-terminal SAA1 fragment and CXCL8 in neutrophil chemotaxis was mediated by FPR2. Hence, proteolytic cleavage of SAA1 by MMP-9 fine tunes the inflammatory capacity of this acute phase protein in that only the synergistic interactions with chemokines remain to prolong the duration of inflammation.
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http://dx.doi.org/10.3389/fimmu.2018.01081DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5994419PMC
July 2019

CXCL4 and CXCL4L1 in cancer.

Cytokine 2018 09;109:65-71

KU Leuven, University of Leuven, Department of Microbiology and Immunology, Rega Institute for Medical Research, Laboratory of Molecular Immunology, B-3000 Leuven, Belgium. Electronic address:

Chemokines regulate leukocyte migration during physiological and pathological conditions. It is currently accepted that these chemotactic cytokines are also important in the development and progression of cancer. CXCL4 and its non-allelic variant CXCL4L1 are two platelet-associated chemokines that have been attributed anti-tumoral activity as a result of their angiostatic potential and the chemotactic activity for anti-tumoral leukocytes. Here we review the role of CXCL4 and CXCL4L1 in cancer, the use of both chemokines as cancer biomarkers and discuss some possible therapeutic opportunities.
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http://dx.doi.org/10.1016/j.cyto.2018.02.022DOI Listing
September 2018

How post-translational modifications influence the biological activity of chemokines.

Cytokine 2018 09;109:29-51

Laboratory of Molecular Immunology, Department of Microbiology and Immunology, Rega Institute for Medical Research, KU Leuven, Leuven, Belgium. Electronic address:

Chemokines are important proteins involved in the regulation of directed leukocyte migration during inflammation and the homeostatic homing of immune cells. In addition, they play a role in angiogenesis, hematopoiesis, organogenesis, tumor growth and metastasis. Therefore, the chemokine/chemokine receptor network is highly complex and needs to be tightly controlled. An important mechanism of fine-tuning chemokine activity and reducing its apparent redundancy is post-translational modification (PTM) of chemokines and their receptors. Under inflammatory conditions, enzymes such as matrix metalloproteinases (MMPs), plasmin, CD13, CD26, and peptidylarginine deiminases (PADs) and protein-modifying agents, such as peroxynitrite, are upregulated and released and may provoke truncation, degradation, nitration or citrullination of chemokines. Most modified chemokines show altered biological activity. This review reports how PTMs influence the biological functions of chemokines, with special attention for the impact beyond chemotaxis.
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http://dx.doi.org/10.1016/j.cyto.2018.02.026DOI Listing
September 2018

The ectoenzyme-side of matrix metalloproteinases (MMPs) makes inflammation by serum amyloid A (SAA) and chemokines go round.

Immunol Lett 2019 01 2;205:1-8. Epub 2018 Jun 2.

KU Leuven, University of Leuven, Department of Microbiology and Immunology, Rega Institute for Medical Research, Laboratory of Molecular Immunology, Herestraat 49 - box 1042, 3000, Leuven, Belgium. Electronic address:

During an inflammatory response, a large number of distinct mediators appears in the affected tissues or in the blood circulation. These include acute phase proteins such as serum amyloid A (SAA), cytokines and chemokines and proteolytic enzymes. Although these molecules are generated within a cascade sequence in specific body compartments allowing for independent action, their co-appearance in space and time during acute or chronic inflammation points toward important mutual interactions. Pathogen-associated molecular patterns lead to fast induction of the pro-inflammatory endogenous pyrogens, which are evoking the acute phase response. Interleukin-1, tumor necrosis factor-α and interferons simultaneously trigger different cell types, including leukocytes, endothelial cells and fibroblasts for tissue-specific or systemic production of chemokines and matrix metalloproteinases (MMPs). In addition, SAA induces chemokines and both stimulate secretion of MMPs from multiple cell types. As a consequence, these mediators may cooperate to enhance the inflammatory response. Indeed, SAA synergizes with chemokines to increase chemoattraction of monocytes and granulocytes. On the other hand, MMPs post-translationally modify chemokines and SAA to reduce their activity. Indeed, MMPs internally cleave SAA with loss of its cytokine-inducing and direct chemotactic potential whilst retaining its capacity to synergize with chemokines in leukocyte migration. Finally, MMPs truncate chemokines at their NH- or COOH-terminal end, resulting in reduced or enhanced chemotactic activity. Therefore, the complex interactions between chemokines, SAA and MMPs either maintain or dampen the inflammatory response.
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http://dx.doi.org/10.1016/j.imlet.2018.06.001DOI Listing
January 2019
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