| 1. |
- Bengtsson, Torbjörn, 1955-, et al.
(författare)
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Role of the actin cytoskeleton during respiratory burst in chemoattractant-stimulated neutrophils
- 2006
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Ingår i: Cell Biology International. - : Wiley. - 1065-6995 .- 1095-8355. ; 30:2, s. 154-163
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Tidskriftsartikel (refereegranskat)abstract
- The aim of this study was to clarify the role of the actin cytoskeleton during chemotactic peptide fMet-Leu-Phe (fMLF)-stimulated respiratory burst in human neutrophil granulocytes. Reactive oxygen species (ROS) was measured as luminol-amplified chemiluminescence (CL) and F-actin content as bodipy phallacidin fluorescence in neutrophils treated with latrunculin B or jasplakinolide, an inhibitor and activator of actin polymerization, respectively. Latrunculin B markedly decreased, whereas jasplakinolide increased, the F-actin content in neutrophils, unstimulated or stimulated with fMLF. Latrunculin B enhanced the fMLF-triggered ROS-production more than tenfold. Jasplakinolide initially inhibited the fMLF-induced CL-response, however, caused a potent second sustained phase (>400% of control). Both actin drugs triggered a substantial CL-response when added 5-25 min after fMLF. This was also valid for chemotactic doses of fMLF, where latrunculin B and jasplakinolide amplified the ROS-production 5-10 times. By using specific signal transduction inhibitors, we found that the NADPH oxidase activation triggered by destabilization of the actin cytoskeleton occurs downstream of phospholipase C and protein kinase C but is mediated by Rho GTPases and tyrosine phosphorylation. In conclusion, rearrangements of the actin cytoskeleton are a prerequisite in connecting ligand/receptor activation, generation of second messengers and assembly of the NADPH oxidase in neutrophil granulocytes. © 2005 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved.
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| 2. |
- Nimeri, G, et al.
(författare)
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The influence of plasma proteins and platelets on oxygen radical production and F-actin distribution in neutrophils adhering to polymer surfaces
- 2002
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Ingår i: Biomaterials. - 0142-9612 .- 1878-5905. ; 23:8, s. 1785-1795
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Tidskriftsartikel (refereegranskat)abstract
- It is well known that blood cell interactions with artificial surfaces might have deleterious effects on host tissue, however, the mechanisms involved are far from understood. In this study, neutrophil-platelet interaction on uncoated or protein-coated polymer surfaces was investigated. Cell spreading, reorganization of actin filaments and release of oxygen metabolites (measured as luminol-amplified chemiluminescence) were used as criteria for cell activation on positively charged, hydrophilic 1,2-diaminocyclohexane, and negatively charged, hydrophobic hexamethylene-disiloxane. The model surfaces were made by radio frequency plasma discharge polymerization. Neutrophil contact with the uncoated polymers induced a prolonged generation of oxygen radicals. Precoating of the polymer surfaces with human serum albumin (HSA) or fibrinogen, markedly reduced neutrophil activation, whereas coating with human immunoglobulin G (IgG), a well-known opsonin, resulted in significantly higher levels of cell activation. Consequently, protein coating overruled the activating effects of the polymer surfaces. The presence of unstimulated or thrombin-stimulated platelets markedly increased the reactivity of neutrophils against fibrinogen- and IgG-coated surfaces. However, neutrophils remained relatively unreactive in the presence of platelets on HSA-treated surfaces. Comparison of the different types of surfaces used, reveals a correlation between the degree of cell spreading, reorganization of the actin cytoskeleton and the amount of oxygen radicals produced. Our results suggest that the acute inflammatory reaction on a biomaterial surface is highly dependent on the nature and composition of the first adsorbed protein layer and the extent of platelet activation.
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| 3. |
- Sjöwall, Christoffer, et al.
(författare)
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Solid-phase classical complement activation by C-reactive protein (CRP) is inhibited by fluid-phase CRP-C1q interaction.
- 2007
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Ingår i: Biochemical and biophysical research communications. - : Elsevier BV. - 0006-291X .- 1090-2104. ; 352:1, s. 251-8
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Tidskriftsartikel (refereegranskat)abstract
- C-reactive protein (CRP) interacts with phosphorylcholine (PC), Fcgamma receptors, complement factor C1q and cell nuclear constituents, yet its biological roles are insufficiently understood. The aim was to characterize CRP-induced complement activation by ellipsometry. PC conjugated with keyhole limpet hemocyanin (PC-KLH) was immobilized to cross-linked fibrinogen. A low-CRP serum with different amounts of added CRP was exposed to the PC-surfaces. The total serum protein deposition was quantified and deposition of IgG, C1q, C3c, C4, factor H, and CRP detected with polyclonal antibodies. The binding of serum CRP to PC-KLH dose-dependently triggered activation of the classical pathway. Unexpectedly, the activation was efficiently down-regulated at CRP levels > 150 mg/L. Using radial immunodiffusion, CRP-C1q interaction was observed in serum samples with high CRP concentrations. We propose that the underlying mechanism depends on fluid-phase interaction between C1q and CRP. This might constitute another level of complement regulation, which has implications for systemic lupus erythematosus where CRP is often low despite flare-ups.
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| 4. |
- Skoglund, Caroline, 1981-, et al.
(författare)
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C-reactive protein and C1q regulate platelet adhesion and activation on adsorbed immunoglobulin G and albumin.
- 2008
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Ingår i: Immunology and cell biology. - : Wiley. - 0818-9641 .- 1440-1711. ; 86:5, s. 466-74
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Tidskriftsartikel (refereegranskat)abstract
- Blood platelets and C-reactive protein (CRP) are both used clinically as markers of ongoing inflammation, and both participate actively in inflammatory responses, although the biological effects are still incompletely understood. Rapidly adhering platelets express receptors for complement factor 1q (C1q) and the Fc part of immunoglobulin G (IgG), and CRP is known to activate/regulate complement via C1q binding, and to ligate FcgammaRs. In the present study, we used normal human IgG pre-adsorbed to a well-characterized methylated surface as a model solid-phase immune complex when investigating the effects of CRP and C1q on platelet adhesion and activation. Protein adsorption was characterized using ellipsometry and polyclonal antibodies, and human serum albumin (HSA) and non-coated surfaces were used as reference surfaces. Platelet adhesion to IgG and HSA was inhibited by both C1q and CRP. Furthermore, CRP (moderately) and C1q (markedly) decreased the spreading of adhering platelets. The combination of C1q and CRP was slightly more potent in reducing cell adhesion to IgG, and also impaired the adhesion to HSA and non-coated surfaces. Platelet production of thromboxane B2 (TXB(2)) was also reduced by C1q both in the presence and absence of CRP, whereas CRP alone had no effect on TXB(2) production. We conclude that CRP and C1q regulate the behaviour of platelets, and that this may be an important immunoregulatory mechanism during inflammatory conditions.
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| 5. |
- Skoglund, Caroline, 1981-, et al.
(författare)
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C-reactive protein inhibit complement-mediated platelet activation suggesting a protective role in atherogenesis
- 2006
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Ingår i: Atherosclerosis Supplements. - Clare, Ireland : Elsevier. - 1567-5688 .- 1878-5050. ; 7:3, s. 284-284
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Objective: C-reactive protein (CRP) represents a powerful predictor of coro- nary artery disease. However, its physiological role is not fully understood. The binding of CRP to its ligand phosphorylcholine (PC) activates the com- plement system via the classical pathway, although limited to the initial stages, i.e. no membrane attack complex is formed. The aim of this study was to chaxacterize CRP-induced complement activation on PC-coated surfaces, and to investigate the regulatory effects of PC-bound crp on complement induced platelet activation.Methods: PC conjugated to keyhole limpet hemocyanin was immobilized to cross-linked fibrinogen on silica particles. Ellipsometry and polyclonal anti- bodies were used to quantify deposition of serum proteins, complement factors and CRP on the surfaces. Washed platelets as well as serum were prepared according to standard protocols. CRP concentrations were measured with a high sensitivity assay. Lumi-aggregometry was used to evaluate the effects of PC-coated particles and CRP on complement-induced platelet aggregation and secretion.Results: Serum (5%) induced platelet aggregation and secretion through complement-dependent mechanisms. PC-coated particles antagonized the complement-mediated platelet activation but only if CRP was present. Inter- estingly, we found that a minor elevation of CRR below 5 rag/1 was sufficient to inhibit platelet activation.Conclusions: We suggest that CRP bound to PC-expressing ligands, e.g. bacteria or modified low-density lipoproteins in an atherosclerotic lesion, modulate complement activation and thereby prevent a harmful platelet activation.
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| 6. |
- Skoglund, Caroline, et al.
(författare)
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C1q regulates collagen-dependent production of reactive oxygen species, aggregation and levels of soluble P-selectin in whole blood
- 2012
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Ingår i: Immunology Letters. - Amsterdam, Netherlands : Elsevier. - 0165-2478 .- 1879-0542. ; 142:1-2, s. 28-33
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Tidskriftsartikel (refereegranskat)abstract
- Blood platelets express several receptors involved in immunity (e.g. complement-, toll-like- and Fc gamma-receptors) and release inflammatory mediators. Furthermore, formation of platelet-leukocyte aggregates has an important role during inflammatory conditions such as coronary artery disease. Thus, apart from their well-known role in haemostasis, platelets are today also recognized as cells with immunomodulatory properties. We have previously reported regulatory effects of complement protein 1q (C1q) on platelet activation in experimental setups using isolated cells. In the present study we have proceeded by investigating effects of C1q on collagen-induced aggregation, production of reactive oxygen species (ROS), formation of platelet-leukocyte aggregates and levels of soluble P-selectin in whole blood. Impedance measurements showed that C1q inhibited collagen-induced aggregation whereas it potentiated the collagen-provoked production of ROS in a luminol-dependent chemiluminescence assay. The effects of C1q on aggregation and ROS-production were dependent upon platelets, as they were no longer observed in presence of the platelet (GpIIb/IIIa) inhibitor Reopro. Furthermore, the levels of soluble P-selectin were found to be lowered upon treatment with C1q prior to addition of collagen. There was also a trend towards a decreased formation of large platelet-leukocyte aggregates in collagen-stimulated whole blood following C1q treatment. In conclusion, our data indicate that C1q could have a role in regulating platelet activation and associated leukocyte recruitment during vessel wall injury. This has implications for inflammatory disorders such as coronary artery disease.
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| 7. |
- Wetterö, Jonas, 1972-, et al.
(författare)
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C1q-independent activation of neutrophils by immunoglobulin M-coated surfaces
- 2001
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Ingår i: Journal of Biomedical Materials Research. - 0021-9304 .- 1097-4636. ; 57:4, s. 550-558
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Tidskriftsartikel (refereegranskat)abstract
- Neutrophil granulocytes are known to rapidly adhere and undergo frustrated phagocytosis upon contact with immunoglobulin and/or complement protein opsonized artificial surfaces. In this study, we examined the relation between serum protein deposition and human neutrophil activation on hydrophobic glass and silicon model surfaces that were coated with immunoglobulin G or M (IgG/IgM), both initiators of the classical complement pathway. Protein adsorption from normal human serum (NHS) was quantified with null-ellipsometry combined with antibody techniques. The neutrophil oxygen radical production was registered by luminol-amplified chemiluminescence (CL) and the morphology, as well as changes in the content of filamentous actin (F-actin), were documented by fluorescence microscopy. Complement factor 3 (C3) bound to both IgG- and IgM-coated surfaces, but surprisingly C1q was found only on IgG-coated surfaces. Both immunoglobulins triggered complement dependent neutrophil activation. However, CL and F-actin accumulation were found sensitive to the presence of C1q in the serum only at the IgG-coated surface. We suggest that spontaneously adsorbed IgM activates the complement system and interacts with neutrophils by C1q-independent mechanisms.
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| 8. |
- Wetterö, Jonas, 1972-, et al.
(författare)
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Complement activation on immunoglobulin G-coated hydrophobic surfaces enhances the release of oxygen radicals from neutrophils through an actin-dependent mechanism
- 2000
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Ingår i: Journal of Biomedical Materials Research. - 0021-9304 .- 1097-4636. ; 51:4, s. 742-751
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Tidskriftsartikel (refereegranskat)abstract
- Neutrophil granulocytes are among the first cells to encounter a plasma protein-coated implant and may through frustrated phagocytosis release toxic oxidative species. We used two model surfaces, hydrophobic and hydrophilic glass, to investigate the effects of plasma immunoglobulin G (IgG)-complement interactions for neutrophil adhesion and respiratory burst. The respiratory burst was measured with luminol-amplified chemiluminescence and cell adhesion was determined by labeling neutrophils with 2′, 7′-bis-(carboxy-ethyl)-5(6)-carboxyfluorescein. We demonstrate that the IgG-triggered neutrophil adhesion and oxygen radical production is augmented in the presence of normal human serum, in particular on hydrophobic surfaces, indicating that complement factors enhance the neutrophil activation. We propose that the complement factors C3, C5a, and C1q are especially important for this amplification, but factor B is probably not. Disturbance of the actin filament dynamics with cytochalasin B or jasplakinolide blocked the neutrophil radical generation on all surfaces. However, these drugs did not affect the number of adherent neutrophils. We suggest that there is a synergistic interaction between adsorbed IgG, and the complement system, which amplifies the neutrophil acute inflammatory responses through a dynamic actin cytoskeleton on synthetic surfaces.
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| 9. |
- Wetterö, Jonas, 1972-, et al.
(författare)
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On the binding of complement to solid artificial surfaces in vitro
- 2002
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Ingår i: Biomaterials. - 0142-9612 .- 1878-5905. ; 23:4, s. 981-991
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Tidskriftsartikel (refereegranskat)abstract
- Since the realization of a complement activation capacity by artificial surfaces upon contact with blood, a common belief has evolved that charged nucleophilic surface groups such as amine (–NH2) and hydroxyl (–OH) react with and eventually bind to the internal thioester in complement factor 3 (C3). A covalent amide or ester linkage is thereby supposed to form between C3b and the surface itself. In this report, we present complement surface binding data by null-ellipsometry for two nucleophilic surfaces (–NH2 and –OH), for surfaces with immunoglobulin G (IgG) covalently bound, and for IgG spontaneously pre-adsorbed to hydrophobic silicon. The results reveal that the plasma proteins that were deposited during complement activation became eluted by sodium dodecyl sulfate. Hence the direct covalent binding between C3 and solid nucleophilic surfaces seems to be only of moderate importance, at least during shorter serum incubations. This strongly suggests that the prevalent covalent linkage model between solid artificial surfaces and C3b is not accurate. Instead we suggest a more pronounced role for C3 associations to other adsorbed proteins and/or electrostatic and hydrophobic protein–surface interactions.
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