Role of complement and Fcγ receptors in the protective activity of the long pentraxin PTX3 against Aspergillus fumigatus

Blood ◽  
2010 ◽  
Vol 116 (24) ◽  
pp. 5170-5180 ◽  
Author(s):  
Federica Moalli ◽  
Andrea Doni ◽  
Livija Deban ◽  
Teresa Zelante ◽  
Silvia Zagarella ◽  
...  
Keyword(s):  
Aspergillus Fumigatus ◽  
Alternative Pathway ◽  
Complement Receptor ◽  
Protective Activity ◽  
Opsonic Activity ◽  
Reactive Protein ◽  
Complement Receptor 3 ◽  
Dependent Pathway

AbstractPentraxin 3 (PTX3) is a soluble pattern recognition molecule playing a nonredundant role in resistance against Aspergillus fumigatus. The present study was designed to investigate the molecular pathways involved in the opsonic activity of PTX3. The PTX3 N-terminal domain was responsible for conidia recognition, but the full-length molecule was necessary for opsonic activity. The PTX3-dependent pathway of enhanced neutrophil phagocytic activity involved complement activation via the alternative pathway; Fcγ receptor (FcγR) IIA/CD32 recognition of PTX3-sensitized conidia and complement receptor 3 (CR3) activation; and CR3 and CD32 localization to the phagocytic cup. Gene targeted mice (ptx3, FcR common γ chain, C3, C1q) validated the in vivo relevance of the pathway. In particular, the protective activity of exogenous PTX3 against A fumigatus was abolished in FcR common γ chain-deficient mice. Thus, the opsonic and antifungal activity of PTX3 is at the crossroad between complement, complement receptor 3-, and FcγR-mediated recognition. Because short pentraxins (eg, C-reactive protein) interact with complement and FcγR, the present results may have general significance for the mode of action of these components of the humoral arm of innate immunity.

scholarly journals β-Glucan enhances complement-mediated hematopoietic recovery after bone marrow injury

Blood ◽  
2006 ◽  
Vol 107 (2) ◽  
pp. 835-840 ◽  
Author(s):  
Daniel E. Cramer ◽  
Daniel J. Allendorf ◽  
Jarek T. Baran ◽  
Richard Hansen ◽  
Jose Marroquin ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Mouse Serum ◽  
Complement Receptor ◽  
Dependent Manner ◽  
Complement Receptor 3 ◽  
Total Body ◽  
Stimulating Factor ◽  
Sublethal Irradiation ◽  
Classic Pathway

AbstractMyelotoxic injury in the bone marrow (BM) as a consequence of total body irradiation (TBI) or granulocyte colony-stimulating factor (G-CSF) mobilization results in the deposition of iC3b on BM stroma (stroma-iC3b). In the present study, we have examined how stroma-iC3b interacts with hematopoietic progenitor cells (HPCs) and the role of complement (C) and complement receptor 3 (CR3) in BM injury/repair. We demonstrate here that stroma-iC3b tethers HPCs via the inserted (I) domain of HPC complement receptor 3 (CR3, CD11b/CD18, Mac-1). Following irradiation, stroma-iC3b was observed in the presence of purified IgM and normal mouse serum (NMS), but not serum from Rag-2-/- mice, implicating a role for antibody (Ab) and the classic pathway of C activation. Furthermore, a novel role for soluble yeast β-glucan, a ligand for the CR3 lectin-like domain (LLD), in the priming of CR3+ HPC is suggested. Soluble yeast β-glucan could enhance the proliferation of tethered HPCs, promote leukocyte recovery following sublethal irradiation, and increase the survival of lethally irradiated animals following allogeneic HPC transplantation in a CR3-dependent manner. Taken together, these observations suggest a novel role for C, CR3, and β-glucan in the restoration of hematopoiesis following injury. (Blood. 2006;107:835-840)

Abstract 277: Elevated C-Reactive Protein Contributes to Preeclampsia via Kinin Signaling Pathways

Hypertension ◽  
2013 ◽  
Vol 62 (suppl_1) ◽  
Author(s):  
Nicholas Parchim ◽  
Wei Wang ◽  
Takayuki Iriyama ◽  
Chen Liu ◽  
Athar H Siddiqui ◽  
...  
Keyword(s):  
Acute Phase Reactant ◽  
Receptor Activation ◽  
C Reactive Protein ◽  
Direct Role ◽  
Human Trophoblast ◽  
Reactive Protein ◽  
Induced Hypertension ◽  
Pregnant Mice

Preeclampsia (PE) is a serious pregnancy disease characterized by hypertension and proteinuria. Despite intensive research efforts, the underlying cause of PE remains a mystery. PE is, however, associated with abnormalities of the immune system. Here we report that the levels of C-reactive protein (CRP), an important acute phase reactant, were significantly elevated in the plasma of human with PE at the third trimester. Next, we found that CRP protein levels in the placentas of PE patients were also significantly increased compared to controls. In an effort to determine the exact role of elevated CRP in PE, we infused CRP into pregnant mice. We found that injection of CRP into pregnant mice induced hypertension (170 mmHg mean systolic vs. 125 mmHg mean systolic control; p<0.05) and proteinuria (25 mg/ug vs 12 mg/ug vehicle; p<0.05), indicating the direct role of CRP in PE. CRP is known to bind with phosphocholine on damaged cell membranes. Recent studies identified that neurokinin B (NKB), a placental enriched neuropeptide and known pathogenic molecule for PE, is phosphocholinated. This posttranslational modification increases its stability and enhances NKB-mediated receptor activation. These findings raise an intriguing hypothesis that CRP may bind with NKB coupled to NK3R activation and contribute to PE. To test this hypothesis, we conducted a pulldown assay, and we found that CRP bound with NKB. Next, using a cellular invasion assay, we revealed that CRP decreased invasion of human trophoblast cells (0.7 to 0.07 invasion index, p<0.05), while treatment with an NK3R selective antagonist, SB222200, ameliorated this shallow invasion. Finally, we provided in vivo evidence that inhibition of NK3R by SB222200 or knockdown of NK3R by specific siRNA in a potent nanoparticle delivery system significantly reduced CRP-induced hypertension and proteinuria in pregnant mice (170 mmHg mean systolic CRP-injected vs. 130 mmHg mean systolic siRNA NK3R; p<0.05 and proteinuria 25 mg/ug vs. 15 mg/ug; p<0.05). Overall, our findings demonstrate that elevated CRP contributes to PE and NKB/NK3R is a novel mechanism underlying CRP-mediated shallow invasion and disease development. These studies suggest novel pathogenic biomarkers and innovative therapeutic targets for PE.

scholarly journals CD44-mediated phagocytosis induces inside-out activation of complement receptor-3 in murine macrophages

Blood ◽  
2007 ◽  
Vol 110 (13) ◽  
pp. 4492-4502 ◽  
Author(s):  
Eric Vachon ◽  
Raiza Martin ◽  
Vivian Kwok ◽  
Vera Cherepanov ◽  
Chung-Wai Chow ◽  
...  
Keyword(s):  
Intracellular Signaling ◽  
Immunofluorescence Microscopy ◽  
Complement Receptor ◽  
Fcγ Receptors ◽  
Intracellular Signaling Pathways ◽  
Large Particles ◽  
Complement Receptor 3 ◽  
Inside Out ◽  
Β2 Integrins

Diverse receptors, including Fcγ receptors and β2 integrins (complement receptor-3 [CR3], CD11b/CD18), have been implicated in phagocytosis, but their distinct roles and interactions with other receptors in particle engulfment are not well defined. CD44, a transmembrane adhesion molecule involved in binding and metabolism of hyaluronan, may have additional functions in regulation of inflammation and phagocytosis. We have recently reported that CD44 is a fully competent phagocytic receptor that is able to trigger ingestion of large particles by macrophages. Here, we investigated the role of coreceptors and intracellular signaling pathways in modulation of CD44-mediated phagocytosis. Using biotinylated erythrocytes coated with specific antibodies (anti-CD44–coated erythrocytes [Ebabs]) as the phagocytic prey, we determined that CD44-mediated phagocytosis is reduced by 45% by a blocking CD11b antibody. Further, CD44-mediated phagocytosis was substantially (42%) reduced in CD18-null mice. Immunofluorescence microscopy revealed that CD11b is recruited to the phagocytic cup. The mechanism of integrin activation and mobilization involved activation of the GTPase Rap1. CD44-mediated phagocytosis was also sensitive to the extracellular concentration of the divalent cation Mg2+ but not Ca2+. In addition, buffering of intracellular Ca2+ did not affect CD44-mediated phagocytosis. Taken together, these data suggest that CD44 stimulation induces inside-out activation of CR3 through the GTPase Rap1.

The Studies in Various Murine Strains with Defects in Activation of Complement Cascade (CC) Reveal Both Pivotal and Pleiotropic Role of CC in Mobilization of Hematopoietic Stem/Progenitor Cells.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 774-774
Author(s):  
Marcin Wysoczynski ◽  
Ryan Reca ◽  
Wu Wan ◽  
Magda Kucia ◽  
Marina Botto ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Molecular Level ◽  
Alternative Pathway ◽  
Classical Pathway ◽  
Complement Cascade ◽  
Hematopoietic Stem ◽  
Alternative Pathways ◽  
Dependent Pathway ◽  
Poor Mobilizers

Abstract We reported that complement cascade (CC) becomes activated in bone marrow (BM) during mobilization of hematopoietic stem/progenitor cells (HSPC) by i) immunoglobulin (Ig)-dependent pathway and/or by ii) alternative Ig-independent pathway and, as result of this, iii) several potent bioactive CC anaphylatoxins (C3a, desArgC3a, C5a and desArgC5a) are released (Blood2003;101,3784; Blood2004;103,2071; Blood2005;105,40). To learn more on the role of CC and innate immunity in this process, we compared mobilization in mice that possess defects in CC activation by i) classical pathway (C1q−/−, Ig-deficient), ii) both classical and alternative pathway (C2fB−/−) and in animals iii) that do not generate CC-derived anaphylatoxins (C3−/−, C5−/−). For mobilization, we employed G-CSF and zymosan that activate classical and alternative pathways of CC, respectively. First, we found by ELISA that CC activation in fact correlates with the level of HSPC mobilization. Next, studies in mice deficient in CC activation revealed that CC plays both pivotal and pleiotropic roles in this process. Accordingly, while C1q−/− and C3−/− mice turned out to be easy mobilizers, mobilization was very poor in Ig-deficient, C2fB−/− and C5−/− mice that demonstrate that C3 and C5 cleavage fragments differently control the mobilization of HSPC. To explain this at molecular level, we found that C3 cleavage fragments (C3a, desArgC3a) directly interact with HSPC and increase their responsiveness to SDF-1 gradient and thus prevent uncontrolled egress of HSPC from BM. It explains why C1q−/− and C3−/− mice that do not generate C3 cleavage fragments in BM release easily HSPC into circulation. In contrast, C5 cleavage fragments (C5a, desArgC5a) increase permeability of BM-endothelium and thus are crucial for the egress of HSPC from BM to occur. This explains why mice that do not activate efficient CC such as Ig-deficient, C2fB−/− and C5−/− animals are poor mobilizers. We conclude that the mobilization of HSPC is i) dependent on C activation by the classical or alternative pathway and balanced differently by C3 and C5 cleavage fragments that enhance retention or promote egress of HSPC respectively. Thus, modulation of C activation in BM may help to develop new more efficient strategies for both HSPC mobilization and their homing/engraftment.

scholarly journals Dectin-1 Is A Major β-Glucan Receptor On Macrophages

2002 ◽  
Vol 196 (3) ◽  
pp. 407-412 ◽  
Author(s):  
Gordon D. Brown ◽  
Philip R. Taylor ◽  
Delyth M. Reid ◽  
Janet A. Willment ◽  
David L. Williams ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Drug Design ◽  
Mannose Receptor ◽  
Therapeutic Drug ◽  
Complement Receptor ◽  
Macrophage Mannose Receptor ◽  
Immunomodulatory Properties ◽  
Complement Receptor 3

Zymosan is a β-glucan– and mannan-rich particle that is widely used as a cellular activator for examining the numerous responses effected by phagocytes. The macrophage mannose receptor (MR) and complement receptor 3 (CR3) have historically been considered the major macrophage lectins involved in the nonopsonic recognition of these yeast-derived particles. Using specific carbohydrate inhibitors, we show that a β-glucan receptor, but not the MR, is a predominant receptor involved in this process. Furthermore, nonopsonic zymosan binding was unaffected by genetic CD11b deficiency or a blocking monoclonal antibody (mAb) against CR3, demonstrating that CR3 was not the β-glucan receptor mediating this activity. To address the role of the recently described β-glucan receptor, Dectin-1, we generated a novel anti–Dectin-1 mAb, 2A11. Using this mAb, we show here that Dectin-1 was almost exclusively responsible for the β-glucan–dependent, nonopsonic recognition of zymosan by primary macro-phages. These findings define Dectin-1 as the leukocyte β-glucan receptor, first described over 50 years ago, and resolves the long-standing controversy regarding the identity of this important molecule. Furthermore, these results identify Dectin-1 as a new target for examining the immunomodulatory properties of β-glucans for therapeutic drug design.

scholarly journals Complement Receptor 3 Blockade Promotes IL-12-Mediated Clearance ofPorphyromonas gingivalisand Negates Its Virulence In Vivo

2007 ◽  
Vol 179 (4) ◽  
pp. 2359-2367 ◽  
Author(s):  
George Hajishengallis ◽  
Muhamad-Ali K. Shakhatreh ◽  
Min Wang ◽  
Shuang Liang
Keyword(s):  
Complement Receptor ◽  
Complement Receptor 3

Immunodeficient Mice Are Poor Mobilizers -Novel Evidence That Demonstrates a Pivotal Role of Complement in Triggering Mobilization of HSPC.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1976-1976 ◽  
Author(s):  
Ryan Reca ◽  
Marcin Wysoczynski ◽  
Richard Hansen ◽  
Magda Kucia ◽  
Anna Janowska-Wieczorek ◽  
...  
Keyword(s):  
B Lymphocytes ◽  
Alternative Pathway ◽  
Normal Serum ◽  
Wild Type ◽  
Hematopoietic Stem ◽  
Alternative Pathways ◽  
Immunodeficient Mice ◽  
Natural Igm ◽  
Dependent Pathway

Abstract Complement (C) is activated by immunoglobulin-dependent classical and immunoglobulin-independent alternative pathways, and we recently reported that the activation of C enhances the responsiveness of hematopoietic stem/progenitor cells (HSPC) to an SDF-1 gradient cascade by releasing C3a and desArgC3a cleavage fragments (Blood2003;101:3784; Blood2004; 103: 2071). We also found that bone marrow (BM) concentration of C3a and desArgC3a increases during mobilization with G-CSF or cyclophosphamide (CY). To explain this phenomenon we envisioned that these mobilizing agents expose a neo-antigen in BM tissue by turning BM into a highly proteolytic microenvironment. As a consequence of this, the newly exposed neo-epitope becomes bound by natural IgM antibodies leading to the activation of the classical C cascade. In our studies to elucidate the role of C activation in triggering the mobilization of HSPC, we found that C is effectively activated in the BM of G-CSF- or CY- mobilized wild-type (wt) but not IgM-deficient (RAG2null) mice. More importantly we found that several immunodeficient murine strains (RAG2null, SCID and Xid) displayed severely reduced G-CSF-induced mobilization of HSPC which we believe is a result of lack of B lymphocytes and complement-activating immunoglobulins. Supporting this, we found T cell depletion in wt mice did not affect mobilization. Moreover, G-CSF-induced mobilization in RAG2null, SCID and Xid animals was restored after infusion of murine inmmunoglobulins. Furthermore, since mobilization by zymosan or sulfated glycans activates C via the alternative immunoglobulin-independent pathway, we focused on the role of C activation during zymosan-induced mobilization. As expected, zymosan-induced immunoglobulin-independent C activation and mobilization of HSPC were unaffected in immunodeficient (RAG2null, SCID and Xid) mice which have a normal serum level of C3. However, these processes were severely reduced in C3-deficient animals. Thus our data strongly support the notion that C and innate immunity is an important trigger of mobilization of HSPC. While G-CSF and cyclophosphamide activate C by a classical IgM-dependent pathway, zymosan and sulfated polyglycans activate it by employing an alternative pathway. In conclusion, mobilization of HSPC could be envisioned as part of a more global immune response that is triggered/mediated by C3 activation/cleavage.

Knockout of the PKC Substrate Pleckstrin Causes Pleomorphic Defects in Platelets, Lymphocytes and Granulocytes.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 394-394
Author(s):  
Lurong Lian ◽  
Yanfeng Wang ◽  
Xinsheng Chen ◽  
Tami Bach ◽  
Laurie Lenox ◽  
...  
Keyword(s):  
T Cells ◽  
Platelet Aggregation ◽  
Alternative Pathway ◽  
Second Messengers ◽  
Pi3k Inhibitor ◽  
Loss Of Function ◽  
Wild Type

Abstract Pleckstrin is a 40 kDa phosphoprotein containing amino- and carboxyl-terminal Pleckstrin Homology (PH) domains separated by a DEP domain. Pleckstrin’s expression is restricted to platelets and leukocytes, and represents approximately 1% of total cellular protein within these cells. Following platelet and leukocyte activation, PKC rapidly phosphorylates pleckstrin inducing it to bind membrane bound phospholipids such as phosphatidylinositol 4,5 bisphosphate (PIP2). Heterologously expressed phosphorylated pleckstrin colocalized with integrins and induces cytoskeletal reorganization. To better define the role of pleckstrin in vivo, we introduced a loss-of-function mutation into the murine pleckstrin gene. Pleckstrin-null mice were present in offspring at a frequency consistent with a Mendelian inheritance pattern. Adult pleckstrin −/− mice had 32% lower platelet counts than their littermates, but exhibited no spontaneous hemorrhage. Given the role of PKC and phospholipid second messengers on cytoskeletal dynamics, and our observations of pleckstrin overexpression in cell lines, we analyzed whether loss of pleckstrin affected cell spreading. Pleckstrin −/− platelets spread extremely poorly upon immobilized fibrinogen, and rarely exhibited broad membrane extensions. Granulocytes from pleckstrin −/− mice also have a spreading defect, as well as impaired ability to generate reactive oxygen species in the response to TNFα. Knockout B-cells, CD4-T-cells, and CD8-T-cells all migrated approximately 30% as efficiently as wild type cells in response to a gradient of SDF-1α in a transwell assay. These data suggest that loss of pleckstrin causes cytoskeletal defects in cells of multiple hematopoietic lineages. Analyzing whether this caused a functional defect, we found that pleckstrin −/− platelets exhibited a 22% dense- and 24% alpha-granule exocytosis defect, and a 35% defect in thrombin-induced calcium entry. In spite of these abnormalities, platelets changed shape and aggregated normally after stimulation with thrombin, ADP, or collagen in vitro. Pleckstrin knockout platelets did have a markedly impaired aggregation response following exposure to the PKC stimulant, PMA. This suggested that pleckstrin is a critical effector for PKC-mediated aggregation, but another pathway is able to compensate for this loss of pleckstrin following agonist stimulation. We reasoned that the alternative pathway might also utilize PIP2-dependent second messengers. Since the phosphorylation of PIP2 by PI3K generates second messengers that also contribute to platelet aggregation, we tested whether PI3K compensated for the loss of pleckstrin. We found that the PI3K inhibitor, LY294002 profoundly impaired the aggregation of pleckstrin knockout platelets in response to stimulation of the thrombin receptor. In contrast, the PI3K inhibitor minimally affected wild type platelets. This demonstrates that second messengers generated by PI3K are able to compensate for loss of pleckstrin. This also demonstrates that thrombin-induced platelet aggregation can be mediated by one of two parallel pathways, one involving PKC and pleckstrin, and the other involving PI3K. Together, our results show that pleckstrin is an essential component of PKC-mediated platelet activation and signals directed to the cytoskeleton.

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