Fibronectin enhances Campylobacter fetus interaction with extracellular matrix components and INT 407 cells

2008 ◽  
Vol 54 (1) ◽  
pp. 37-47 ◽  
Author(s):  
L. L. Graham ◽  
T. Friel ◽  
R. L. Woodman
Keyword(s):  
Extracellular Matrix ◽  
Solid Phase ◽  
Cell Surface Hydrophobicity ◽  
Surface Hydrophobicity ◽  
Bacterial Attachment ◽  
Binding Assays ◽  
Rgd Sequence ◽  
Campylobacter Fetus ◽  
Extracellular Matrix Components

Campylobacter fetus is a recognized pathogen of cattle and sheep that can also infect humans. No adhesins specific for C. fetus have to date been identified; however, bacterial attachment is essential to establish an infecting population. Scanning electron microscopy revealed C. fetus attachment to the serosal surface of human colonic biopsy explants, a location consistent with the presence of the extracellular matrix (ECM). To determine whether the ECM mediated C. fetus adherence, 7 C. fetus strains were assessed in a solid-phase binding assay for their ability to bind to immobilized ECM components. Of the ECM components assayed, adherence to fibronectin was noted for all strains. Attachment to ECM components was neither correlated with S-layer expression nor with cell-surface hydrophobicity. Ligand immunoblots, however, identified the S-layer protein as a major site of fibronectin binding, and modified ECM binding assays revealed that soluble fibronectin significantly enhanced the attachment of S-layer-expressing C. fetus strains to other ECM components. Soluble fibronectin also increased C. fetus adherence to INT 407 cells. This adherence was inhibited when INT 407 cells were incubated with synthetic peptides containing an RGD sequence, indicating that integrin receptors were involved in fibronectin-mediated attachment. Together, this data suggests that C. fetus can bind to immobilized fibronectin and use soluble fibronectin to enhance attachment to other ECM components and intestinal epithelial cells. In vivo, fibronectin would promote bacterial adherence, thereby, contributing to the initial interaction of C. fetus with mucosal and submucosal surfaces.

scholarly journals Functional mapping of SPARC: peptides from two distinct Ca+(+)-binding sites modulate cell shape.

1990 ◽  
Vol 111 (6) ◽  
pp. 3065-3076 ◽  
Author(s):  
T F Lane ◽  
E H Sage
Keyword(s):  
Extracellular Matrix ◽  
Cell Shape ◽  
Solid Phase ◽  
Cell Spreading ◽  
Dependent Manner ◽  
Binding Assays ◽  
Extracellular Matrix Components ◽  
Multiple Regions ◽  
Matrix Components ◽  
Inhibitor Of Protein Synthesis

Using synthetic peptides, we have identified two distinct regions of the glycoprotein SPARC (Secreted Protein Acidic and Rich in Cysteine) (osteonectin/BM-40) that inhibit cell spreading. One of these sites also contributes to the affinity of SPARC for extracellular matrix components. Peptides representing subregions of SPARC were synthesized and antipeptide antibodies were produced. Immunoglobulin fractions of sera recognizing an NH2-terminal peptide (designated 1.1) blocked SPARC-mediated anti-spreading activity. Furthermore, when peptides were added to newly plated endothelial cells or fibroblasts, peptide 1.1 and a peptide corresponding to the COOH terminal EF-hand domain (designated 4.2) inhibited cell spreading in a dose-dependent manner. These peptides exhibited anti-spreading activity at concentrations from 0.1 to 1 mM. The ability of peptides 1.1 and 4.2 to modulate cell shape was augmented by an inhibitor of protein synthesis and was blocked by specific antipeptide immunoglobulins. In addition to blocking cell spreading, peptide 4.2 competed for binding of [125I]SPARC and exhibited differential affinity for extracellular matrix molecules in solid-phase binding assays. The binding of peptide 4.2 to matrix components was Ca+(+)-dependent and displayed specificities similar to those of native SPARC. These studies demonstrate that both anti-spreading activity and affinity for collagens are functions of unique regions within the SPARC amino acid sequence. The finding that two separate regions of the SPARC protein contribute to its anti-spreading activity lead us to propose that multiple regions of the protein act in concert to regulate the interactions of cells with their extracellular matrix.

Cell-extracellular matrix interactions under in vivo conditions during interstitial cell migration in Hydra vulgaris

Development ◽  
1994 ◽  
Vol 120 (2) ◽  
pp. 425-432 ◽  
Author(s):  
X. Zhang ◽  
M.P. Sarras
Keyword(s):  
Extracellular Matrix ◽  
Cell Migration ◽  
Synthetic Peptide ◽  
Interstitial Cell ◽  
Extracellular Matrix Components ◽  
Matrix Interactions ◽  
Apical Half ◽  
Matrix Components ◽  
Extracellular Matrix Interactions

Interstitial cell (I-cell) migration in hydra is essential for establishment of the regional cell differentiation pattern in the organism. All previous in vivo studies have indicated that cell migration in hydra is a result of cell-cell interactions and chemotaxic gradients. Recently, in vitro cell adhesion studies indicated that isolated nematocytes could bind to substrata coated with isolated hydra mesoglea, fibronectin and type IV collagen. Under these conditions, nematocytes could be observed to migrate on some of these extracellular matrix components. By modifying previously described hydra grafting techniques, two procedures were developed to test specifically the role of extracellular matrix components during in vivo I-cell migration in hydra. In one approach, the extracellular matrix structure of the apical half of the hydra graft was perturbed using beta-aminopropionitrile and beta-xyloside. In the second approach, grafts were treated with fibronectin, RGDS synthetic peptide and antibody to fibronectin after grafting was performed. In both cases, I-cell migration from the basal half to the apical half of the grafts was quantitatively analyzed. Statistical analysis indicated that beta-aminopropionitrile, fibronectin, RGDS synthetic peptide and antibody to fibronectin all were inhibitory to I-cell migration as compared to their respective controls. beta-xyloside treatment had no effect on interstitial cell migration. These results indicate the potential importance of cell-extracellular matrix interactions during in vivo I-cell migration in hydra.

Interaction of plakophilins with desmoplakin and intermediate filament proteins: an in vitro analysis

2000 ◽  
Vol 113 (13) ◽  
pp. 2471-2483 ◽  
Author(s):  
I. Hofmann ◽  
C. Mertens ◽  
M. Brettel ◽  
V. Nimmrich ◽  
M. Schnolzer ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Intermediate Filament ◽  
Solid Phase ◽  
Specific Interaction ◽  
Binding Assays ◽  
Intermediate Filament Proteins ◽  
Amino Terminal ◽  
Vitro Analysis

Plakophilin 1 and 2 (PKP1, PKP2) are members of the arm-repeat protein family. They are both constitutively expressed in most vertebrate cells, in two splice forms named a and b, and display a remarkable dual location: they occur in the nuclei of cells and, in epithelial cells, at the plasma membrane within the desmosomal plaques. We have shown by solid phase-binding assays that both PKP1a and PKP2a bind to intermediate filament (IF) proteins, in particular to cytokeratins (CKs) from epidermal as well as simple epithelial cells and, to some extent, to vimentin. In line with this we show that recombinant PKP1a binds strongly to IFs assembled in vitro from CKs 8/18, 5/14, vimentin or desmin and integrates them into thick (up to 120 nm in diameter) IF bundles extending for several microm. The basic amino-terminal, non-arm-repeat domain of PKP1a is necessary and sufficient for this specific interaction as shown by blot overlay and centrifugation experiments. In particular, the binding of PKP1a to IF proteins is saturable at an approximately equimolar ratio. In extracts from HaCaT cells, distinct soluble complexes containing PKP1a and desmoplakin I (DPI) have been identified by co-immunoprecipitation and sucrose density fractionation. The significance of these interactions of PKP1a with IF proteins on the one hand and desmoplakin on the other is discussed in relation to the fact that PKP1a is not bound - and does not bind - to extended IFs in vivo. We postulate that (1) effective cellular regulatory mechanisms exist that prevent plakophilins from unscheduled IF-binding, and (2) specific desmoplakin interactions with either PKP1, PKP2 or PKP3, or combinations thereof, are involved in the selective recruitment of plakophilins to the desmosomal plaques.

scholarly journals Fibulin-1 mediates platelet adhesion via a bridge of fibrinogen

Blood ◽  
1996 ◽  
Vol 88 (7) ◽  
pp. 2569-2577 ◽  
Author(s):  
S Godyna ◽  
M Diaz-Ricart ◽  
WS Argraves
Keyword(s):  
Extracellular Matrix ◽  
Vascular Injury ◽  
Whole Blood ◽  
Platelet Adhesion ◽  
Solid Phase ◽  
Blood Perfusion ◽  
General Mechanism ◽  
Binding Assays ◽  
Solid Phase Binding ◽  
Coated Surfaces

Fibulin-1 is a component of the extracellular matrix that surrounds vascular smooth muscle. This observation, along with the recent finding that fibulin-1 can bind fibrinogen (J Biol Chem 270:19458, 1995), prompted investigation into the potential role of fibulin-1 as a thrombogenic agent. In perfusion chamber assays, platelets in whole blood under flow conditions attached and spread on surfaces coated with fibulin-1. This adhesion was completely blocked by fibulin-1 antibodies. Platelets free of plasma did not attach to fibulin-1 coated surfaces; however, with the addition of fibrinogen, platelet adhesion to fibulin-1 took place. When detergent extracts of platelets were subjected to fibulin-1-Sepharose affinity chromatography, the integrin alpha IIb beta 3 was selected. Solid phase binding assays using purified components showed that integrin alpha IIb beta 3 could not bind directly to fibulin-1 but in the presence of fibrinogen the integrin bound to fibulin-1-coated surfaces. Monoclonal alpha IIb beta 3 antibodies capable of blocking its interaction with fibrinogen completely blocked platelet adhesion to fibulin-1 in both whole blood perfusion and static adhesion assays. The results show that fibulin-1 can support platelet attachment via a bridge of fibrinogen to the platelet integrin alpha IIb beta 3. When fibroblast monolayers containing extracellular matrix-incorporated fibulin-1 were used as adhesion substrates, platelet adhesion in the presence of fibrinogen could be inhibited by 30% using antibodies to fibulin-1. Following vascular injury, fibulin-1 present in the extracellular matrix of the vessel wall may therefore interact with plasma fibrinogen and promote platelet adhesion, leading to the formation of a platelet plug. Thus, fibulin-1 joins the list of matrix proteins including collagens I and IV and fibronectin that mediate platelet adhesion via a plasma protein bridge. This bridging phenomenon may represent a general mechanism by which platelets interact with exposed subendothelial matrices following vascular injury.

scholarly journals Bartonella henselae Pap31, an Extracellular Matrix Adhesin, Binds the Fibronectin Repeat III13 Module

2006 ◽  
Vol 74 (5) ◽  
pp. 2513-2521 ◽  
Author(s):  
S. M. Dabo ◽  
A. W. Confer ◽  
B. E. Anderson ◽  
Snehalata Gupta
Keyword(s):  
Extracellular Matrix ◽  
Binding Site ◽  
Binding Protein ◽  
Umbilical Vein ◽  
Bartonella Henselae ◽  
Bacterial Attachment ◽  
Dependent Manner ◽  
Host Interaction ◽  
Cloned Gene

ABSTRACT Bartonella henselae wound-associated infections suggest involvement of extracellular matrix molecules in adhesion and invasion. Pap31 was previously identified as a hemin-binding protein. Our recent studies suggest the protein is an adhesin that is recognized by the host's immune systems. In this study we examined the interactions of B. henselae Pap31 with fibronectin (Fn), heparin (Hep), and human umbilical vein endothelial cells (HUVECs). The cloned gene was expressed in Escherichia coli, and the purified Pap31 protein elicited strong antibody responses in mice and was reactive with rabbit anti-live B. henselae and mouse anti-Pap31 antibodies by Western blotting. Pap31 bound to immobilized Fn and to HUVECs in a dose-dependent manner and to Hep. Fn fragment-binding assays identified the Hep-1 and Hep-2 binding domains of human Fn and in particular the 12-13FnIII repeat module as primary binding sites for this adhesin. Furthermore, Pap31 binding to the above Fn fragments could be inhibited by Hep, suggesting a common binding site involving the 13FnIII repeat module on the Hep-2 domain of Fn. Adherence of intact B. henselae to HUVECs was inhibited by increasing concentrations of anti-Pap31 antibodies. In addition, purified Pap31 coprecipitated effectively with Fn and anti-Fn antibodies. Taken together, these data suggest that Pap31 is an Fn-binding protein mediating the B. henselae-host interaction(s), and they implicate the 13FnIII repeat module as an important binding site for this adhesin on the Fn molecule. These interactions may be important initial steps leading to bacterial attachment and colonization that promote the establishment of B. henselae infections in vivo.

scholarly journals Agrin-induced reorganization of extracellular matrix components on cultured myotubes: relationship to AChR aggregation.

1990 ◽  
Vol 111 (3) ◽  
pp. 1161-1170 ◽  
Author(s):  
R M Nitkin ◽  
T C Rothschild
Keyword(s):  
Extracellular Matrix ◽  
Type Iv Collagen ◽  
Neuromuscular Junctions ◽  
Synaptic Organization ◽  
Type Iv ◽  
Extracellular Matrix Components ◽  
Cultured Myotubes ◽  
Matrix Components ◽  
Induced Aggregation

Agrin, an extracellular matrix-associated protein extracted from synapse-rich tissues, induces the accumulation of acetylcholine receptors (AChRs) and other synaptic components into discrete patches on cultured myotubes. The appearance of agrin-like molecules at neuromuscular junctions suggests that it may direct synaptic organization in vivo. In the present study we examined the role of extracellular matrix components in agrin-induced differentiation. We used immunohistochemical techniques to visualize the spatial and temporal distribution of laminin, a heparan sulfate proteoglycan (HSPG), fibronectin, and type IV collagen on cultured chick myotubes during agrin-induced aggregation of AChRs. Myotubes displayed significant amounts of laminin and HSPG, lesser amounts of type IV collagen, and little, if any, fibronectin. Agrin treatment caused cell surface laminin and HSPG to patch, while collagen and fibronectin distributions were generally unaffected. Many of the agrin-induced laminin and HSPG patches colocalized with AChR patches, raising the possibility of a causal relationship between matrix patching and AChR accumulations. However, patching of AChRs (complete within a few hours) preceded that of laminin or HSPG (not complete until 15-20 h), making it unlikely that matrix accumulations initiate AChR patching at agrin-induced sites. Conversely, when AChR patching was blocked by treatment with anti-AChR antibody mAb 35, agrin was still able to effect patching of laminin and HSPG. Taken together, these findings suggest that agrin-induced accumulations of AChR and laminin/HSPG are not mechanistically linked.

New insights into the structural elements involved in the skin haemorrhage induced by snake venom metalloproteinases

2010 ◽  
Vol 104 (09) ◽  
pp. 485-497 ◽  
Author(s):  
Ana Oliveira ◽  
Adriana Paes Leme ◽  
Amanda Asega ◽  
Antonio Camargo ◽  
Jay Fox ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Snake Venom ◽  
Solid Phase ◽  
Structural Elements ◽  
Binding Assays ◽  
Domain Organisation ◽  
Solid Phase Binding ◽  
A Minor ◽  
Von Willebrand

SummaryHaemorrhage induced by snake venom metalloproteinases (SVMPs) is a complex phenomenon resulting in capillary disruption and extravasation. This study analysed structural elements important for the interaction of four Bothrops jararaca SVMPs of different domain organisation and glycosylation levels with plasma and extracellular matrix proteins: HF3 (P-III class) is highly glycosylated and ~80 times more haemorrhagic than bothropasin (P-III class), which has a minor carbohydrate moiety; BJ-PI (P-I class) is not haemorrhagic and the DC protein is composed of disintegrin-like/cysteine-rich domains of bothropasin. HF3, bothropasin and BJ-PI showed different degradation profiles of fibrinogen, fibronectin, vitronectin, von Willebrand factor, collagens IV and VI, laminin and Matrigel™; however, only bothropasin degraded collagen I. In solid-phase binding assays HF3 and bothropasin interacted with fibrinogen, fibronectin, laminin, collagens I and VI; the DC protein bound only to collagens I and VI; however, no binding of BJ-PI to these proteins was detected. N-deglycosylation caused loss of structural stability of bothropasin and BJ-PI but HF3 remained intact, although its haemorrhagic and fibrinogenolytic activities were partially impaired. Nevertheless, N-deglycosylated HF3 bound with higher affinity to collagens I and VI, although its proteolytic activity upon these collagens was not enhanced. This study demonstrates that features of carbohydrate moieties of haemorrhagic SVMPs may play a role in their interaction with substrates of the extracellular matrix, and the ability of SVMPs to degrade proteins in vitro does not correlate to their ability to cause haemorrhage, suggesting that novel, systemic approaches are necessary for understanding the mechanism of haemorrhage generation by SVMPs.

Surface hydrophobicity of acidophilic heterotrophic bacterial cells in relation to their adhesion on minerals

1991 ◽  
Vol 37 (9) ◽  
pp. 692-696 ◽  
Author(s):  
B. K. Chakrabarti ◽  
P. C. Banerjee
Keyword(s):  
Heterotrophic Bacteria ◽  
Ph Value ◽  
Manganese Nodule ◽  
Cell Surface Hydrophobicity ◽  
Surface Hydrophobicity ◽  
Bacterial Attachment ◽  
Bacterial Cells ◽  
Iron Pyrite ◽  
Acidophilic Bacteria ◽  
Hydrophilic Nature

The cell-surface hydrophobicity of acidophilic heterotrophic bacteria originating from mines varied with the pH of the suspending medium and with the growth temperature. Adhesion of these bacterial cells on mineral particles depended upon the hydrophobic (or hydrophilic) nature of both the cells and the minerals. A strong correlation between these properties was usually observed at different pH values of the suspending medium. At a certain pH value, bacterial attachment depended upon the particle size of the minerals. Key words: hydrophobicity, acidophilic bacteria, Acidiphilium cryptum, Acidiphilium symbioticum, adhesion, manganese nodule, chalcopyrite, iron pyrite.

scholarly journals Laminin promotes neuritic regeneration from cultured peripheral and central neurons.

1983 ◽  
Vol 97 (6) ◽  
pp. 1882-1890 ◽  
Author(s):  
M Manthorpe ◽  
E Engvall ◽  
E Ruoslahti ◽  
F M Longo ◽  
G E Davis ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Neurite Growth ◽  
Sensory Ganglia ◽  
Culture Methods ◽  
Fetal Rat ◽  
Extracellular Matrix Components ◽  
Ciliary Ganglion Neurons ◽  
Ganglion Neurons

The ability of axons to grow through tissue in vivo during development or regeneration may be regulated by the availability of specific neurite-promoting macromolecules located within the extracellular matrix. We have used tissue culture methods to examine the relative ability of various extracellular matrix components to elicit neurite outgrowth from dissociated chick embryo parasympathetic (ciliary ganglion) neurons in serum-free monolayer culture. Purified laminin from both mouse and rat sources, as well as a partially purified polyornithine-binding neurite promoting factor (PNPF-1) from rat Schwannoma cells all stimulate neurite production from these neurons. Laminin and PNPF-1 are also potent stimulators of neurite growth from cultured neurons obtained from other peripheral as well as central neural tissues, specifically avian sympathetic and sensory ganglia and spinal cord, optic tectum, neural retina, and telencephalon, as well as from sensory ganglia of the neonatal mouse and hippocampal, septal, and striatal tissues of the fetal rat. A quantitative in vitro bioassay method using ciliary neurons was used to (a) measure and compare the specific neurite-promoting activities of these agents, (b) confirm that during the purification of laminin, the neurite-promoting activity co-purifies with the laminin protein, and (c) compare the influences of antilaminin antibodies on the neurite-promoting activity of laminin and PNPF-1. We conclude that laminin and PNPF-1 are distinct macromolecules capable of expressing their neurite-promoting activities even when presented in nanogram amounts. This neurite-promoting bioassay currently represents the most sensitive test for the biological activity of laminin.

scholarly journals Regulation of Tissue Fibrosis by the Biomechanical Environment

2013 ◽  
Vol 2013 ◽  
pp. 1-10 ◽  
Author(s):  
Wayne Carver ◽  
Edie C. Goldsmith
Keyword(s):  
Extracellular Matrix ◽  
Intracellular Signaling ◽  
Mechanical Load ◽  
Mechanical Force ◽  
Mechanical Signals ◽  
Organ Systems ◽  
Extracellular Matrix Components ◽  
Matrix Components

The biomechanical environment plays a fundamental role in embryonic development, tissue maintenance, and pathogenesis. Mechanical forces play particularly important roles in the regulation of connective tissues including not only bone and cartilage but also the interstitial tissues of most organs.In vivostudies have correlated changes in mechanical load to modulation of the extracellular matrix and have indicated that increased mechanical force contributes to the enhanced expression and deposition of extracellular matrix components or fibrosis. Pathological fibrosis contributes to dysfunction of many organ systems. A variety ofin vitromodels have been utilized to evaluate the effects of mechanical force on extracellular matrix-producing cells. In general, application of mechanical stretch, fluid flow, and compression results in increased expression of extracellular matrix components. More recent studies have indicated that tissue rigidity also provides profibrotic signals to cells. The mechanisms whereby cells detect mechanical signals and transduce them into biochemical responses have received considerable attention. Cell surface receptors for extracellular matrix components and intracellular signaling pathways are instrumental in the mechanotransduction process. Understanding how mechanical signals are transmitted from the microenvironment will identify novel therapeutic targets for fibrosis and other pathological conditions.

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