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At least 1,531 records · Page 85Linked to original sources
PubMed1993

Interleukin 1-induced cancer cell/endothelial cell adhesion in vitro and its relationship to metastasis in vivo: role of vessel wall 13-HODE synthesis and integrin expression.

Previously, we have demonstrated that stimulation of endothelial cells (ECs) with interleukin-1 alpha (IL-1 alpha) enhances the synthesis and expression of the vitronectin receptor (VnR), promotes VnR-dependent adhesion of human A549 adenocarcinoma cells to ECs, and is associated with decreased EC 13-hydroxyoctadecadienoic acid (13-HODE) synthesis in vitro. To determine whether these observations are relevant in vivo, we examined the acute retention and subsequent metastasis of intravenously-injected B16F10 melanoma cells in murine lungs, in relation to vessel wall 13-HODE. In C57BL/6 mice pretreated with IL-1 alpha, vessel wall 13-HODE was decreased and B16F10 lung entrapment and metastasis were increased. The latter two events were blocked by pretreating the animals with the GRGDS peptide. These data suggest a relationship between vessel wall 13-HODE synthesis, adhesion molecule expression, and adhesion of B16F10 cells to the endothelium.

Animals↗
PubMed1992

Localization of the cytoadhesin integrins in the human cornea. An immunohistochemical study using monoclonal antibodies.

Cell-matrix interactions play a fundamental role in normal and pathological conditions. They can be mediated by the cytoadhesin subgroup of the integrin superfamily of adhesion molecules. Its members include the vitronectin receptor (VNR) and the platelet glycoprotein IIb/IIIa (GP IIb/IIIa). Both receptors are composed of an alpha-chain (alpha v and alpha IIb, respectively) coupled to a beta 3-chain. Using in situ immunohistochemistry and monoclonal antibodies, the authors studied the distribution of GP IIIa (common beta 3-chain), GP IIb/IIIa (alpha IIb-chain) and VNR (alpha v-chain) in normal and pathological corneal tissues. In the normal cornea, the limbal vascular endothelium was weakly alpha v-positive. Occasionally, faint and granular staining was seen in the epithelium. In the pathological samples, an upregulated expression of the alpha v-chain was noticed on the corneal epithelium as well as on fibroblasts and corneal endothelium. The alpha IIb and beta 3-chains were consistently absent. These data suggest that expression of the VNR-alpha v-chain in the human cornea is modulated by soluble factors released during inflammation and wound healing. Dissociation of expression of the alpha v and beta 3-chains suggests usage of an alternative beta-chain by the VNR-alpha v-chain.

Antibodies, Monoclonal↗
PubMed1994

An immunohistochemical study of the extracellular matrix in oral squamous cell carcinoma and its association with invasive and metastatic potential.

The expression of extracellular matrices (ECMs) laminin (LN), type IV collagen (IV C), heparan-sulphate proteoglycan (HS-PG), fibronectin (FN), tenascin (TN), decorin and vitronectin (VN) was examined immunohistochemically in 112 primary tumours and 29 metastatic cervical lymph nodes in oral squamous cell carcinoma (OSCC). In highly invasive primary tumours, the expression of LN, IV C and HS-PG in the basement membrane along the tumour-stroma borderline and the expression of decorin and VN in the tumour stroma at the invasive site were all significantly decreased. The expression of FN and TN in the tumour stroma at the same site was markedly increased. In peritumour stroma in metastatic lymph nodes, LN, IV C, HS-PG, decorin and VN were weakly expressed, while FN and TN were strongly expressed. Thus, the staining pattern of the ECMs in the metastatic lymph nodes was similar to that in highly invasive primary tumours. Furthermore, in primary tumours of metastatic cases, the expression of LN, IV C, HS-PG, decorin and VN obviously decreased, while the expression of FN and TN increased when compared with those of the non-metastatic cases. The investigation of ECMs in OSCC was valuable in predicting tumour behaviour.

Carcinoma, Squamous Cell↗
PubMed1996

Lily cofactor-independent phosphoglycerate mutase: purification, partial sequencing, and immunolocalization.

A cofactor-independent phosphoglycerate mutase (PGAM-i) was isolated to homogeneity from monocotyledonous Lilium longiflorum Thunb. Two-dimensional (2D) polyacrylamide gel electrophoresis resolved three PGAM-i forms. This enzyme was originally identified by cross-reactivity to antibodies affinity-purified from 2D gels using human vitronectin (VN). Antibody produced against a denatured protein spot from a 2D gel did not recognize VN protein, but partial protein and DNA sequencing showed similarity of the former protein to maize PGAM-i. Immunoblots from roots, styles, leaves, and anthers showed the presence of PGAM-i in all tissues examined; it was isolated predominantly from the soluble cell fraction, with some present in the insoluble cell fraction. Immunoelectron microscopy demonstrated its localization in the cytoplasm and plastids in root cells near the apical meristem. In addition, immunogold labeling detected signals from the nucleus. The immunohistochemical localization of the enzyme in the nucleus, as well as in the cytosol and plastids, indicates that lily PGAM-i might have multiple functions in the cell.

Amino Acid Sequence↗
PubMed1991

Activation of blood coagulation and fibrinolysis in vibration syndrome.

The pathophysiology of peripheral circulatory disturbance in patients presenting with vibration syndrome was studied from the viewpoint of blood coagulation. Plasma levels of fibronectin (FN), vitronectin (VN), thrombin-antithrombin III complex (TAT), and alpha 2-plasmin inhibitor-plasmin complex (PIC) were measured in 23 subjects who showed no evidence of vibration-induced white finger [VWF(-) group] and in 24 patients who presented with VWF [VWF(+) group]. In the VWF(-) group, plasma FN concentrations were elevated but plasma TAT and PIC levels were within the normal ranges. In the VWF(+) group, plasma FN concentrations were normal but plasma TAT and PIC levels were significantly elevated. In both groups, plasma VN concentrations were similar to those in normal controls. For purposes of comparison, 32 patients presenting with diabetes mellitus were also studied. They were divided into 2 groups, 13 subjects who showed no evidence of angiopathy [complication(-) group] and 19 patients who presented with angiopathy [complication(+) group]. In the complication(+) group, plasma TAT and PIC concentrations were significantly elevated, as in the VWF(+) group. These results suggest that in vibration syndrome, vibration, cold stimulus, or other factors first injure the vascular endothelium, resulting in a rise in plasma FN, and that in the VWF(+) group, augmentation of coagulation and fibrinolysis induces a state of compensated disseminated intravascular coagulation (DIC).

Adult↗
PubMed1995

Flagellum-mediated adhesion of Trypanosoma congolense to bovine aorta endothelial cells.

We studied the interaction between Trypanosoma congolense and bovine aorta endothelial (BAE) cell monolayers. Our findings suggest that trypanosomes adhere predominantly to the flattened, peripheral cell surface domains as well as to filamentous endothelial outgrowths that are present during in vitro cultivation in non-confluent monolayers. Adhesion is mediated exclusively by the flagellum in a distinct geometrical order with respect to the flagellar cytoskeleton. Thus, it is possible to define exactly the trypanosomal cell surface domain involved in the attachment process. After 24-48 h of cultivation on monolayers, trypanosomes start to develop short, filopodia-like flagellar protrusions, which serve as additional elements in assisting parasite attachment. Small filaments (3-5 nm) also serve as cross-links between flagellar and endothelial cell surface membranes. Lectin-gold labeling shows that these cross-links contain sialic acid residues. In vitro assays confirm that sialic acid is involved in the adhesion process, whereas the extracellular matrix (ECM) proteins fibronectin, collagen, laminin and vitronectin are not. The presence of T. congolense exhibits a mitogenic effect on BAE cells.

Adhesiveness↗
PubMed1992

Adherence of pathogenic and non-pathogenic Entamoeba histolytica strains to neutrophils.

The adherence of polymorphonuclear leukocytes (PMNs) to eight pathogenic and nine nonpathogenic strains of Entamoeba histolytica was examined. No difference between pathogenic and nonpathogenic strains was found. The addition of different carbohydrates confirmed the importance of the 170-kDa lectin of E. histolytica in binding to PMNs, corroborated by the finding that treatment of PMNs with galactosidase inhibited adherence. Inhibition of the microfilament system of E. histolytica using cytochalasin B resulted in a loss of adherence to PMNs. Inhibition of the microtubule system using nocodazole did not affect adherence. Preincubation of the trophozoites with serum resulted in enhanced adherence, but the serum factor responsible for this effect could not be identified. Fibronectin, vitronectin, integrins (CD11/CD18 molecules), complement, and mannose-binding protein did not seem to mediate adherence between E. histolytica and PMNs. In summary, these results indicate that defective adherence mechanisms are not a common feature of nonpathogenic E. histolytica strains.

Acetylgalactosamine↗
PubMed1988

A serum-free primary culture system for studying cell-substrate interactions during newt epidermal cell migration.

To study the interaction of migrating newt epidermal cells with purified extracellular matrix (ECM) molecules we have developed an in vitro migration assay using pieces of newt skin explanted onto culture dishes coated with various ECM molecules and cultured for 18 h in defined serum-free medium. Newt epidermal cells migrate out from explants placed on dishes coated with either collagen, vitronectin, fibronectin, or fibrinogen but not on albumin-coated or uncoated dishes. Explant outgrowth on collagen was best in CEM 2000 medium diluted to 60% of mammalian osmolarity. Other media such as RPMI 1640 or Ex-Cell 300, diluted similarly, may also be used although in our hands CEM 2000 always allowed more migration. We found no migration on collagen when skin explants were incubated in Holtfreter's solution (an amphibian saline solution that we have previously shown allows reepithelialization on amputated newt limbs). Supplementation of Holtfreter's solution with glucose did not improve its ability to support migration. By testing various supplement combinations in conjunction with CEM 2000 and RPMI 1640 we found that neither serum, insulin, selenium, transferrin, nor L-glutamine is required for explant outgrowth. Of the additives tested, outgrowth was stimulated only by insulin. Epidermal cell outgrowth on collagen was inhibited by both puromycin and cycloheximide, indicating the necessity for protein synthesis in this system. Whether the effects of these protein synthesis inhibitors are specifically on migration-related events or on general metabolic requirements is not clear. Inasmuch as there was no correlation (r = -0.227) between DNA synthesis (measured by incorporation of tritiated thymidine) and the amount of outgrowth, we believe that our assay is a measure of cell migration alone rather than a combination of mitosis and migration. This explant outgrowth system represents a new and relatively simple assay that can be used in the study of cell-substrate interactions during newt epidermal cell migration over extracellular matrix molecules in a defined serum-free environment.

Animals↗
PubMed2003

Plasminogen activation in neurofibromatosis 2-associated and sporadic schwannomas.

BACKGROUND: Schwannomas are usually benign tumours which occur sporadically or in association with neurofibromatosis 2 (NF2), an autosomal dominant disorder. Invasiveness and higher proliferative potential compared to sporadic tumours are features of NF2-associated schwannomas. METHOD: We studied urokinase (uPA), tissue-type plasminogen activator (tPA), urokinase receptor (uPAR) and plasminogen activator inhibitor-1 (PAI-1) expression by in situ hybridization and by immunohistochemistry in 14 NF2 and 15 sporadic patients with 34 schwannomas. uPAR and vitronectin immunohistochemistry were also studied. Three sural nerve specimens were included as Schwann cell controls. FINDINGS: Both schwannoma groups expressed prominent levels of uPA and tPA. Semiquantitative analysis of the in situ hybridization and immunoreactivity demonstrated that NF2 schwannomas expressed less PAI-1 at the mRNA level than sporadic schwannomas (score 1.63+/-0.41 vs. 2.05+/-0.75) and less total PAI-1 at the antigen level (score 1.55+/-0.66 vs. 2.07+/-0.56). PAI-1 was mostly in a free form in NF2 schwannomas compared to the sporadic counterparts (score 1.85+/-0.73 vs. 1.46+/-0.58), whereas there was less uPAR antigen in NF2 schwannomas than in the sporadic counterparts (score 1.18+/-0.49 vs. 1.68+/-0.56). Sural nerve Schwann cells did not express detectable level of PAI-1 and at the most a minor amount of tPA. CONCLUSIONS: Schwann cells of tumour cell origin, both in sporadic and NF2 schwannomas, expressed elevated levels of plasminogen activators and PAI-1 compared to normal suralic nerve Schwann cells. Furthermore, there seemed to be an imbalance in the PA-PAI-1 system in NF2-associated schwannomas. Although our methods are more descriptive than quantitative, we suggest that the somewhat more aggressive behavior of NF2-associated schwannomas compared to sporadic schwannomas may be based on the local proteolytic activity.

Antibodies, Monoclonal↗
PubMed2001

Establishment and characterization of macrophage-like cell lines expressing osteoclast-specific markers.

Osteoclasts differentiate from hematopoietic precursors of the monocyte/macrophage lineage to mononuclear preosteoclasts and multinuclear mature osteoclasts. In the present study, we report on the establishment of macrophage like cell lines from mouse bone marrow by coculturing bone marrow cells with mouse chondrocytes. Isolated clones (MLC-6 and MLC-7 cells) expressed fully differentiated osteoclast markers, such as calcitonin receptors, vitronectin receptors, tartrate-resistant acid phosphatase and vacuolar H+ -ATPase, in the absence of osteoclast differentiation factor/osteoprotegerin ligand/RANKL/TRANCE, which was essential for osteoclast differentiation. Most clones also maintained expression of a macrophage-associated protein, namely non-specific esterase. Both MLC-6 and MLC-7 cells released cathepsin K into the culture medium. Both clones resolved dentine slices when cocultured with the osteoblast cell line ST2. The cloned cell lines are considered to be useful tools in the study of osteoclast differentiation.

Animals↗
PubMed2004

Breast cancer cells with inhibition of p38alpha have decreased MMP-9 activity and exhibit decreased bone metastasis in mice.

p38 belongs to a family of mitogen-activated protein kinases, which transfer extracellular signals into intracellular responses. p38 is also frequently detected in clinical breast cancer specimens, but its role as a prognostic factor is not known. Of the various p38 isoforms, p38alpha has been shown to mediate the in vitro invasiveness of breast cancer cells through up-regulation of urokinase plasminogen activator (uPA). We studied the role of p38alpha in breast cancer bone metastases, using dominant negative blockade approach. Human MDA-MB-231 breast cancer clones stably expressing dominant negative p38alpha (p38/AF) exhibited decreased basal MMP-9 activity. TGF-beta1-induced MMP-9 activity was also blunted in these clones, as compared with controls in which TGF-betal up-regulated MMP-9 activity. Consistent with these findings, SB202190, a specific p38 inhibitor, also inhibited TGF-beta1-induced MMP-9 activity in parental cells. The p38/AF clones exhibited also reduced uPA production after growth on vitronectin and decreased cell motility, as compared with controls. VEGF production levels in all the studied clones were similar. The p38/AF clone, which had similar in vitro growth rate as the control pcDNA3 clone, formed significantly less bone metastases in a mouse model, as compared with the control clone. In conclusion, inhibition of the p38alpha pathway results in decreased MMP-9 activity, impaired uPA expression and decreased motility, all of which may contribute to the decreased formation of bone metastasis.

Animals↗
PubMed1993

Fibrinolytic components in fetal membranes and amniotic fluid.

OBJECTIVE: We evaluated fibrinolytic components in plasma and amniotic fluid of pregnant women and in postpartum fetal membranes. STUDY DESIGN: Fibrinolytic parameters in amniotic fluid and plasma were measured by means of enzyme-linked immunosorbent assays. Fetal membranes collected after spontaneous labor at term were analyzed by immunohistochemical methods with immunospecific antibodies against fibrinolytic components. RESULTS: Amniotic fluid contained high plasminogen activator inhibitor-1 concentrations but had low activity. Strong staining for plasminogen activator inhibitor-1 and vitronectin was observed in chorionic trophoblasts and moderate staining in decidual connective tissue. Strong staining for plasminogen activator inhibitor-2 was seen in decidual cells. Although prominent staining of plasminogen activators and plasminogen were observed in the amniotic epithelium, virtually no plasminogen activator inhibitor-1, plasminogen activator inhibitor-2, or alpha 2-plasmin inhibitor staining was detected. CONCLUSION: The delicate balance of fibrinolytic activators and inhibitors in fetal membranes and amniotic fluid may contribute to the triggering of membrane rupture at term.

Amniotic Fluid↗
PubMed1990

Biotinylated antibodies bound to streptavidin beads: a versatile solid matrix for immunoassays.

Streptavidin was covalently bound to commercially available polyacrylamide beads (3-10 microns diameter) by peptide bond formation between the carboxyl groups on the solid matrix and the amino groups of the soluble protein. Biotinylated antibody or lectin was linked to the polyacrylamide beads via the streptavidin molecules. Immunoassays for human IgA1, IgA2, IgE, and vitronectin were developed utilizing the antibody or lectin as a capture ligand. The protein being assayed was quantitated colorimetrically at 492 nm via horseradish peroxidase-conjugated antibodies.

Acrylic Resins↗
PubMed1996

Centrifugal pump and heparin coating improves cardiopulmonary bypass biocompatibility.

BACKGROUND: Centrifugal pumps are being used increasingly for short-term extracorporeal circulation purposes such as during heart operations. Whether the centrifugal pump improves the cardiopulmonary bypass biocompatibility has not been fully documented. METHODS: A roller pump (n = 20) was compared in vivo with a centrifugal pump (n = 20) in groups of patients in which cardiopulmonary bypass circuits that were either totally heparin coated (Carmeda BioActive Surface; n = 20) or uncoated (n = 20) were used. We expected the heparin coating to attenuate blood activation, thus possibly making the comparison of the two pumps easier with respect to their different blood activation potentials. Samples of blood plasma, obtained during cardiopulmonary bypass from low-risk coronary artery bypass grafting patients, were analyzed for hemolysis (plasma haemoglobin), complement activation (C3bc and the terminal complement complex), a complement lytic inhibitor (vitronectin), coagulation activation (fibrinopeptide A), granulocyte activation (lactoferrin), and platelet activation (beta-thromboglobulin). RESULTS: The concentrations of terminal complement complex, lactoferrin, and beta-thromboglobulin were significantly lower in association with heparin-coated surfaces. The concentration of plasma hemoglobin was significantly lower in association with the centrifugal pump. In uncoated circuits, the beta-thromboglobulin level was significantly higher in association with the roller pump than with the centrifugal pump, but this significant reduction in the beta-thromboglobulin level did not hold true for the heparin-coated circuit group. CONCLUSIONS: A heparin-coated cardiopulmonary bypass surface reduces the blood activation potential during cardiopulmonary bypass, and the centrifugal pump causes less hemolysis than the roller pump.

Adult↗
PubMed1991

Effect of cyclic RGD peptide on cell adhesion and tumor metastasis.

Several kinds of cyclic peptides containing an L-arginine-glycine-L-aspartic acid RGD sequence were synthesized by the liquid phase method, and we investigated their effects on the attachment of mouse B16 melanoma cells onto fibronectin-coated well. Cyclo (GRGDSPA) inhibited the cell attachment at a 20-fold lower concentration than the linear form. The cell adhesion was inhibited by the synthetic peptides with the following relative order of activity: cyclo (GRGDSPA) much greater than cyclo (GRGD) greater than cyclo (RGDS), cyclo (GRGDSP) greater than cyclo (GRGDS) greater than cyclo (RGDSP), cyclo (RGDSPA). Cyclo (GRGDSPA) was more effective at inhibiting cell attachment to vitronectin than it was at competing with fibronectin attachment, as reported in the case of GRGDSP. Moreover, cyclo (GRGDSPA) significantly reduced the formation of colonies in mice injected with B16-FE7 melanoma cells.

Amino Acid Sequence↗
PubMed1987

Immobilized serotonin: a novel substrate for cell culture.

Baby hamster kidney cells, bovine aortal endothelial cells, bovine smooth muscle cells, and chick embryo fibroblasts were all observed to attach and grow on serotonin which had been immobilized by covalent coupling to agarose beads. While growth and morphology of cells on immobilized serotonin appeared normal, a change in cell function may have occurred since the pattern of polypeptides expressed by these cells was different from that of cells grown on two other substrates: immobilized fibronectin and tissue culture plastic. By changing the composition of the fetal calf serum proteins in the growth medium it was shown that cells attach directly to immobilized fibronectin without mediation by medium components. In contrast, cells were found not to attach directly to immobilized serotonin but to attach indirectly via factors absorbed onto immobilized serotonin from fetal calf serum. The major component of this cell attachment activity was shown not to be fibronectin and was identified following separation by SDS-PAGE, electroblotting, and cell binding on nitrocellulose filters. The cell attachment activity compromises a major protein species of Mr 70,000 which is the molecular size of the recently identified serum spreading factor also called vitronectin.

Adsorption↗
PubMed1991

Multiple integrins share the ability to induce elevation of intracellular pH.

Previous work has shown that adhesion of anchorage-dependent cells to fibronectin via integrin alpha 5 beta 1 leads to activation of the Na-H antiporter and a rise in intracellular pH (pHi). We now show that adhesion of bovine capillary endothelial cells (BCE) to fibrinogen; collagens type III, IV, and V; laminin; and vitronectin; ligands that bind other members of the integrin family, resulted in significant elevations in pHi. Other ligands (basic fibroblast growth factor, concanavalin A, and thrombin), which bind cells when immobilized on plastic, but that do not bind integrins and do not support cell growth, do not elevate pHi. Adhesion to an antibody against integrin alpha v beta 3 also elevates pHi. Adhesion of peripheral human T lymphocytes to an antibody against the integrin LFA-1 induced a rise in pHi. Antibodies to CD2 or ICAM-2 had only slight effects on pHi, whereas an antibody to the T cell receptor complex that strongly activates T cells induced a large increase in pHi. We conclude that elevation of pHi by integrins is specific and is a property shared by many members of the integrin family.

Animals↗
PubMed1991

Cytoskeletal reorganizations in human umbilical vein endothelial cells as a result of cytokine exposure.

Treatment of HUVECs in culture with several cytokines and phorbol esters caused reorganizations of the actin and microtubule networks, as well as a redistribution of focal contract proteins. However, expression of the cytoskeletal proteins which link cells, via integrins, to the substrate, was not significantly affected. Indirect immunofluorescence microscopy of endothelial cells after treatment with interleukin-1 alpha and beta, gamma-interferon, tumor necrosis factor (TNF), phorbol 12-myristate 13-acetate, and phorbol 12,13-dibutyrate allowed us to observe reductions in the areas of cell-cell contact, redistribution of the stress fiber network, and concomitant changes in focal contacts. Microtubule arrays in TNF-treated cells became bundled. Phorbol esters induced formation of microtubule organizing centers not seen in resting or TNF-treated HUVECs. Talin was distributed along stress fibers and not exclusively in focal contacts. Vitronectin receptor was observed in focal contacts, occasionally at cell-cell contacts, and in vesicular structures close to the lumenal surface, after both types of treatment. Although these morphological changes were easily observed by indirect immunofluorescence, no quantitative differences in specific cytoskeletal proteins were detected by immunoblots and [35S]cysteine metabolic labeling experiments.

Cell Adhesion Molecules↗