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Cell Binding (cell + binding)
Selected AbstractsLectin-Based Drug Design: Combined Strategy to Identify Lead Compounds using STD NMR Spectroscopy, Solid-Phase Assays and Cell Binding for a Plant Toxin ModelCHEMMEDCHEM, Issue 3 2010Abstract The growing awareness of the sugar code,i.e. the biological functionality of glycans,is leading to increased interest in lectins as drug targets. The aim of this study was to establish a strategic combination of screening procedures with increased biorelevance. As a model, we used a potent plant toxin (viscumin) and lactosides synthetically modified at the C6/C6, positions and the reducing end aglycan. Changes in the saturation transfer difference (STD) in NMR spectroscopy, applied in inhibition assays, yielded evidence for ligand activity and affinity differences. Inhibitory potency was confirmed by the blocking of lectin binding to a glycoprotein-bearing matrix. In cell-based assays, iodo/azido-substituted lactose derivatives were comparatively active. Interestingly, cell-type dependence was observed, indicating the potential of synthetic carbohydrate derivative to interact with lectins in a cell-type (glycan profile)-specific manner. These results are relevent to research into human lectins, glycosciences, and beyond. [source] Secretion of IL-12 by murine macrophages activated by immunoglobulin receptor-mediated internalization of the surface coat of Trichinella spiralis larvaePARASITE IMMUNOLOGY, Issue 3 2000Modha Trichinella spiralis larvae incubated with a rabbit antiserum raised against the larval surface coat bound murine macrophages to the parasite surface. Cell binding was not observed without the antisurface coat serum, or with incubation of larvae in normal rabbit serum, or with antibodies to keyhole limpet haemocyanin which identify a cryptic T. spiralis larval antigen. Cell adherence to the larval surface was lost by treatment of the cells with the lysosomotropic drug primaquine, implicating a receptor-mediated mechanism. Cells adhering to the parasite surface internalized parasite surface coat material, which was subsequently concentrated into endosomes. Culture supernatants from these cells contained enhanced levels of IL-12. Thus, the initial Th1 response to T. spiralis infection may be explained by these data. [source] Evolutions, Revolutions and Trends in Biomaterials Science , A Perspective,ADVANCED ENGINEERING MATERIALS, Issue 12 2007D. Jandt Abstract Despite a number of shortcomings biomaterials for implants have not changed much during the last decades. Yet, there is a revolution ongoing in fundamental biomaterials science research, which introduces new concepts and may help to answer a number of open questions. Bioactive and biomimetic nanomaterials in zero, one or two dimensions may be at the brink of transfer to clinical testing and application. These materials will in the future intervene actively in biological processes, such as protein adsorption, cell binding and growth. [source] Combining adenoviral oncolysis with temozolomide improves cell killing of melanoma cellsINTERNATIONAL JOURNAL OF CANCER, Issue 12 2007Christina Quirin Abstract Oncolytic Adenoviruses are emerging agents for treatment of cancer by tumor-restricted virus replication, cell lysis and virus spread. Clinical studies with first generation oncolytic adenoviruses have revealed that an increased potency is warranted in order to achieve therapeutic efficacy. One approach towards this end is to combine adenoviral oncolysis with chemotherapy. Here, a fundamental requirement is that chemotherapy does not interfere with adenovirus replication in cancer cells. We have previously developed a melanoma-targeted oncolytic adenovirus, Ad5/3.2xTyr, which features tyrosinase promoter regulated replication and enhanced cell entry into melanoma cells. In this study, we investigated a combination treatment of melanoma cells with Ad5/3.2xTyr and temozolomide (TMZ), which produces the same active metabolite as Dacarbazine/DTIC, the standard chemotherapy for advanced melanoma. We report that TMZ does not inhibit adenovirus replication in melanoma cells. Additive or synergistic cell killing of melanoma cells, dependent on the cell line used, was observed. Enhanced cell binding was not responsible for synergism of adenoviral oncolysis and TMZ treatment. We rather observed that higher numbers of virus genomes are produced in TMZ-treated cells, which also showed a cell cycle arrest in the G2 phase. Our results have important implications for the clinical implementation of adenoviral oncolysis for treatment of malignant melanoma. It suggests that such studies are feasible in the presence of TMZ or DTIC chemotherapy and recommends the investigation of a viro-chemo combination therapy. © 2007 Wiley-Liss, Inc. [source] Insulin-like growth factor (IGF) binding protein-3 regulation of IGF-I is altered in an acidic extracellular environmentJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2001Kimberly E. Forsten While extracellular acidification within solid tumors is well-documented, how reduced pH impacts regulation of insulin-like growth factor-I (IGF-I) has not been studied extensively. Because IGF-I receptor binding is affected by IGF binding proteins (IGFBPs), we examined how pH impacted IGFBP-3 regulation of IGF-I. IGF-I binding in the absence of IGFBP-3 was diminished at reduced pH. Addition of IGFBP-3 reduced IGF-I cell binding at pH 7.4 but increased surface association at pH 5.8. This increase in IGF-I binding at pH 5.8 corresponded with an increase in IGFBP-3 cell association. This, however, was not due to an increase in affinity of IGFBP-3 for heparin at reduced pH although both heparinase III treatment and heparin addition reduced IGFBP-3 enhancement of IGF-I binding. An increase in IGF-I binding to IGFBP-3, though, was seen at reduced pH using a cell-free assay. We hypothesize that the enhanced binding of IGF-I at pH 5.8 is facilitated by increased association of IGFBP-3 at this pH and that the resulting cell associated IGF-I is IGFBP-3 and not IGF-IR bound. Increased internalization and nuclear association of IGF-I at pH 5.8 in the presence of IGFBP-3 was evident, yet cell proliferation was reduced by IGFBP-3 at both pH 5.8 and 7.4 indicating that IGFBP-3-cell associated IGF-I does not signal the cell to proliferate and that the resulting transfer of bound IGF-I from IGF-IR to IGFBP-3 results in diminished proliferation. Solution binding of IGF-I by IGFBP-3 is one means by which IGF-I-induced proliferation is inhibited. Our work suggests that an alternative pathway exists by which IGF-I and IGFBP-3 both associate with the cell surface and that this association inhibits IGF-I-induced proliferation. © 2001 Wiley-Liss, Inc. [source] Involvement of complement receptor 3 (CR3) and scavenger receptor in macrophage responses to wear debrisJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 11 2006Diptendu S. Rakshit Abstract The ability of prosthetic wear debris to induce pro-inflammatory responses in macrophages is widely appreciated, but little is known about the molecular mechanisms involved in particle recognition. Specifically, the nature of the cell surface receptors that interact with wear debris is poorly understood. Elucidating the identities of these receptors and how they interact with different types of wear debris are critical to understanding how wear debris initiates periprosthetic osteolysis. We examined the involvement of opsonization, complement receptor 3 (CR3), and scavenger receptor A (SRA), in responses to polymethylmethacrylate (PMMA) and titanium wear particles. Serum dependence of pro-inflammatory responses to PMMA and titanium was tested, and serum proteins that adhered to these two types of particles were identified. Several serum proteins, including known opsonins such as C3bi and fibronectin, adhered to PMMA but not titanium, and serum was required for pro-inflammatory signaling induced by PMMA, but not by titanium. Phagocytosis of PMMA and titanium by macrophages was demonstrated by flow cytometry. Blocking CR3 specifically inhibited phagocytosis of PMMA by macrophages, whereas blocking SRA specifically inhibited titanium uptake. Direct involvement of CR3 and SRA in cell,particle interaction was assessed by expression of these receptors in nonphagocytic HEK293 cells. CR3 specifically induced cell binding to PMMA particles and adhesion to PMMA-coated plates, while SRA specifically induced binding to titanium particles and adhesion to titanium-coated plates. Taken together, these results suggest involvement of opsonization, complement, and integrin receptors, including CR3 and fibronectin receptors, in PMMA action, and an involvement of scavenger receptors in responses to titanium. © 2006 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 24:2036,2044, 2006 [source] Adaptation of the Lyme disease spirochaete to the mammalian host environment results in enhanced glycosaminoglycan and host cell bindingMOLECULAR MICROBIOLOGY, Issue 5 2003Nikhat Parveen Summary The Lyme disease spirochaete, Borrelia burgdorferi, is transmitted to mammals by Ixodes ticks and can infect multiple tissues. Host cell attachment may be critical for tissue colonization, and B. burgdorferi cultivated in vitro recognizes heparin- and dermatan sulphate-related glycosaminoglycans (GAGs) on the surface of mammalian cells. To determine whether growth of the spirochaete in the mammalian host alters GAG binding, we assessed the cell attachment activities of B. burgdorferi grown in vitro or in dialysis membrane chambers implanted intraperitoneally in rats. Host-adapted B. burgdorferi exhibited approximately threefold better binding to purified heparin and dermatan sulphate and to GAGs expressed on the surface of cultured endothelial cells. Three B. burgdorferi surface proteins, Bgp, DbpA and DbpB, have been demonstrated previously to bind to GAGs or to GAG-containing molecules, and we show here that recombinant derivatives of each of these proteins were able to bind to purified heparin and dermatan sulphate. Immunofluorescent staining of in vitro -cultivated or host-adapted spirochaetes revealed that DbpA and DbpB were present on the bacterial surface at higher levels after host adaptation. Recombinant Bgp, DbpA and DbpB each partially inhibited attachment of host-adapted B. burgdorferi to cultured mammalian cells, consistent with the hypothesis that these proteins may promote attachment of B. burgdorferi during growth in the mammalian host. Nevertheless, the partial nature of this inhibition suggests that multiple pathways promote mammalian cell attachment by B. burgdorferi in vivo. Given the observed increase in cell attachment activity upon growth in the mammalian host, analysis of host-adapted bacteria will facilitate identification of the cell binding pathways used in vivo. [source] Biological activity, membrane-targeting modification, and crystallization of soluble human decay accelerating factor expressed in E. coliPROTEIN SCIENCE, Issue 9 2004Jennifer White CHAPS, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate; GPI, glycophosphatidyl inositol; PpDAF, human DAF1,4 expressed in Pichia pastoris, N glycosylated and with an oligohistidine tag; EcDAF, nonglycosylated human DAF 1,4 expressed in Escherichia coli; nDAF, human native glycosylated (GPI-anchored) DAF from erythrocytes; EcDAF-MP, soluble E. coli human DAF linked through a C-terminal cysteine to the myristoylated peptide APT542; PCR, polymerase chain reaction; SCR, short consensus repeat; TCEP, Tris- (2-carboxyethyl) phosphine Abstract Decay-accelerating factor (DAF, CD55) is a glycophosphatidyl inositol-anchored glycoprotein that regulates the activity of C3 and C5 convertases. In addition to understanding the mechanism of complement inhibition by DAF through structural studies, there is also an interest in the possible therapeutic potential of the molecule. In this report we describe the cloning, expression in Escherichia coli, isolation and membrane-targeting modification of the four short consensus repeat domains of soluble human DAF with an additional C-terminal cysteine residue to permit site-specific modification. The purified refolded recombinant protein was active against both classical and alternative pathway assays of complement activation and had similar biological activity to soluble human DAF expressed in Pichia pastoris. Modification with a membrane-localizing peptide restored cell binding and gave a large increase in antihemolytic potency. These data suggested that the recombinant DAF was correctly folded and suitable for structural studies as well as being the basis for a DAF-derived therapeutic. Crystals of the E. coli -derived protein were obtained and diffracted to 2.2 Å, thus permitting the first detailed X-ray crystallography studies on a functionally active human complement regulator protein with direct therapeutic potential. [source] Identification of repertoires of surface antigens on leukemias using an antibody microarrayPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 11 2003Larissa Belov Abstract We have previously described a microarray of cluster of differentiation (CD) antibodies that enables concurrent determination of more than 60 CD antigens on leukocytes. This procedure does not require protein purification or labeling, or a secondary detection system. Whole cells are captured by a microarray of 10 nL antibody dots immobilized on a nitrocellulose film on a microscope slide. Distinct patterns of cell binding are observed for different leukemias or lymphomas. These haematological malignancies arise from precursor cells of T- or B-lymphocytic, or myeloid lineages of hematopoiesis. The dot patterns obtained from patients are distinct from those of peripheral blood leukocytes from normal subjects. This microarray technology has recently undergone a number of refinements. The microarray now contains more CD antibodies, and a scanner for imaging dot patterns and software for data analysis provide an extensive immunophenotype sufficient for diagnosis of common leukemias. The technology is being evaluated for diagnosis of leukemias with parallel use of conventional diagnostic criteria. [source] Thrombin-activatable carboxypeptidase B cleavage of osteopontin regulates neutrophil survival and synoviocyte binding in rheumatoid arthritisARTHRITIS & RHEUMATISM, Issue 10 2009Shadi A. Sharif Objective Osteopontin (OPN) is a proinflammatory cytokine that plays an important role in the pathogenesis of rheumatoid arthritis (RA). OPN can be cleaved by thrombin, resulting in OPN-R and exposing the cryptic C-terminal ,4,1 and ,9,1 integrin,binding motif (SVVYGLR). Thrombin-activatable carboxypeptidase B (CPB), also called thrombin-activatable fibrinolysis inhibitor, removes the C-terminal arginine from OPN-R, generating OPN-L and abrogating its enhanced cell binding. We undertook this study to investigate the roles of OPN-R and OPN-L in synoviocyte adhesion, which contributes to the formation of invasive pannus, and in neutrophil survival, which affects inflammatory infiltrates in RA. Methods Using specifically developed enzyme-linked immunosorbent assays, we tested the synovial fluid of patients with RA, osteoarthritis (OA), and psoriatic arthritis (PsA) to determine OPN-R, OPN-L, and full-length OPN (OPN-FL) levels. Results Elevated levels of OPN-R and OPN-L were found in synovial fluid samples from RA patients, but not in samples from OA or PsA patients. Increased levels of OPN-R and OPN-L correlated with increased levels of multiple inflammatory cytokines, including tumor necrosis factor , and interleukin-6. Immunohistochemical analyses revealed robust expression of OPN-FL, but only minimal expression of OPN-R, in RA synovium, suggesting that cleaved OPN is released into synovial fluid. In cellular assays, OPN-FL, and to a lesser extent OPN-R and OPN-L, had an antiapoptotic effect on neutrophils. OPN-R augmented RA fibroblast-like synoviocyte binding mediated by SVVYGLR binding to ,4,1, whereas OPN-L did not. Conclusion Thrombin activation of OPN (resulting in OPN-R) and its subsequent inactivation by thrombin-activatable CPB (generating OPN-L) occurs locally within inflamed joints in RA. Our data suggest that thrombin-activatable CPB plays a central homeostatic role in RA by regulating neutrophil viability and reducing synoviocyte adhesion. [source] | ||||||||||||||||