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Integrin
Kinds of Integrin Terms modified by Integrin Selected AbstractsA Novel Approach to Observing Synergy Effects of PHSRN on Integrin,RGD Binding Using Intelligent Surfaces,ADVANCED MATERIALS, Issue 16 2008Mitsuhiro Ebara A novel assay for measuring time-dependant ligand-receptor affinity changes is developed based on a peptide-immobilized temperature-responsive surface, as schematically illustrated in the figure. The grafted thermoresponsive polymer acts as an "on-off" switch for mediating integrin,peptide bonding. At temperatures above and below the lower critical solution temperature (LCST), the peptides are accessible and shielded from integrin access, respectively. [source] Integrin signaling through FAK in the regulation of mammary stem cells and breast cancerIUBMB LIFE, Issue 4 2010Jun-Lin Guan Abstract Focal adhesion kinase (FAK) is a cytoplasmic tyrosine kinase identified as a key mediator of intracellular signaling by integrins, a major family of cell surface receptors for extracellular matrix, in the regulation of different cellular functions in a variety of cells. Upon activation by integrins through disruption of an autoinhibitory mechanism, FAK undergoes autophosphorylation and forms a complex with Src and other cellular proteins to trigger downstream signaling through its kinase activity or scaffolding function. A number of integrins are identified as surface markers for mammary stem cells (MaSCs), and both integrins and FAK are found to play crucial roles in the maintenance of MaSCs in studies using mouse models, suggesting that integrin signaling through FAK may serve as a functional marker for MaSCs. Consistent with previous studies linking increased expression and activation of FAK to human breast cancer, these findings suggest a novel cellular mechanism of FAK promotion of mammary tumorigenesis by maintaining the pools of MaSCs as targets of oncogenic transformation. Furthermore, FAK inactivation in mouse models of breast cancer also reduced the pool of mammary cancer stem cells (MaCSCs), decreased their self-renewal in vitro, and compromised their tumorigenicity and maintenance in vivo, suggesting a potential role of integrin signaling through FAK in breast cancer growth and progression through its functions in MaCSCs. This review discusses these recent advances and future studies into the mechanism of integrin signaling through FAK in breast cancer through regulation of MaCSCs that may lead to development of novel therapies for this deadly disease. © 2010 IUBMB IUBMB Life, 62(4): 268,276, 2010 [source] Bone Mineralization and Osteoblast Differentiation Are Negatively Modulated by Integrin ,v,3JOURNAL OF BONE AND MINERAL RESEARCH, Issue 2 2001Su-Li Cheng Abstract Numerous bone matrix proteins can interact with ,v-containing integrins including ,v,3. To elucidate the net effects of the interaction between these proteins and ,v,3 on osteoblast function, we developed a murine osteoblastic cell line that overexpressed human ,v,3. Human ,v,3-integrin was expressed on cell membrane, in which its presence did not alter the surface level of endogenous mouse ,v,3. The expressed human ,v,3 was functional because cell adhesion to osteopontin was increased and this increment was abolished by antibody against human ,v,3. The proliferation rate of cells overexpressing ,v,3 (,v,3-cells) was increased whereas matrix mineralization was decreased. To elucidate the mechanisms leading to inhibition of matrix mineralization, the expression of proteins important for mineralization was analyzed. Alkaline phosphatase activity and the expression of osteocalcin, type I collagen, and bone sialoprotein (BSP) were decreased whereas osteopontin was stimulated in ,v,3-cells. The regulation of osteopontin, osteocalcin, and BSP expression was mediated via transcriptional mechanism because their promoter activities were altered. Examination of molecules involved in integrin signaling indicated that activator protein-1 (AP-1) and extracellular signal-regulated kinase (Erk) activities were enhanced whereas c-jun N -terminal kinase (JNK) activity was decreased in ,v,3-cells. The activity of p38 and the levels of focal adhesion kinase (FAK) and vinculin were not altered. Moreover, the adhesions of ,v,3-cells to type I collagen and fibronectin were inhibited, which was attributed to decreased ,1-integrin levels on cell surface. In conclusion, overexpressing ,v,3-integrin in osteoblasts stimulated cell proliferation but retarded differentiation, which were derived via altered integrin-matrix interactions, signal transduction, and matrix protein expression. [source] Integrin ,v,3 is involved in stimulated migration of vascular adventitial fibroblasts by basic fibroblast growth factor but not platelet-derived growth factorJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 1 2001Guizhen Liu Abstract We examined the effects of basic fibroblast growth factor (bFGF) and platelet-derived growth factor (PDGF) on the migration of vascular adventitial fibroblasts (VAFs) isolated from rat aortic adventitiae. Both bFGF and PDGF significantly stimulated VAF migration in vitro. An antibody to rat ,3 integrin reduced bFGF-stimulated migration in a dose dependent manner. Moreover, VAF migration was inhibited in the presence of cyclic RGD (cRGD) peptide. However, PDGF-directed migration was blocked only by equivalent cRGD peptide but not by antibody to ,3 integrin. These data suggest that ,v,3 integrin mediates VAF migration stimulated by bFGF and that chemoattractant directed migration may be through distinct integrins. J. Cell. Biochem. 83: 129,135, 2001. © 2001 Wiley-Liss, Inc. [source] Expression of RNAs encoding for , and , integrin subunits in periodontitis and in cyclosporin A gingival overgrowthJOURNAL OF CLINICAL PERIODONTOLOGY, Issue 11 2003A.-L. Bolcato-Bellemin Abstract Background: Variation of integrin expression in healthy and diseased gingiva revealed a potential biological role for these cell matrix receptors during gingival remodeling. Aim: Here we determined the level of RNA and tissue localization of different integrin subunits in periodontitis and cyclosporin A-induced gingival overgrowth. Methods: The level of expression was determined by Reverse Transcriptase Polymerase Chain Reaction in 12 periodontitis-affected patients, four patients exhibiting severe cyclosporin A-induced gingival overgrowth and seven healthy patients as controls. Results: The RNA encoding for ,1, ,2 and ,5 integrin subunits were reduced in periodontitis gingiva. The reduction observed was stronger in cyclosporin A-treated patients as compared to the healthy controls, while RNA encoding for ,1 subunit was increased. The RNA encoding for ,6 integrin was only reduced in cyclosporin A-treated gingiva. Immunohistochemistry showed that i) integrin ,2 expression is restricted to the gingival epithelium of cyclosporin A-treated patients, ii) the reduction of ,6 integrin expression in cyclosporin A-treated gingiva is due to loss of expression at focal contacts and iii) ,1 integrin is evenly distributed in the three populations with an intensity decrease in periodontitis and cyclosporin A-treated gingiva. Conclusion: Taken together these results showed a role for the integrin receptors in periodontal diseases and cyclosporin A-induced gingival overgrowth. Zusammenfassung Die Variation der Integrin Expression bei gesunder und erkrankter Gingiva zeigt eine potentielle biologische Rolle für diese Zellmatrixrezeptoren während der gingivalen Erneuerung. Wir bestimmten hier die Level von RNA und die Gewebelokalisation von unterschiedlichen Integrin Untereinheiten bei Parodontitis und Cyclosporin A induzierter gingivaler Wucherung. Die Level der Expression wurden mit der reversen Transscriptase Polymerase Kettenreaktion bei 12 Parodontitis-Patienten, 4 Patienten mit schwerer Cyclosporin A induzierter gingivaler Wucherung und sieben gesunden Kontrollpatienten bestimmt. Die kodierende RNA für ,1, ,2 und ,5 Integrin Untereinheiten waren in der Gingiva mit Parodontitis reduziert. Die beobachtete Reduktion war stärker bei den mit Cyclosporin A behandelten Patienten verglichen mit den gesunden Kontrollen, während kodierende RNA für ,1 Untereinheiten erhöht war. Die kodierende RNA für ,6 Integrin war nur bei der Cyclosporin A behandelten Gingiva reduziert. Die Immunhistochemie zeigte (i) die Integrin ,2 Expression ist auf das gingivale Epithel von Cyclosporin A behandelten Patienten beschränkt, (ii) die Reduktion von ,6 Integrin Expression bei Cyclosporin A behandelter Gingiva ist die Folge von fokalen Expressionverlusten und (iii) ,1 Integrin ist gleichmäßig verteilt in den drei Populationen mit einer Intensitätsabnahme bei Parodontitis und Cyclosporin A behandelter Gingiva. Zusammenfassend zeigen die Ergebnisse eine Rolle für die Integrin Rezeptoren bei den parodontalen Erkrankungen und Cyclosporin A induzierter gingivalen Wucherung. Résumé La variation d'expression des intégrines dans les tissus gingivaux sain et pathologique a démontré le rôle biologique potentiel de ces récepteurs de la matrice extracellulaire au cours du remodelage tissulaire gingival. La quantité d'ARN et la localisation tissulaire de certaines sous-unités d'intégrines dans la parodontite et l'hyperplasie gingivale induite par la ciclosporine A ont été déterminées. Le niveau d'expression a étéévalué par transcription inverse des ARN et réaction de polymérisation en chaine chez douze patients atteints de parodontite, quatre patients présentant une hyperplasie gingivale sévère induite par la ciclosporine A et sept patients sains ayant servi de témoins. L'expression des ARN codant pour les sous-unités ,1, ,2 et ,5était diminuée dans le tissu gingival atteint de parodontite. La diminution observée était plus importante chez les patients traités par la ciclosporine A, comparée aux témoins sains alors que l'expression de l'ARN codant pour la sous-unité,1était augmentée. L'expression de l'ARN codant pour la sous-unité,6était diminuée uniquement dans le tissu gingival traité par la ciclosporine A. L'immohistochimie a montré que (1) l'expression de la sous-unité,2 est limitée à l'épithélium gingival des patients traités par la ciclosporine A, (2) la diminution de l'expression de la sous-unité,6 dans le tissu gingival traité par la ciclosporine A est due à une perte des contacts focaux et (3) la sous-unité,1 est répartie de manière uniforme dans les trois groupes avec une diminution de l'intensité dans les cas de parodontite et d'hyperplasie gingivale induite par la ciclosporine A. Ces résultats montrent un rôle des récepteurs de type intégrine dans la pathologie parodontale et l'hyperplasie gingivale induite par la ciclosporine A. [source] Integrin ,3,1 interacts with I1PP2A/lanp and phosphatase PP1JOURNAL OF NEUROSCIENCE RESEARCH, Issue 8 2006Diana Mutz Abstract Integrin ,3,1 is a receptor for the extracellular matrix component laminin 5. To elucidate possible signaling pathways induced by integrin ,3,1, we looked for proteins that interact with the cytoplasmic part of the ,3A integrin subunit. We identified several multifunctional proteins by affinity chromatography and subsequent MALDI-TOF-MS and focused on the inhibitor 1 of serine/threonine phosphatase PP2A (I1PP2A, synonym: lanp) which also plays a role during the development of the mouse cerebellum. I1PP2A/lanp colocalizes with the ,3A integrin subunit in differentiated PC12 cells in the cell body and in neurites as well as in Purkinje cells of mouse cerebellum. Overexpression of GFP-I1PP2A/lanp in PC12 cells leads to markedly reduced neurite length on laminin 5 after induction with nerve growth factor. By affinity chromatography the protein phosphatase PP1 can also be identified as a ,3A/cyto-binding protein. PP1 and integrin ,3,1 can be pulled down by GST-I1PP2A/lanp from cell lysates of differentiated and undifferentiated PC12 cells. The phosphatase binds to the cytoplasmic membrane-proximal conserved GFFKR motif of the , integrin subunit, whereas I1PP2A/lanp requires a longer sequence for binding. PP1 but not PP2A is able to dephosphorylate precipitated integrin ,3,1 in vitro. Furthermore, PP1 releases phosphate from T1046 of phosphopeptides that mimic the phosphorylation consensus sequence in the cytoplasmic part of the ,3A integrin subunit. These data suggest that I1PP2A/lanp forms a complex with PP1 and the ,3A integrin subunit and might possibly regulate the phosphorylation status of integrin ,3,1 and/or integrin downstream targets. © 2006 Wiley-Liss, Inc. [source] Differential localization of laminin ,2 and integrin ,4 in primary cultures of the rat gingival epitheliumJOURNAL OF PERIODONTAL RESEARCH, Issue 1 2006Michie Tanno Objectives:, The aim of this study was to investigate the differential immunolocalization of laminin ,2 and integrin ,4 in primary cultures of the rat gingival epithelium. Methods:, The gingival epithelium was obtained from Sprague-Dawley rats and was cultured in serum-free keratinocyte growth medium (DK-SFM). Western blotting analysis, immunofluorescence, confocal laser scanning microscopy (CLSM), and immuno-gold labeling for laminin ,2 and integrin ,4 were employed. CLSM images for laminin and integrin were analyzed in horizontal (x,y axis) and in vertical (x,z axis) sections. Results:, Both laminin ,2 and integrin ,4 were detected by Western blot analysis in the gingival epithelium. Immunolocalization of laminin ,2 was distinct in the cytoplasm to form one or two irregular rings in gingival epithelial cells. By contrast, integrin ,4 was localized diffusely in the cytoplasm. F-actin (indicating actin filaments) was clearly discernible at the periphery of the cytoplasm to form a cellular fringe. In x,z axis images obtained by CLSM, laminin ,2 was recognized as large foci in the most inner portion just above the basal plasma membrane. Integrin ,4 existed in the area where F-actin was labeled surrounding the membrane. Immuno-electron microscopy showed that 10nm colloidal gold particles indicating laminin ,2 were mainly localized at the extracellular portion and in the peripheral cytoplasm, whereas integrin ,4 was distributed in the cytoplasm close to the basal plasma membrane but not in extracellular regions. Conclusions:, In primary cultures of the rat gingival epithelium, both laminin ,2 and integrin ,4 may be produced by the epithelium, and irregular rings of laminin ,2 are formed in areas where gingival cells adhere to the extracellular matrix. [source] Ethanol Increases Fetal Human Neurosphere Size and Alters Adhesion Molecule Gene ExpressionALCOHOLISM, Issue 2 2008Sharada D. Vangipuram Background:, Ethanol (ETOH) consumption by pregnant women can result in Fetal Alcohol Spectrum Disorder (FASD). To date, the cellular targets and mechanisms responsible for FASD are not fully characterized. Our aim was to determine if ETOH can affect fetal human brain-derived neural progenitor cells (NPC). Methods:, Neural progenitor cells were isolated by positive selection from normal second trimester fetal human brains (n = 4) and cultured, for up to 72 hours, in mitogenic media containing 0, 1, 10, or 100 mM ETOH. From 48 to 72 hours in culture, neurospheres generated in these conditions were filmed using time-lapse video microscopy. At the end of 72 hours, neurosphere diameter and roundness were measured using videographic software. Mitotic phase analysis of cell-cycle activity and apoptotic cell count were also performed at this time, by flow cytometry using propidium iodide (PI) staining. Real-time PCR was used to estimate expression of genes associated with cell adhesion pathways. Results:, Neurosphere diameter correlated positively (r = 0.87) with increasing ETOH concentrations. There was no significant difference in cell-cycle activity and no significant increase in apoptosis with increasing ETOH concentrations. Time-lapse video microscopy showed that ETOH (100 mM) reduced the time for neurosphere coalescence. Real-time PCR analysis showed that ETOH significantly altered the expression of genes involved in cell adhesion. There was an increase in the expression of , and , Laminins 1, , Integrins 3 and 5, Secreted phosphoprotein1 and Sarcoglycan ,. No change in the expression of , Actin was observed while the expression of , Integrin 2 was significantly suppressed. Conclusions:, ETOH had no effect on NPC apoptosis but, resulted in more rapid coalescence and increased volume of neurospheres. Additionally, the expression of genes associated with cell adhesion was significantly altered. ETOH induced changes in NPC surface adhesion interactions may underlie aspects of neurodevelopmental abnormalities in FASD. [source] Platelet integrin ,IIb,3: activation mechanismsJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 7 2007Y.-Q. MA Summary., Integrin ,IIb,3 plays a critical role in platelet aggregation, a central response in hemostasis and thrombosis. This function of ,IIb,3 depends upon a transition from a resting to an activated state such that it acquires the capacity to bind soluble ligands. Diverse platelet agonists alter the cytoplasmic domain of ,IIb,3 and initiate a conformational change that traverses the transmembrane region and ultimately triggers rearrangements in the extracellular domain to permit ligand binding. The membrane-proximal regions of ,IIb and ,3 cytoplasmic tails, together with the transmembrane segments of the subunits, contact each other to form a complex which restrains the integrin in the resting state. It is unclasping of this complex that induces integrin activation. This clasping/unclasping process is influenced by multiple cytoplasmic tail binding partners. Among them, talin appears to be a critical trigger of ,IIb,3 activation, but other binding partners, which function as activators or suppressors, are likely to act as co-regulators of integrin activation. [source] Initiating and potentiating role of platelets in tissue factor-induced thrombin generation in the presence of plasma: subject-dependent variation in thrombogram characteristicsJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 3 2004K. Vanschoonbeek Summary., The hemostatic activity of plasma is determined by platelet activation and coagulation, which processes are mutually stimulatory. We studied this interaction by measuring the cleavage of fluorescent thrombin substrate in platelet-rich plasma (PRP), using the calibrated thrombogram method. In freshly isolated human plasma, thrombin formation triggered by tissue factor was fully dependent on the presence of platelets. It was abolished by annexin A5, indicating dependence on phosphatidylserine (PS) exposure at activated platelets. Comparison of plasmas from various subjects showed considerable interindividual variation in total amount of thrombin generation, regardless of whether platelets or PS-containing phospholipids were present. Integrin ,IIb,3 antagonists and ADP receptor blockage, but not aspirin, decreased the rate of thrombin generation (thrombin peak level) and extended the time of onset. Platelet inhibition with cAMP-elevating agents decreased the thrombin-forming rate, but surprisingly shortened the onset time. Stimulation of platelets with agonists of Gi/q-coupled receptors and, to a larger extent, with collagen or Ca2+ -ionophore increased the rate of thrombin generation and shortened its onset. In PRP from donors with low and high generation, platelet inhibitors and activators were similarly effective. Taken together, these results indicate that, in tissue factor-triggered PRP, PS exposure on activated platelets regulates both onset and rate of thrombin generation. However, coagulant activity rather than platelet activation determines the total amount of thrombin formed, i.e. the endogenous thrombin potential. Thus, kinetics of thrombin generation in PRP are controlled by platelet inhibitors and agonists, but the process is restricted in amount by the subject-dependent variation in coagulation. [source] An Integrin and Rho GTPase-Dependent Pinocytic Vacuole Mechanism Controls Capillary Lumen Formation in Collagen and Fibrin MatricesMICROCIRCULATION, Issue 1 2003GEORGE E. DAVIS ABSTRACT A major question that remains unanswered concerning endothelial cell (EC) morphogenesis is how lumens are formed in three-dimensional extracellular matrices (ECMs). Studies from many laboratories have revealed a critical role for an ECM-integrin-cytoskeletal signaling axis during EC morphogenesis. We have discovered a mechanism involving intracellular vacuole formation and coalescence that is required for lumen formation in several in vitro models of morphogenesis. In addition, a series of studies have observed vacuoles in vivo during angiogenic events. These vacuoles form through an integrin-dependent pinocytic mechanism in either collagen or fibrin matrices. In addition, we have shown that the Cdc42 and Rac1 guanosine triphosphatases (GTPases), which control actin and microtubule cytoskeletal networks, are required for vacuole and lumen formation. These GTPases are also known to regulate integrin signaling and are activated after integrin-matrix interactions. Furthermore, the expression of green fluorescent protein-Rac1 or -Cdc42 chimeric proteins in ECs results in the targeting of these fusion proteins to intracellular vacuole membranes during lumen formation. Thus, a matrix-integrin-cytoskeletal signaling axis involving both the Cdc42 and Rac1 GTPases regulates the process of EC lumen formation in three-dimensional collagen or fibrin matrices. [source] Integrin ,IIb,3:ligand interactions are linked to binding-site remodelingPROTEIN SCIENCE, Issue 8 2006Roy R. Hantgan Abstract This study tested the hypothesis that high-affinity binding of macromolecular ligands to the ,IIb,3 integrin is tightly coupled to binding-site remodeling, an induced-fit process that shifts a conformational equilibrium from a resting toward an open receptor. Interactions between ,IIb,3 and two model ligands,echistatin, a 6-kDa recombinant protein with an RGD integrin-targeting sequence, and fibrinogen's ,-module, a 30-kDa recombinant protein with a KQAGDV integrin binding site,were measured by sedimentation velocity, fluorescence anisotropy, and a solid-phase binding assay, and modeled by molecular graphics. Studying echistatin variants (R24A, R24K, D26A, D26E, D27W, D27F), we found that electrostatic contacts with charged residues at the ,IIb/,3 interface, rather than nonpolar contacts, perturb the conformation of the resting integrin. Aspartate 26, which interacts with the nearby MIDAS cation, was essential for binding, as D26A and D26E were inactive. In contrast, R24K was fully and R24A partly active, indicating that the positively charged arginine 24 contributes to, but is not required for, integrin recognition. Moreover, we demonstrated that priming,i.e., ectodomain conformational changes and oligomerization induced by incubation at 35°C with the ligand-mimetic peptide cHarGD,promotes complex formation with fibrinogen's ,-module. We also observed that the ,-module's flexible carboxy terminus was not required for ,IIb,3 integrin binding. Our studies differentiate priming ligands, which bind to the resting receptor and perturb its conformation, from regulated ligands, where binding-site remodeling must first occur. Echistatin's binding energy is sufficient to rearrange the subunit interface, but regulated ligands like fibrinogen must rely on priming to overcome conformational barriers. [source] Proteomic study of DBA/2J mice retina: Down-regulation of Integrin ,7 correlated with retinal ganglion cell deathPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 21 2009Takashi Kanamoto Abstract To identify and determine the function of the proteins associated with the death of retinal ganglion cells (RGCs) in DBA/2J mice, an animal model of glaucoma, retinas of DBA/2J mice, were analyzed by proteomics at 5-, 7-, and 11-months-of-age. The proteins showing significant alterations were selected for identification by MS and 18 proteins were differentially expressed and the identified proteins included cell membrane receptors and proteins associated with intracellular signaling pathways. Among of identified proteins, the expression of Integrin ,7 at 7-months-of-age was decreased by about 89% of that at 5-months-of-age. Integrin ,7 was expressed in the RGCs. The effect of glutamate toxicity on the expression pattern of Integrin ,7 in a RGC line was also investigated and the glutamate-induced death of RGC was inhibited by the RNA knockdown of Integrin ,7. Our data showed also that the expression of 18 proteins in the DBA/2J was significantly altered in DBA2 mice and down-regulation of Integrin ,7 may have a protective effect on glutamate-induced death of RGCs. [source] Osteopontin stimulates invasion of NCI-h295 cells but is not associated with survival in adrenocortical carcinoma,THE JOURNAL OF PATHOLOGY, Issue 2 2009Dirk Weismann Abstract Gene array studies indicated that osteopontin (OPN) mRNA is highly expressed in adrenocortical carcinomas (ACCs). OPN enhances invasiveness, proliferation, and metastasis formation, and is associated with poor survival in some malignant diseases. Integrin ,v,3 has been shown to mediate OPN effects on invasion. In this study, we demonstrated OPN and integrin ,v,3 expression in normal adrenal glands and benign adenomas, with staining seen exclusively in adrenocortical cells as well as even stronger staining in ACC. Western blot analysis confirmed overexpression of OPN in ACC (p < 0.01). With Matrigel invasion assays, we have shown that OPN greatly stimulates the invasiveness of NCI-h295 cells (>six-fold increase, p < 0.001). Transfection with integrin ,v,3 further increased invasiveness after OPN stimulation (p < 0.001). This increase was reversed by the addition of an anti-integrin ,3 antibody, indicating a functional relationship of OPN and integrin ,v,3 in ACC. With tissue arrays, we confirmed high OPN expression in 147 ACC samples. However, no association with survival was seen in Kaplan-Meier analysis including 111 patients with primary tumours graded for OPN staining and follow-up data available. In conclusion, our in vitro data indicate that OPN and integrin ,v,3 may act as a functional complex facilitating the invasiveness of adrenocortical tumours. This relationship remains of relevance to our understanding of carcinogenesis, but further studies are needed to address the physiological and pathological function of OPN in adrenal tissue. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [source] ORIGINAL ARTICLE: Two Different Homing Pathways Involving Integrin ,7 and E-selectin Significantly Influence Trafficking of CD4 Cells to the Genital Tract Following Chlamydia muridarum InfectionAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 6 2009Kathleen A. Kelly Problem,Chlamydia trachomatis causes STI and reproductive dysfunction worldwide which is not preventable with antibiotics. Identifying a population of endocervical T cells to target in vaccine development would enhance efficacy. Method of study, Trafficking of murine CD4+ lymphocytes to Chlamydia muridarum infected genital tract (GT) tissue in vivo was measured using adoptive transfer studies of fluorescent CD4+ T cells from integrin ,7,/, mice or mice which lack E-selectin on endothelial cells. Results, Murine in vivo migration studies showed that lack of ,4,7 or E-selectin significantly reduced trafficking of CD4 T cells to the GT of mice infected with C. muridarum. Conclusion, CD4+ T cells use at least two different adhesive mechanisms involving an integrin of the mucosal homing pathway and selectin pathway to accumulate in the GT during C. muridarum infection. [source] ORIGINAL ARTICLE: The Combination of the Gastrointestinal Integrin (,4,7) and Selectin Ligand Enhances T-Cell Migration to the Reproductive Tract During Infection with Chlamydia trachomatisAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 6 2009Kathleen A. Kelly Problem,Chlamydia trachomatis causes sexually transmitted infection and reproductive dysfunction worldwide. Identifying a population of endocervical T-cells to target in vaccine development is likely to enhance efficacy of a vaccine and reduce reproductive tract dysfunction. Method of study, Endocervical samples were obtained from young women and flow cytometric analysis was used to identify lymphocytes that appeared in the genital tract in response to sexually transmitted bacterial infections caused by C. trachomatis. Results, Increased numbers of ,4,7+CLA+ memory T-cells, a unique T-cell phenotype, were found in the endocervix of human female subjects infected with C. trachomatis. Conclusion A unique population of memory T lymphocytes expressing both ,4,7 and CLA gain access to reproductive tract tissues during a sexually transmitted infection with C. trachomatis and should be considered in development of vaccines against sexually transmitted infections. [source] Transmembranstruktur von Integrin ,IIb,3 , Bedeutung für die Signalübertragung,ANGEWANDTE CHEMIE, Issue 36 2009Martin Hoefling Kontakt nach draußen: Zellen in Mehrzellern regulieren ihre Adhäsion durch Integrinrezeptoren, die Signale bidirektional zwischen dem Zellinneren und -äußeren übertragen. Die Transmembrandomänen (blau im Bild; rot/grün: Integrin/Talin-Komplexe) von Integrinen sind der Dreh- und Angelpunkt der Signalweiterleitung. Vor kurzem wurden Strukturen dieser Domänen vorgestellt, die den Mechanismus der Signalweiterleitung in neuem Licht erscheinen lassen. [source] Skin wound healing in diabetic ,6 integrin-deficient miceAPMIS, Issue 10 2010JASPER N. JACOBSEN Jacobsen JN, Steffensen B, Häkkinen L, Krogfelt KA, Larjava HS. Skin wound healing in diabetic ,6 integrin-deficient mice. APMIS 2010; 118: 753,64. Integrin ,v,6 is a heterodimeric cell surface receptor, which is absent from the normal epithelium, but is expressed in wound-edge keratinocytes during re-epithelialization. However, the function of the ,v,6 integrin in wound repair remains unclear. Impaired wound healing in patients with diabetes constitutes a major clinical problem worldwide and has been associated with the accumulation of advanced glycated endproducts (AGEs) in the tissues. AGEs may account for aberrant interactions between integrin receptors and their extracellular matrix ligands such as fibronectin (FN). In this study, we compared healing of experimental excisional skin wounds in wild-type (WT) and ,6-knockout (,6,/,) mice with streptozotocin-induced diabetes. Results showed that diabetic ,6,/, mice had a significant delay in early wound closure rate compared with diabetic WT mice, suggesting that ,v,6 integrin may serve as a protective role in re-epithelialization of diabetic wounds. To mimic the glycosylated wound matrix, we generated a methylglyoxal (MG)-glycated variant of FN. Keratinocytes utilized ,v,6 and ,1 integrins for spreading on both non-glycated and FN-MG, but their spreading was reduced on FN-MG. These findings indicated that glycation of FN and possibly other integrin ligands could hamper keratinocyte interactions with the provisional matrix proteins during re-epithelialization of diabetic wounds. [source] Expression of Integrin ,v,3 in Gliomas Correlates with Tumor Grade and Is not Restricted to Tumor VasculatureBRAIN PATHOLOGY, Issue 3 2008Oliver Schnell MD Abstract In malignant gliomas, the integrin adhesion receptors seem to play a key role for invasive growth and angiogenesis. However, there is still a controversy about the expression and the distribution of ,v,3 integrin caused by malignancy. The aim of our study was to assess the extent and pattern of ,v,3 integrin expression within primary glioblastomas (GBMs) compared with low-grade gliomas (LGGs). Tumor samples were immunostained for the detection of ,v,3 integrin and quantified by an imaging software. The expression of ,v,3 was found to be significantly higher in GBMs than in LGGs, whereby focal strong reactivity was restricted to GBMs only. Subsequent analysis revealed that not only endothelial cells but also, to a large extent, glial tumor cells contribute to the overall amount of ,v,3 integrin in the tumors. To further analyze the integrin subunits, Western blots from histologic sections were performed, which demonstrated a significant difference in the expression of the ,3 integrin subunit between GBMs and LGGs. The presented data lead to new insights in the pattern of ,v,3 integrin in gliomas and are of relevance for the inhibition of ,v,3 integrin with specific RGD peptides and interfering drugs to reduce angiogenesis and tumor growth. [source] Up-regulation of integrin ,3 in radioresistant pancreatic cancer impairs adenovirus-mediated gene therapyCANCER SCIENCE, Issue 10 2009Takuya Egami Adenovirus-mediated gene therapy is a promising approach for the treatment of pancreatic cancer. We previously reported that radiation enhanced adenovirus-mediated gene expression in pancreatic cancer, suggesting that adenoviral gene therapy might be more effective in radioresistant pancreatic cancer cells. In the present study, we compared the transduction efficiency of adenovirus-delivered genes in radiosensitive and radioresistant cells, and investigated the underlying mechanisms. We used an adenovirus expressing the hepatocyte growth factor antagonist, NK4 (Ad-NK4), as a representative gene therapy. We established two radioresistant human pancreatic cancer cell lines using fractionated irradiation. Radiosensitive and radioresistant pancreatic cancer cells were infected with Ad-NK4, and NK4 levels in the cells were measured. In order to investigate the mechanisms responsible for the differences in the transduction efficiency between these cells, we measured expression of the genes mediating adenovirus infection and endocytosis. The results revealed that NK4 levels in radioresistant cells were significantly lower (P < 0.01) than those in radiosensitive cells, although there were no significant differences in adenovirus uptake between radiosensitive cells and radioresistant cells. Integrin ,3 was up-regulated and the Coxsackie virus and adenovirus receptor was down-regulated in radioresistant cells, and inhibition of integrin ,3 promoted adenovirus gene transfer. These results suggest that inhibition of integrin ,3 in radioresistant pancreatic cancer cells could enhance adenovirus-mediated gene therapy. (Cancer Sci 2009; 100: 1902,1907) [source] N-Aryl-,-lactams as Integrin ,v,3 Antagonists.CHEMINFORM, Issue 38 2004Celia Dominguez Abstract For Abstract see ChemInform Abstract in Full Text. [source] 111Indium-labelled human gut-derived T cells from healthy subjects with strong in vitro adhesion to MAdCAM-1 show no detectable homing to the gut in vivoCLINICAL & EXPERIMENTAL IMMUNOLOGY, Issue 1 2004J. KELSEN SUMMARY Integrin ,4,,7 is the principal gut-homing receptor, and it is assumed that expression of this specific integrin directs lymphocytes to the gut in vivo. Adoptive cellular immunotherapy against inflammatory bowel disease (IBD) may depend on the expression of integrin ,4,,7 to accomplish local delivery of intravenously injected regulatory T cells in inflamed gut mucosa. The present study aimed to investigate whether in vitro expanded human T cells from the colonic mucosa maintain integrin expression, show in vitro adhesion and retain in vivo gut-homing properties during cultivation. Whole colonic biopsies from healthy subjects were cultured in the presence of interleukin-2 (IL-2) and IL-4. The integrin expression of the cultured T cells was determined by flow cytometry and in vitro adhesion was assessed in a mucosal addressin cell adhesion molecule 1 (MAdCAM-1) adhesion assay. We studied the homing pattern after autologous infusion of 3 × 108 111Indium (111In)-labelled T cells in five healthy subjects using scintigraphic imaging. The cultured CD4+CD45RO+ gut-derived T cells express higher levels of integrin ,4,,7 than peripheral blood lymphocytes (PBLs) and show strong adhesion to MAdCAM-1 in vitro, even after 111In-labelling. Scintigraphic imaging, however, showed no gut-homing in vivo. After prolonged transit through the lungs, the T cells migrated preferentially to the spleen, liver and bone marrow. In conclusion, it is feasible to infuse autologous T cells cultured from the gut mucosa, which may be of interest in adoptive immunotherapy. Despite high expression of the gut-homing integrin ,4,,7 and adhesion to MAdCAM-1 in vitro, evaluation by 111In-scintigraphy demonstrated no gut-homing in healthy individuals. [source] Lowered albumin extravasation rate in heart but not in other organs in ,3-integrin-deficient miceACTA PHYSIOLOGICA, Issue 4 2009Ø. S. Svendsen Abstract Aim:, The vascular protein permeability is dependent on the integrity of the vascular wall. The heart capillaries in male mice lacking ,3 integrins have an immature phenotype. Previously, we have demonstrated a role for ,v,3 integrins in control of interstitial fluid pressure (Pif) and thereby in the fluid flux during inflammation. We wanted to explore a possible role for ,v,3 integrins in controlling capillary protein permeability during control situation and inflammation. Methods:, We performed double-tracer and microdialysis experiments on ,3-integrin-deficient mice and wild type control mice. We also measured blood pressure and heart rate in the two mice strains. Results:, We found reduced albumin extravasation (during 25 min) in the heart capillaries (0.053 ± 0.003 vs. 0.087 ± 0.009 mL g,1 dw, P < 0.05), and an increased cardiac mass/body weight (5.3 × 10,3 ± 0.3 × 10,3 vs. 3.8 × 10,3 ± 0.1 × 10,3, P < 0.01) in the ,3-integrin-deficient mice (n = 6) compared with the controls (n = 6). Heart rate and blood pressure were the same in mice with and without ,3-integrins. No difference in permeability was found in other tissues studied, or under local inflammation. Conclusion:, These results show a function for the ,v,3 integrin in the regulation of protein permeability, selective for the heart capillaries. [source] Correlation of dystrophin,glycoprotein complex and focal adhesion complex with myosin heavy chain isoforms in rat skeletal muscleACTA PHYSIOLOGICA, Issue 4 2009S. Masuda Abstract Aim:, The dystrophin,glycoprotein complex (DGC) and focal adhesion complex (FAC) are transmembrane structures in muscle fibres that link the intracellular cytoskeleton to the extracellular matrix. DGC and FAC proteins are abundant in slow-type muscles, indicating the structural reinforcement which play a pivotal role in continuous force output to maintain posture for long periods. The aim of the present study was to examine the expression of these structures across fast-type muscles containing different myosin heavy chain (MHC) isoform patterns which reflect the fatigue-resistant characteristics of skeletal muscle. Methods:, We measured the expression of dystrophin and ,1 integrin (representative proteins of DGC and FAC respectively) in plantaris, extensor digitorum longus, tibialis anterior, red and white portions of gastrocnemius, superficial portion of vastus lateralis and diaphragm, in comparison with soleus (SOL) and cardiac muscle from rats. Results:, The expression of dystrophin and ,1 integrin correlated positively with the percentage of type I, IIa and IIx MHC isoforms and negatively with that of type IIb MHC isoform in fast-type skeletal muscles, and their expression was abundant in SOL and cardiac muscle. Conclusion:, Our results support the idea that DGC and FAC are among the factors that explain the fatigue-resistant property not only of slow-type but also of fast-type skeletal muscles. [source] Overexpression of CD49f in precursor B-cell acute lymphoblastic leukemia: Potential usefulness in minimal residual disease detectionCYTOMETRY, Issue 2 2009Joseph A. DiGiuseppe Abstract Background: The persistence of minimal residual disease (MRD) following therapy is an established prognostic factor in precursor B-cell acute lymphoblastic leukemia (pB-ALL). Detection of MRD in pB-ALL by flow cytometric immunophenotyping requires demonstration of abnormal antigen expression in leukemic B-cell precursors relative to that of normal B-cell precursors. The gene encoding CD49f (integrin ,-6) is one of several whose overexpression in pB-ALL at diagnosis has been associated with the subsequent detection of MRD. However, whether CD49f might be a useful reagent in the immunophenotypic detection of MRD in pB-ALL has not been evaluated. Methods: We evaluated CD49f expression by 4-color flow cytometry in normal B-cell precursors, and in a series of cases of pB-ALL, both at diagnosis and at intervals following the initiation of therapy. Results: In 10 control marrow samples, CD49f was undetectable or extremely dim in all but a minor subset of normal CD19+ B-lineage cells, whereas in 11 of 15 cases (73%) of pB-ALL, CD49f was moderate or bright at diagnosis, and persisted or became brighter after initiation of therapy. MRD detected using CD49f corresponded precisely with that obtained using a standard panel of antibodies, and permitted the detection of leukemic populations comprising as little as 0.02% of cells. Of the four pB-ALL cases in which CD49f was undetectable or dim at diagnosis, MRD was detected in two; in one of these, CD49f expression was substantially increased in the leukemic cells that persisted following initiation of therapy. Conclusions: CD49f is commonly overexpressed in p-B-ALL, and represents a potentially useful marker for the immunophenotypic detection of MRD. © 2008 Clinical Cytometry Society How to cite this article: DiGiuseppe JA, Fuller SG, Borowitz MJ. Overexpression of CD49f in precursor B-cell acute lymphoblastic leukemia: potential usefulness in minimal residual disease detection. Cytometry Part B 2008. [source] Characterization of L-plastin interaction with beta integrin and its regulation by micro-calpain,CYTOSKELETON, Issue 5 2010E. Le Goff Abstract Recent evidences suggest that plastin/fimbrin is more than a simple actin cross-linking molecule. In this context and based on the fact that other members of the same family interact with transmembrane proteins, such as integrins, we have investigated a possible interaction between L-plastin and integrins. By combining coimmunoprecipitation of endogenous proteins and in vitro techniques based on solid phase and solution assays, we demonstrate that L-plastin is an additional binding partner for the ,-chain of integrin and confirmed that both proteins display some colocalization. We then show that L-plastin binds to the cytoplasmic domain of ,1 integrin and to ,1 and ,2 peptides. Using recombinant L-plastin domains, we demonstrate that the integrin-binding sites are not located in NH2 terminal part of L-plastin but rather in the two actin-binding domains. Using pull-down, cross-linking experiments, and enzyme-linked immunosorbent assay, we show that the L-plastin/integrin complex is regulated by ,-calpain cleavage and is not directly dissociated by calcium. Indeed, despite the ability of calpain to cleave both proteins, only the cleavage of , integrin hindered the formation of the L-plastin/integrin complex. We discuss these results in the light of the three-dimensional structure of the actin-binding domains of L-plastin. © 2010 Wiley-Liss, Inc. [source] The effect of combined hypergravity and microgrooved surface topography on the behaviour of fibroblastsCYTOSKELETON, Issue 7 2006W. A. Loesberg Abstract This study evaluated in vitro the differences in morphological behaviour between fibroblast cultured on smooth and microgrooved substrata (groove depth: 1 ,m, width: 1, 2, 5, 10 ,m), which undergo artificial hypergravity by centrifugation (10, 24 and 50 g; or 1 g control). The aim of the study was to clarify which of these parameters was more important to determine cell behaviour. Morphological characteristics were investigated using scanning electron microscopy and fluorescence microscopy in order to obtain qualitative information on cell spreading and alignment. Confocal laser scanning microscopy visualised distribution of actin filaments and vinculin anchoring points through immunostaining. Finally, expression of collagen type I, fibronectin, and ,1 - and ,1 -integrin were investigated by PCR. Microscopy and image analysis showed that the fibroblasts aligned along the groove direction on all textured surfaces. On the smooth substrata (control), cells spread out in a random fashion. The alignment of cells cultured on grooved surfaces increased with higher g-forces until a peak value at 25 g. An ANOVA was performed on the data, for all main parameters: topography, gravity force, and time. In this analysis, all parameters proved significant. In addition, most gene levels were reduced by hypergravity. Still, collagen type 1 and fibronectin are seemingly unaffected by time or force. From our data it is concluded that the fibroblasts primarily adjust their shape according to morphological environmental cues like substratum surface whilst a secondary, but significant, role is played by hypergravity forces. Cell Motil. Cytoskeleton 2006. © 2006 Wiley-Liss, Inc. [source] Effects of substrate stiffness on cell morphology, cytoskeletal structure, and adhesionCYTOSKELETON, Issue 1 2005Tony Yeung Abstract The morphology and cytoskeletal structure of fibroblasts, endothelial cells, and neutrophils are documented for cells cultured on surfaces with stiffness ranging from 2 to 55,000 Pa that have been laminated with fibronectin or collagen as adhesive ligand. When grown in sparse culture with no cell-cell contacts, fibroblasts and endothelial cells show an abrupt change in spread area that occurs at a stiffness range around 3,000 Pa. No actin stress fibers are seen in fibroblasts on soft surfaces, and the appearance of stress fibers is abrupt and complete at a stiffness range coincident with that at which they spread. Upregulation of ,5 integrin also occurs in the same stiffness range, but exogenous expression of ,5 integrin is not sufficient to cause cell spreading on soft surfaces. Neutrophils, in contrast, show no dependence of either resting shape or ability to spread after activation when cultured on surfaces as soft as 2 Pa compared to glass. The shape and cytoskeletal differences evident in single cells on soft compared to hard substrates are eliminated when fibroblasts or endothelial cells make cell-cell contact. These results support the hypothesis that mechanical factors impact different cell types in fundamentally different ways, and can trigger specific changes similar to those stimulated by soluble ligands. Cell Motil. Cytoskeleton 60:24,34, 2005. © 2004 Wiley-Liss, Inc. [source] Early molecular events in the assembly of the focal adhesion-stress fiber complex during fibroblast spreadingCYTOSKELETON, Issue 3 2004Baruch Zimerman Cell adhesion to the extracellular matrix triggers the formation of integrin-mediated contact and reorganization of the actin cytoskeleton. Examination of nascent adhesions, formed during early stages of fibroblast spreading, reveals a variety of forms of actin-associated matrix adhesions. These include: (1) small (,1 ,m), dot-like, integrin-, vinculin-, paxillin-, and phosphotyrosine-rich structures, with an F-actin core, broadly distributed over the ventral surfaces of the cells; (2) integrin-, vinculin-, and paxillin-containing "doublets" interconnected by short actin bundles; (3) arrays of actin-vinculin complexes. Such structures were formed by freshly plated cells, as well as by cells recovering from latrunculin treatment. Time-lapse video microscopy of such cells, expressing GFP-actin, indicated that long actin cables are formed by an end-to-end lining-up and apparent fusion of short actin bundles. All these structures were prominent during cell spreading, and persisted for up to 30,60 min after plating. Upon longer incubation, they were gradually replaced by stress fibers, associated with focal adhesions at the cell periphery. Direct examination of paxillin and actin reorganization in live cells revealed alignment of paxillin doublets, forming long and highly dynamic actin bundles, undergoing translocation, shortening, splitting, and convergence. The mechanisms underlying the assembly and reorganization of actin-associated focal adhesions and the involvement of mechanical forces in regulating their dynamic properties are discussed. Cell Motil. Cytoskeleton 58:143,159, 2004. © 2004 Wiley-Liss, Inc. [source] Myosin-mediated cytoskeleton contraction and Rho GTPases regulate laminin-5 matrix assemblyCYTOSKELETON, Issue 2 2004Gregory W. deHart Abstract Laminin-5 is a major structural element of epithelial tissue basement membranes. In the matrix of cultured epithelial cells, laminin-5 is arranged into intricate patterns. Here we tested a hypothesis that myosin II-mediated actin contraction is necessary for the proper assembly of a laminin-5 matrix by cultured SCC12 epithelial cells. To do so, the cells were treated with ML-7, a myosin II light chain kinase inhibitor, or Y-27632, an inhibitor of Rho-kinase (ROCK), both of which block actomyosin contraction. Under these conditions, laminin-5 shows an aberrant localization in dense patches at the cell periphery. Since ROCK activity is regulated by the small GTPase Rho, this suggests that members of the Rho family of GTPases may also be important for laminin-5 matrix assembly by SCC12 cells. We confirmed this hypothesis since SCC12 cells expressing mutant proteins that inhibit RhoA, Rac, and Cdc42 assemble the same aberrant laminin-5 protein arrays as drug-treated cells. We have also evaluated the organization of the laminin-5 receptors ,3,1 and ,6,4 integrin and hemidesmosome proteins in ML-7- and Y-27632-treated cells or in cells in which RhoA, Rac, and Cdc42 activity were inhibited. In all instances, ,3,1 and ,6,4 integrin heterodimers, as well as hemidesmosome proteins, localize precisely with laminin-5 in the matrix of the cells. In summary, our results provide evidence that myosin II-mediated actin contraction and the activity of Rho GTPases are necessary for the proper organization of a laminin-5 matrix and localization of hemidesmosome protein arrays in epithelial cells. Cell Motil. Cytoskeleton 57:107,117, 2004. © 2004 Wiley-Liss, Inc. [source] | |||||||||||||||||||||||||||||||||||||