SEARCH · PubMed Health

Results for “Integrin alpha5beta1”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 667 records · Page 37Linked to original sources

PubMed2005

Three-dimensional migration of human adult dermal fibroblasts from collagen lattices into fibrin/fibronectin gels requires syndecan-4 proteoglycan.

Fibroblast migration from the peri-wound collagenous stroma into the fibrin-laden wound is critical for granulation tissue formation and subsequent healing. Previously we found that fibroblast transmigration from a collagen matrix into a fibrin matrix required fibronectin (FN). Integrins alpha4beta1, alpha5beta1, and alphavbeta3 and dermatan sulfate CD44 were required for this invasive migration. Here we demonstrated that syndecan-4, a transmembrane heparan sulfate (HS) proteoglycan, known to bind FN, is also required for fibroblast invasive migration of a fibrin/FN gel. This conclusion was based on fibroblast migration using two independent means of disrupting syndecan-4: heparinase degradation of HS glycosaminoglycans or suppression of syndecan-4 core protein with antisense oligodeoxynucleotides. Isolated syndecan-4 from these fibroblasts bound Hep II recombinant constructs FN III12-V15>FN III12-15>FN III12-14 but did not bind the IIICS (V) domain. Furthermore, platelet-derived growth factor (PDGF), which is required to stimulate fibroblast migration, markedly increased cell levels of syndecan-4 core protein in a time and concentration-dependent fashion. PDGF also induced upregulation of syndecan-4 at transcriptional level as determined by RT-PCR. These results demonstrate that syndecan-4 is essential for fibroblast invasive migration into fibrin clot and that PDGF, the stimulus for migration, induces increased syndecan-4 core protein expression.

Adult↗

PubMed2004

Good and poor mobilizing patients differ in mobilized CD34+ cell adhesion molecule profiles.

BACKGROUND: Alterations in expression of adhesion molecules are important in the trafficking of hematopoietic progenitors and probably in the mobilization process. Relatively little and conflicting data are currently available on the differences in expression between good and poor mobilizing patients. STUDY DESIGN AND METHODS: In this study, the expression of eight adhesion molecules on the collected CD34+ cells from 36 patients undergoing mobilization was determined. RESULTS: Good mobilizing patients, defined as those who collected their target in one apheresis procedure, had significantly fewer cells that expressed CD11a (LFA-1) and CD54 (ICAM-1) and borderline fewer that expressed CD11c, CD49d (VLA-4), and CD49d (VLA-5). No differences were detected in CD11b (Mac-1), CD15s (sLe(x)), or CD62L (L-selectin). Linear regression analysis identified number of prior chemotherapy courses and expression of CD11a (LFA-1) as independent predictive factors for mobilization efficiency. Good and poor mobilizing patients had approximately the same number of total CD34+ cells collected and little difference in times to engraftment. CONCLUSIONS: CD11a (LFA-1) expression inversely correlates with mobilization efficiency. Elucidation of the mechanism(s) underlying these observations will require further study.

Adolescent↗

PubMed2005

Arginine-glycine-aspartic acid (RGD)-containing peptides inhibit the force production of mouse papillary muscle bundles via alpha 5 beta 1 integrin.

Integrins are considered to be an important mechanosensor in cardiac myocytes. To test whether integrins can influence cardiac contractile function, the force-frequency relationships of mouse papillary muscle bundles were measured in the presence or absence of a synthetic integrin-binding peptide, GRGDNP (gly-arg-gly-asp-asn-pro). Results demonstrate that in the presence of an arginine-glycine-aspartic acid (RGD)-containing synthetic peptide, contractile force was depressed significantly by, 28% at 4 Hz, 37.7% at 5 Hz and 20% at 10 Hz (n = 6, P < 0.01). Treatment of myofibres with either protease-generated fragments of denatured collagen (Type I) or denatured collagen that contain the RGD motif, also reduced force production significantly. An integrin-activating antibody for beta(1) integrin inhibited the force similar to synthetic RGD peptide. Function-blocking integrin antibodies for alpha(5) and beta(1) integrins reversed the effect of the RGD-containing peptide, and alpha(5) integrin also reversed the effect of proteolytic fragments of denatured collagen on contractile force, whereas experiments with function-blocking antibody for beta(3) integrin did not reverse the effect of RGD peptide. Force-[Ca(2)(+)](i) measurements showed that the depressed rate of force generation observed in the presence of the RGD-containing peptide was associated with reduced [Ca(2)(+)](i). Data analyses further demonstrated that force per unit of Ca(2)(+) was reduced, suggesting that the myofilament activation process was altered. In addition, inhibition of PKC enzyme using the selective, cell-permeable inhibitor Ro-32-0432, reversed the activity of RGD peptide on papillary muscle bundles. In conclusion, these data indicate that RGD peptide, acting via alpha(5)beta(1) integrin, depresses the force production from papillary muscle bundles, partly associated with changes in [Ca(2)(+)](i) and the myofilament activation processes, that is modulated by PKCepsilon.

Animals↗

PubMed2003

Fibronectin bound to the surface of Staphylococcus aureus induces association of very late antigen 5 and intracellular signaling factors with macrophage cytoskeleton.

Staphylococcus aureus Cowan I and a clinically isolated coagulase-negative Staphylococcus strain, S. saprophyticus 10312, were found to have two fibronectin binding proteins, FnBPA and FnBPB. While both staphylococci bound to serum fibronectin to a similar extent, fibronectin binding significantly increased the phagocytic activity of macrophages against S. aureus (by ca. 150%) but not against S. saprophyticus. This enhancing effect of fibronectin was inhibited by an RGD sequence-containing peptide and also by anti-very late antigen 5 antibody. This suggests that the effect is mediated by very late antigen 5 expressed on macrophages. In macrophages ingesting fibronectin-bound Cowan I, alpha(5) and beta(1) chains were associated with the cytoskeleton. Cytosolic signaling factors such as paxillin, c-Src, and c-Csk were also associated with the cytoskeleton. On the contrary, beta(3) integrin transiently disappeared from the cytoskeleton when macrophages ingested the fibronectin-treated S. aureus Cowan I. Furthermore, the Src kinase family tyrosine kinase Lyn dissociated from the cytoskeleton. These cellular components did not respond in a fibronectin-dependent manner when macrophages phagocytosed S. saprophyticus. This means that only fibronectin-treated S. aureus Cowan I induces the accumulation of very late antigen 5, which in turn induces the association of paxillin and tyrosine kinases. It is thought that the phagocytic activity of macrophages against fibronectin-treated S. aureus was increased by signaling via the activation of very late antigen 5.

Animals↗

PubMed2004

Role of cell-cell communication in inhibiting butyric acid-induced T-cell apoptosis.

We have previously demonstrated that human gingival fibroblasts rescue butyric acid-induced T-cell apoptosis via proinflammatory cytokines such as interleukin 6 (IL-6) and IL-11, which are produced by fibroblasts stimulated with butyric acid. In this study, we determined if T-cell adhesion to human gingival fibroblasts influenced the susceptibility of T cells to butyric acid-induced apoptosis. We have shown that the number of Jurkat T cells adherent to gingival fibroblasts (Gin-1 cells) was significantly increased by the addition of butyric acid. All Jurkat cells that adhered to Gin-1 cells remained viable, while the nonadherent Jurkat cells dropped into apoptosis. The increase in T-cell adhesion to fibroblasts was also observed when Jurkat cells, but not Gin-1 cells, were pretreated with butyric acid. The expression levels of CD44, very late antigen 2 (VLA-2) and VLA-5 but not of leukocyte function-associated antigen 1 (LFA-1) and VLA-4 on Jurkat cells were increased following treatment with butyric acid. Furthermore, pretreatment of butyric acid-sensitized Jurkat cells with monoclonal antibodies against CD44, VLA-2, and VLA-5, but not LFA-1 and VLA-4, followed by coculture with Gin-1 cells inhibited T-cell adhesion to fibroblasts and increased apoptosis of nonadherent T cells after coculture of gingival fibroblasts and Jurkat cells. These results indicate that T-cell adherence to fibroblasts is enhanced by butyric acid and that butyric acid-induced T-cell apoptosis is down-regulated by T-cell adhesion to gingival fibroblasts through an interaction with the adhesion molecules CD44, VLA-2, and VLA-5 expressed on T cells stimulated with butyric acid.

Antibodies, Monoclonal↗

PubMed2004

Mechanotransduction by integrin is essential for IL-6 secretion from endothelial cells in response to uniaxial continuous stretch.

We previously reported that uniaxial continuous stretch in human umbilical vein endothelial cells (HUVECs) induced interleukin-6 (IL-6) secretion via IkappaB kinase (IKK)/nuclear factor-kappaB (NF-kappaB) activation. The aim of the present study was to clarify the upstream signaling mechanism responsible for this phenomenon. Stretch-induced IKK activation and IL-6 secretion were inhibited by application of alpha(5)beta(1) integrin-inhibitory peptide (GRGDNP), phosphatidylinositol 3-kinase inhibitor (LY-294002), phospholipase C-gamma inhibitor (U-73122), or protein kinase C inhibitor (H7). Although depletion of intra- or extracellular Ca(2+) pool using thapsigargin (TG) or EGTA, respectively, showed little effect, a TG-EGTA mixture significantly inhibited stretch-induced IKK activation and IL-6 secretion. An increase in the intracellular Ca(2+) concentration ([Ca(2+)](i)) upon continuous stretch was observed even in the presence of TG, EGTA, or GRGDNP, but not in a solution containing the TG-EGTA mixture, indicating that both integrin activation and [Ca(2+)](i) rise are crucial factors for stretch-induced IKK activation and after IL-6 secretion in HUVECs. Furthermore, while PKC activity was inhibited by the TG-EGTA mixture, GRGDNP, LY-294002, or U-73122, PLC-gamma activity was retarded by GRGDNP or LY-294002. These results indicate that continuous stretch-induced IL-6 secretion in HUVECs depends on outside-in signaling via integrins followed by a PI3-K-PLC-gamma-PKC-IKK-NF-kappaB signaling cascade. Another crucial factor, [Ca(2+)](i) increase, may at least be required to activate PKC needed for NF-kappaB activation.

Calcium↗

PubMed2004

Lipopolysaccharide induces expression of fibronectin alpha 5 beta 1-integrin receptors in human monocytic cells in a protein kinase C-dependent fashion.

LPS is an outer-membrane glycolipid component of gram-negative bacteria known for its fervent ability to activate monocytic cells and for its potent proinflammatory capabilities. In addition, LPS triggers the release of cytokines and chemokines as well as cell-cell adhesion molecules. We postulate that LPS may also affect the expression of matrix-binding integrin receptors, thereby modulating cell-adhesive functions in monocytic cells. To test this hypothesis, we investigated the effects of LPS on the expression of the integrin alpha(5)beta(1), a fibronectin receptor, in a human monocytic cell line (U937) as well as in isolated human peripheral blood mononuclear cells (PBMCs). We found that LPS increased the expression of alpha(5)beta(1) receptors and enhanced the adherence of U937 cells and PBMCs to fibronectin-coated surfaces; this was blocked by anti-alpha(5)beta(1) antibodies. LPS increased alpha(5)-subunit mRNA accumulation in a dose- and time-dependent manner. The induction by LPS occurred, at least in part, at the level of gene transcription as indicated by experiments using alpha(5) intact and deletion promoter constructs. LPS-induced alpha(5) gene transcription was associated with rapid induction of conventional PKC-alpha protein and activity, was blocked by PKC inhibitors, and was mimicked by lipid A. Finally, we found that an anti-CD14 antibody was able to inhibit the LPS response. Overall, the data suggest that LPS stimulates alpha(5) gene transcription via CD14 and PKC-dependent signals to enhance the expression of functional alpha(5)beta(1) receptors in monocytic cells. This process may help stimulate monocytic cell activation and facilitate their migration into fibronectin-containing tissues during infection.

Cell Adhesion↗

PubMed2004

Integrin fibronectin receptors in matrix metalloproteinase-1-dependent invasion by breast cancer and mammary epithelial cells.

Integrins contribute to progression in many cancers, including breast cancer. For example, the interaction of alpha(5)beta(1) with plasma fibronectin causes the constitutive invasiveness of human prostate cancer cells. Inhibition of this process reduces tumorigenesis and prevents metastasis and recurrence. In this study, naturally serum-free basement membranes were used as invasion substrates. Immunoassays were used to compare the roles of alpha(5)beta(1) and alpha(4)beta(1) fibronectin receptors in regulating matrix metalloproteinase (MMP)-1-dependent invasion by human breast cancer and mammary epithelial cells. We found that a peptide consisting of fibronectin PHSRN sequence, Ac-PHSRN-NH(2), induces alpha(5)beta(1)-mediated invasion of basement membranes in vitro by human breast cancer and mammary epithelial cells. PHSRN-induced invasion requires interstitial collagenase MMP-1 activity and is suppressed by an equimolar concentration of a peptide consisting of the LDV sequence of the fibronectin connecting segment, Ac-LHGPEILDVPST-NH(2), in mammary epithelial cells, but not in breast cancer cells. This sequence interacts with alpha(4)beta(1), an integrin that is often down-regulated in breast cancer cells. Immunoblotting shows that the PHSRN peptide stimulates MMP-1 production by serum-free human breast cancer and mammary epithelial cells and that the LDV peptide represses PHSRN-stimulated MMP-1 production only in mammary epithelial cells. Furthermore, PHSRN stimulates MMP-1 activity in breast cancer cells and mammary epithelial cells with a time course that closely parallels invasion induction. Thus, down-regulation of surface alpha(4)beta(1) during oncogenic transformation may be crucial for establishment of the alpha(5)beta(1)-induced, MMP-1-dependent invasive phenotype of breast cancer cells.

Amino Acid Sequence↗

PubMed2005

Defective p38 mitogen-activated protein kinase signaling impairs chemotaxic but not proliferative responses to stromal-derived factor-1alpha in acute lymphoblastic leukemia.

The chemokine stromal-derived factor-1alpha (SDF-1alpha) regulates leukemic cell motility and proliferation; however, the importance of these functions in the growth and dissemination of leukemia is unclear. We examined SDF-1alpha-mediated responses of cells from 27 cases of acute lymphoblastic leukemia (ALL). Although cells from the majority of cases showed chemotactic and proliferative responses to SDF-1alpha, a subset of cases did not undergo chemotaxis in response to SDF-1alpha, while still demonstrating dependence on SDF-1alpha for proliferation in stroma-supported cultures. This chemotactic defect was associated with an absence of phosphorylation of p38 mitogen-activated protein kinase (MAPK) induced by SDF-1alpha, and of SDF-1alpha-induced augmentation of beta(1) integrin-mediated adhesion. Signaling through phosphoinositide 3-kinase and MEK was not affected. No correlation was observed between CXCR4 expression and chemotactic function, in vitro migration into bone marrow stromal layers, and engraftment of leukemic cells in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. This study suggests that signaling through p38 MAPK is required for ALL cell chemotaxis but not for proliferation, and that the loss of a chemotactic response to SDF-1alpha does not impede engraftment in NOD/SCID mice.

Adolescent↗

PubMed2002

VLA-2 and VLA-5 cell adhesion molecules expression in CD34+ cells from umbilical cord blood and from bone marrow.

BACKGROUND/AIMS: Umbilical cord blood contains a large number of early hematopoietic cells with high proliferating capacity, that has been used as an alternative to bone marrow transplantation. The aim of this study is to investigate the number of two cell adhesion molecules in cord blood and in bone marrow. METHODS: We investigated two integrins, named VLA-2 and VLA-5 (Very Late Appearing Antigen), expressed in the surface of CD34+ cells. The CD34+ cells, isolated with MACS CD34+ isolation kit, were labelled with the appropriate monoclonal antibodies. RESULTS: Cell adhesion molecules showed highly expressed in both cord blood and bone marrow CD34+ cells. CONCLUSION: There are no significant differences between the two sources of CD34+ populations.

Antigens, CD34↗

PubMed2003

Permissive effect of fibronectin on collagen gel contraction mediated by bovine trabecular meshwork cells.

PURPOSE: The effect of fibronectin on the contractility of trabecular meshwork (TM) cells was investigated. METHODS: The contractility of bovine TM cells was evaluated by culture of the cells in a collagen gel and measurement of the change in the diameter of the gel under various conditions. The formation of stress fibers and the localization of integrin alpha5 and beta1 chains (which together form a fibronectin receptor) in bovine TM cells were investigated by laser confocal microscopy of cells stained with phalloidin and antibodies to the integrin subunits. RESULTS: The addition of fibronectin to collagen gels containing bovine TM cells induced marked gel contraction in a time- and concentration-dependent manner. Cytochalasin D (an inhibitor of microfilament formation) and the peptide GRGDSP (Gly-Arg-Gly-Asp-Ser-Pro), a fibronectin receptor antagonist, each inhibited this effect of fibronectin, whereas nocodazole (an inhibitor of microtubule polymerization) and the control peptide GRGESP (Gly-Arg-Gly-Glu-Ser-Pro) did not. Furthermore, fibronectin induced the spreading of cells, the formation of actin stress fibers, and the expression of integrin alpha5 in the collagen gel-embedded TM cells. CONCLUSIONS: Fibronectin promotes collagen gel contraction mediated by bovine TM cells. Moreover, the formation of actin stress fibers and upregulation of integrin alpha5 appear to contribute to this permissive effect of fibronectin. The interaction of fibronectin with TM cells may thus be a determinant of the contractility of TM tissue.

Actins↗

PubMed2006

Expression of the gene encoding poly(ADP-ribose) polymerase-1 is modulated by fibronectin during corneal wound healing.

PURPOSE: Poly(ADP-ribose) polymerase (PARP)-1 is a nuclear enzyme essential in several cellular functions such as DNA repair, DNA transcription, carcinogenesis, and apoptosis. Expression of the PARP-1 gene is mainly dictated by the transcription factor Sp1. Fibronectin (FN), a component from the extracellular matrix transiently expressed at high levels during wound healing of the corneal epithelium, was reported to exert a positive influence on expression of the alpha5 integrin subunit gene promoter by altering the state of Sp1 phosphorylation, a process that depended on the activation of the ERK signaling pathway. The present study was undertaken to investigate whether PARP-1 gene expression might be similarly regulated by FN through the same signaling pathways and attempted to link expression of this gene to corneal wound healing in vitro. METHODS: Expression of PARP-1, Sp1/Sp3, ERK1/2, phospho-ERK1/2, P38 and phospho-P38 was monitored by Western blot in cultures of rabbit corneal epithelial cells (RCECs) grown on FN in the presence of inhibitors of the MAPK, PI3K, and P38 signaling pathways. Electrophoretic mobility shift assays (EMSAs) were conducted to assess the binding of Sp1 and Sp3 in nuclear extracts from RCECs grown on FN in the presence of inhibitors. Plasmids bearing the PARP-1 promoter fused to the CAT reporter gene were also transfected into RCECs grown under similar culture conditions to assess the influence of these inhibitors on PARP-1 promoter activity. RESULTS: Expression of PARP-1, Sp1, and Sp3 increased considerably in RCECs grown on FN and translated into increased binding of Sp1 and Sp3 to their DNA target sites. In addition, FN increased PARP-1 promoter activity in a cell-density-dependent manner. Inhibition of both the MAPK and the PI3K pathways entirely abolished these properties. CONCLUSIONS: PARP-1 gene expression was strongly activated by FN through alterations in the phosphorylation state of Sp1 and Sp3 that resulted from the activation of the MAPK and PI3K signaling pathways, thereby suggesting that PARP-1 may play a critical function during the highly proliferative phase that characterizes wound healing of the corneal epithelium.

Animals↗

PubMed2005

Exogenous administration of Substance P enhances wound healing in a novel skin-injury model.

Soft tissue injury accounts for approximately 44% of all wounds in both the military and civilian populations. Following injury to soft tissue, Substance P (SP) and other neuropeptides are released by cutaneous neurons and modulate the function of immunocompetent and inflammatory cells, as well as epithelial and endothelial cells. The interaction between these components of the nervous system and multiple target cells affecting cutaneous repair is of increasing interest. In this report, we describe the effects of SP on wound repair in a novel, laser-induced, skin-wound model. Gross and histologic examination of laser-induced injury revealed that exogenously administered SP affects wound healing via neurite outgrowth, in addition to adhesion molecule and neurokinin-1 receptor involvement in vivo. All SP effects were decreased by pretreatment with Spantide II, an SP antagonist. The elucidation of SP-mediating mechanisms is crucial to firmly establishing the involvement and interaction of the peripheral nervous system and the immune system in cutaneous repair. Findings presented here suggest that SP participates in the complex network of mediators involved in cutaneous inflammation and wound healing.

Animals↗

PubMed2002

A highly sensitive strategy for SCID-repopulating cell assay by direct injection of primitive human hematopoietic cells into NOD/SCID mice bone marrow.

To measure the ability of human hematopoietic stem cells (HSCs), the SCID-repopulating cell (SRC) assay has been widely used. Conventionally, human HSCs are transplanted into a nonobese diabetic/severe combined immunodeficient (NOD/SCID) mouse via a tail vein. However, those cells must go through various obstacles until they reach the mouse marrow environment, which could explain the generally low homing efficiency in this system. Thus, the capability of HSCs may not be studied accurately by this intravenous transplantation method. In our attempt to reveal actual SRC potential, ie, self-renewal and multilineage differentiation in recipient bone marrow, we introduced cells into mouse marrow directly (intrabone marrow [iBM]) to minimize the effect of factors that may interfere with the homing of HSCs and compared the results obtained by intravenous and iBM methods. When cord blood CD34(+)CD38(-) cells were transplanted in NOD/SCID mice by iBM, a 15-fold higher frequency of SRC, 1 in 44 CD34(+)CD38(-) cells, was achieved compared with 1 in 660 by the intravenous method. Furthermore, the iBM transplant showed high levels of engraftment in the secondary transplantation. Pretreatment of CD34(+) cells with antibodies that block either very late antigen 4 (VLA-4) or VLA-5 reduced engraftment partially, whereas blockage of both molecules resulted in complete inhibition of engraftment, which suggests that VLA-4 and VLA-5 are involved in different processes in engraftment or have complementary roles. Our results indicate that the iBM injection strategy is a more sensitive and direct way to measure the capability of human SRCs and is useful to investigate the interaction of HSCs and marrow environment in vivo.

Animals↗

PubMed2003

Genetic evidence for convergence of c-Kit- and alpha4 integrin-mediated signals on class IA PI-3kinase and the Rac pathway in regulating integrin-directed migration in mast cells.

Mast cells play a critical role in host defense against a number of pathogens. Increased mast cell infiltration has been described in allergic asthma, in rheumatoid arthritis, and during helminthes infection. Despite the importance of mast cells in allergic disease and defense against infection, little is known about the mechanisms by which mast cells migrate to various tissues under steady state conditions or during infection or inflammation. Here, we show that activation of c-Kit by its ligand, stem cell factor (SCF), cooperates with alpha4 integrin in inducing directed migration of mast cells on fibronectin. A reduction in migration and activation of a small G protein, Rac, was observed in mast cells derived from class IA phosphoinositide-3 kinase (PI-3kinase)-deficient mice in response to SCF stimulation and in mast cells expressing the dominant-negative Rac (RacN17), as well as in mast cells deficient in the hematopoietic-specific small G protein, Rac2. In addition, a PI-3kinase inhibitor inhibited alpha4- as well as SCF-induced migration in a dose-dependent fashion. In contrast, a mitogen-activated protein kinase (MAPK) inhibitor had little effect. Consistent with the pharmacologic results, abrogating the binding of the p85alpha subunit of class IA PI-3kinase to c-Kit also resulted in inhibition of SCF-induced migration on fibronectin. These genetic and biochemical data demonstrate that both c-Kit and alpha4 integrin signaling are linked to class IA PI-3kinase and Rac pathways and regulate integrin-directed (haptotactic) migration in mast cells.

Animals↗

PubMed2003

IgE alone stimulates mast cell adhesion to fibronectin via pathways similar to those used by IgE + antigen but distinct from those used by Steel factor.

We recently demonstrated that immunoglobulin E (IgE), in the absence of cross-linking agents, activates signaling pathways in healthy murine bone marrow-derived mast cells (BMMCs) and that this activation enhances BMMC survival, at least in part, via secretion of autocrine-acting cytokines. We report herein that IgE alone also triggers the adhesion of both BMMCs and connective tissue mast cells (CTMCs) to the connective tissue component, fibronectin (FN). This adhesion occurs to the same extent as that triggered by optimal levels of Steel factor (SF) or IgE + antigen (IgE + Ag) and is mediated by an increased avidity of the integrin very late antigen 5 (VLA-5). Moreover, this IgE-induced adhesion, which is prolonged compared with that elicited by SF or IgE + Ag, requires phosphatidylinositol 3-kinase (PI3K), phospholipase C gamma (PLCgamma), and extracellular calcium but not extracellular-regulated kinase (Erk) or p38. Interestingly, we found, using the calcium channel blocker, 2-APB (2-aminoethoxydiphenyl borate) and Lyn-/- BMMCs that both IgE- and IgE + Ag-induced adhesion to FN require extracellular calcium entry, whereas SF does not. Furthermore, our data suggest that FN acts synergistically with IgE to prolong intracellular phosphorylation events and to enhance IgE-induced inflammatory cytokine production and BMMC survival.

3T3 Cells↗

PubMed2004

The sphingosine 1-phosphate receptor agonist FTY720 supports CXCR4-dependent migration and bone marrow homing of human CD34+ progenitor cells.

The novel immunosuppressant FTY720 activates sphingosine 1-phosphate receptors (S1PRs) that affect responsiveness of lymphocytes to chemokines such as stromal cell-derived factor 1 (SDF-1), resulting in increased lymphocyte homing to secondary lymphoid organs. Since SDF-1 and its receptor CXCR4 are also involved in bone marrow (BM) homing of hematopoietic stem and progenitor cells (HPCs), we analyzed expression of S1PRs and the influence of FTY720 on SDF-1/CXCR4-mediated effects in human HPCs. By reverse transcriptase-polymerase chain reaction (RT-PCR), S1PRs were expressed in mobilized CD34+ HPCs, particularly in primitive CD34+/CD38- cells. Incubation of HPCs with FTY720 resulted in prolonged SDF-1-induced calcium mobilization and actin polymerization, and substantially increased SDF-1-dependent in vitro transendothelial migration, without affecting VLA-4, VLA-5, and CXCR4 expression. In nonobese diabetic-severe combined immunodeficient (NOD/SCID) mice, the number of CD34+/CD38- cells that homed to the BM after 18 hours was significantly raised by pretreatment of animals and cells with FTY720, tending to result in improved engraftment. In addition, in vitro growth of HPCs (week-5 cobblestone area-forming cells [CAFCs]) was 2.4-fold increased. We conclude that activation of S1PRs by FTY720 increases CXCR4 function in HPCs both in vitro and in vivo, supporting homing and proliferation of HPCs. In the hematopoietic microenvironment, S1PRs are involved in migration and maintenance of HPCs by modulating the effects of SDF-1.

Antigens, CD↗

PubMed2002

Identification of CD9 extracellular domains important in regulation of CHO cell adhesion to fibronectin and fibronectin pericellular matrix assembly.

CD9, a 24-kDa member of the tetraspanin family, influences cellular growth and development, activation, adhesion, and motility. Our investigation focuses on the hypothesis that the CD9 second extracellular loop (EC2) is important in modulating cell adhesive events. Using a Chinese hamster ovary (CHO) cell expression system, we previously reported that CD9 expression inhibited cell adhesion to fibronectin and fibronectin matrix assembly. For the first time, a functional epitope on CD9 EC2 that regulates these processes is described. Binding of mAb7, an EC2-specific anti-CD9 monoclonal antibody, reversed the CD9 inhibitory activity on CHO cell adhesion and fibronectin matrix assembly. This reversal of cell phenotype also was observed in CHO cells expressing CD9 EC2 truncations. Furthermore, our data showed that the EC2 sequence (173)LETFTVKSCPDAIKEVFDNK(192) was largely responsible for the CD9-mediated CHO cell phenotype. Two peptides, (135)K-V(172) (peptide 5b) and (168)P-I(185) (peptide 6a), selectively blocked mAb7 binding to soluble CD9 and to CD9 on intact cells. These active peptides reversed the influence of CD9 expression on CHO cell adhesion to fibronectin. In addition, confocal microscopy revealed that CD9 colocalized with the integrin alpha(5)beta(1) and cytoskeletal F-actin in punctate clusters on the cell surface, particularly at the cell margins. Immunoprecipitation studies confirmed CD9 association with beta(1) integrin. The cellular distribution and colocalization of focal adhesion kinase and alpha-actinin with cytoskeletal actin was also influenced by CD9 expression. Thus, CD9 may exhibit its effect by modulating the composition of adhesive complexes important in facilitating cell adhesion and matrix assembly.

Actins↗