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C G Gahmberg

Publications and source records attributed to C G Gahmberg.

At least 19 recordsLinked to original sources
PubMed1995

Intercellular adhesion molecule-2 (CD102) binds to the leukocyte integrin CD11b/CD18 through the A domain.

The interactions between the leukocyte-specific beta 2-integrins cluster of differentiation (CD) Ag CD11/CD18 and their ligands, the intercellular adhesion molecules (ICAMs), play important roles in many adhesion-dependent leukocyte functions. ICAM-1 is known to be a ligand for both CD11a/CD18 and CD11b/CD18. ICAM-2, whose two extracellular Ig domains show the highest homology to the two NH2-terminal domains of ICAM-1, has been previously shown to be a ligand for CD11a/CD18. We recently found that a 22-amino acid CD11a/CD18-binding peptide, P1, derived from the first domain of ICAM-2, also binds to purified CD11b/CD18. In the present study, we demonstrate that the ICAM-2 protein interacts with CD11b/CD18, and the binding is through the CD11b A domain.

Animals↗
PubMed1995

Treatment with okadaic acid reveals strong threonine phosphorylation of CD18 after activation of CD11/CD18 leukocyte integrins with phorbol esters or CD3 antibodies.

The CD11/CD18 leukocyte integrins comprise three heterodimers involved in leukocyte adhesion. CD11/CD18 avidity may be regulated intracellularly, and the CD18 polypeptide has previously been shown to become phosphorylated in leukocytes after phorbol ester stimulation. The importance of phosphorylation in the regulation of CD11/CD18 avidity has, however, remained unclear. We have now activated T cells using phorbol esters, CD3, and CD44 Abs. Both phorbol ester and CD3 treatment activated protein kinase C. CD18 was shown to become more stably phosphorylated after phorbol ester treatment and more transiently so after CD3 stimulation. The phosphorylation was strongly augmented by okadaic acid, a serine/threonine phosphatase inhibitor. While phorbol ester treatment caused phosphorylation mainly on serine, in okadaic acid-pretreated cells, both phorbol ester treatment as well as CD3 stimulation revealed strong threonine phosphorylation. Since earlier mutational studies have demonstrated the functional importance of cytoplasmic threonine residues in CD18, the threonine phosphorylation reported here indicates the role of threonine phosphorylation in the regulation of CD11/CD18 avidity.

Alkaloids↗
PubMed1995

Mutation of the cytoplasmic domain of the integrin beta 3 subunit. Differential effects on cell spreading, recruitment to adhesion plaques, endocytosis, and phagocytosis.

The cytoplasmic domain of the beta subunit of the alpha IIb beta 3 integrin is required for cell spreading on fibrinogen. Here we report that deletion of six amino acids from the COOH terminus of the beta 3 (I757TYRGT) totally abolished cell spreading and formation of adhesion plaques, whereas retaining Ile757 partially preserved these functions. We further found that substitution of Tyr747 with Ala also abolished alpha IIb beta a-mediated cell spreading. The effects of these and other mutations on additional functions of alpha IIb beta 3 were also studied. Progressive truncations of beta 3, in which stop codons were inserted at amino acid positions 759-756, caused partial defects in the recruitment of alpha IIb beta 3 to preestablished adhesion plaques and a gradual decrease in the ability of alpha IIb beta 3 to mediate internalization of fibrinogen-coated particles. The Tyr747-->Ala substitution mutant was almost totally inactive in both of these assays. Point mutations at Tyr759, and at a conserved area close to the transmembrane domain of beta 3, decreased integrin recruitment to preestablished adhesion plaques but allowed alpha IIb beta 3-mediated formation of these structures and partial cell spreading. Deletion of the cytoplasmic domain of beta 3 did not affect the constitutive endocytosis of alpha IIb beta 3.

Amino Acid Sequence↗
PubMed1995

Activation of natural killer cell migration by leukocyte integrin-binding peptide from intracellular adhesion molecule-2 (ICAM-2).

Intracellular adhesion molecule-2 (ICAM-2), one of the ligands of CD11a/CD18 (LFA-1), is mainly expressed on endothelial and hematopoietic cells. The biological significance of ICAM-2 has remained unclear. Previous findings have shown that a peptide from ICAM-2, spanning residues 21-42 from the first immunoglobulin domain, enhances natural killer (NK) cell cytotoxicity and induces T cell aggregation. We have now studied the effect of the same ICAM-2 peptide on NK cell migration in the Boyden chamber assay. The peptide significantly increased NK cell migration up to 215 +/- 21%, as compared to migration of control cells (100%), and the induction was inhibited by anti-CD11a monoclonal antibodies. The ICAM-2 peptide also induced polymerization of F-actin at the leading edge of migratory NK cells. Cross-linking of CD11a/CD18 receptors with anti-CD11a or anti-CD18 monoclonal antibodies and secondary antibodies resulted in receptor recycling, increased migration, and actin polymerization, but led to slight inhibition of cytotoxicity. The ICAM-2 peptide did not induce such a receptor recycling. Phosphotyrosine immunoblotting experiments showed that the ICAM-2 peptide increased the phosphorylation of 150- and 35-kDa proteins. During cross-linking with antibodies, only the 150-kDa protein showed increased phosphorylation. The results show that depending on the type of CD11a/CD18 receptor ligation different kinds of signals are transduced in NK cells. These signals may either trigger only locomotion, or both locomotion and cytotoxicity. Based on these findings, a major function for ICAM-2 on endothelium may be triggering of migration of adhering leukocytes.

Actins↗
PubMed1995

Expression and characterization of a B cell growth promoting polypeptide derived from the 12 kDa B cell growth factor gene (BCGF 1).

The expression and partial purification of recombinant 12 kDa B cell growth factor are reported. The polypeptide was derived from the genomic sequence of the gene (BCGF 1) which is here shown to be a single copy gene that localizes to human chromosome 16. When expressed as a glutathione S-transferase fusion protein in E. coli, the protein appears as a 38 kDa polypeptide in Western blot analysis using a peptide antibody. The purified fusion protein stimulates the proliferation of activated human B cells in a dose-dependent manner, and the active site resides within the 104 carboxy-terminal amino acids. The availability of biologically active recombinant 12 kDa B cell growth factor will enable its evaluation in B cell growth regulation, and provides a new means of in vitro culturing of human B lymphocytes.

Amino Acid Sequence↗
PubMed1995

A CD44 monoclonal antibody differentially regulates CD11a/CD18 binding to intercellular adhesion molecules CD54, CD102 and CD50.

We have made a monoclonal anti-CD44 antibody which is able to activate the leukocyte integrin CD11a/CD18. Activated T cells strongly aggregated, and the aggregation was shown to be intercellular adhesion molecule (ICAM)-1 (CD54) and ICAM-2 (CD102) dependent. Using purified ICAM coated on plastic, only binding to ICAM-1 was increased by the CD44 antibody, whereas activation by phorbol ester increased binding to both ICAM-1 and ICAM-3. The binding to ICAM-2 was not affected by either treatment. These findings show that the CD11a/CD18 integrin can be activated in a ligand-specific manner by engagement of CD44.

Antibodies, Monoclonal↗
PubMed1995

A peptide derived from the intercellular adhesion molecule-2 regulates the avidity of the leukocyte integrins CD11b/CD18 and CD11c/CD18.

beta 2 integrin (CD11a,b,c/CD18)-mediated cell adhesion is required for many leukocyte functions. Under normal circumstances, the integrins are nonadhesive, and become adhesive for their cell surface ligands, the intercellular adhesion molecules (ICAMs), or soluble ligands such as fibrinogen and iC3b, when leukocytes are activated. Recently, we defined a peptide derived from ICAM-2, which specifically binds to purified CD11a/CD18. Furthermore, this peptide strongly induces T cell aggregation mainly mediated by CD11a/CD18-ICAM-1 interaction, and natural killer cell cytotoxicity. In the present study, we show that the same ICAM-2 peptide also avidly binds to purified CD11b/CD18, but not to CD11c/CD18. This binding can be blocked by the CD11b antibody OKM10. The peptide strongly stimulates CD11b/CD18-ICAM-1-mediated cell aggregations of the monocytic cell lines THP-1 and U937. The aggregations are energy and divalent cation-dependent. The ICAM-2 peptide also induces CD11b/CD18 and CD11c/CD18-mediated binding of THP-1 cells to fibrinogen and iC3b coated on plastic. These findings indicate that in addition to induction of CD11a/CD18-mediated cell adhesion, the ICAM-2 peptide may also serve as a "trigger" for high avidity ligand binding of other beta 2 integrins.

Actin Cytoskeleton↗
PubMed1994

Sialyl Lewis(x)- and L-selectin-dependent site-specific lymphocyte extravasation into renal transplants during acute rejection.

Kidney allograft rejection is an inflammatory process dominated by lymphocytes. During rejection lymphocytes preferentially adhere to the peritubular capillary endothelium (PTCE), which acquires morphological features common to high endothelium. These observations indicate that PTCE is the site of lymphocyte entry into the rejecting renal allograft. Of the identified endothelial adhesion molecules, ICAM-1 was already expressed on the endothelium of normal kidneys, and its expression was strongly enhanced during rejection without site-specific restriction. VCAM-1 was not expressed on the endothelium of normal or syngeneic kidneys, but its expression was induced during allograft rejection not only in PTCE, but occasionally also on the endothelium of larger vessels. Sialyl Lewisx (sLex) showed a very restricted pattern of expression; endothelium was sLex-negative both in control and syngeneic kidneys. On the other hand, PTCE reacted strongly with anti-sLex antibody in allografts. When kidney frozen sections were treated with sialidase the binding of lymphocytes decreased by 70%. Low-dose chymotrypsin treatment of lymphocytes, known to remove L-selectin from the lymphocyte surface, decreased their binding to PTCE by 60%. Likewise lymphocyte adhesion to PTCE was inhibited by 70% by anti-sLex- and anti-L-selectin-antibodies and by sLex tetrasaccharide. Finally PTCE in the allografts, but not in syngeneic grafts or normal kidneys, bound an L-selectin-IgG fusion protein, indicating that ligands for L-selectin were induced during rejection.

Animals↗
PubMed1994

Tyrosine phosphorylation of CD45 phosphotyrosine phosphatase by p50csk kinase creates a binding site for p56lck tyrosine kinase and activates the phosphatase.

Src family protein tyrosine kinases (PTKs) play an essential role in antigen receptor-initiated lymphocyte activation. Their activity is largely regulated by a negative regulatory tyrosine which is a substrate for the activating action of the CD45 phosphotyrosine phosphatase (PTPase) or, conversely, the suppressing action of the cytosolic p50csk PTK. Here we report that CD45 was phosphorylated by p50csk on two tyrosine residues, one of them identified as Tyr-1193. This residue was not phosphorylated by T-cell PTKs p56lck and p59fyn. Tyr-1193 was phosphorylated in intact T cells, and phosphorylation increased upon treatment with PTPase inhibitors, indicating that this tyrosine is a target for a constitutively active PTK. Cotransfection of CD45 and csk into COS-1 cells caused tyrosine phosphorylation of CD45 in the intact cells. Tyrosine-phosphorylated CD45 bound p56lck through the SH2 domain of the kinase. Finally, p50csk-mediated phosphorylation of CD45 caused a severalfold increase in its PTPase activity. Our results show that direct tyrosine phosphorylation of CD45 can affect its activity and association with Src family PTKs and that this phosphorylation could be mediated by p50csk. If this is also true in the intact cells, it adds a new dimension to the physiological function of p50csk in T lymphocytes.

Amino Acid Sequence↗
PubMed1993

Structural study of the sugar chains of human leukocyte common antigen CD45.

The leukocyte cell surface glycoprotein CD45 is a protein tyrosine phosphatase and is involved in signal transduction mediated by the T cell antigen receptor. The asparagine-linked sugar chains were released as oligosaccharides from purified CD45 by hydrazinolysis. Approximately 6 mol of sugar chains was released from 1 mol of CD45. These sugar chains were converted to radioactive oligosaccharides by reduction with NaB3H4 and separated into neutral and acidic fractions by paper electrophoresis. All of the acidic oligosaccharides were converted to neutral ones by digestion with sialidase, indicating that they are sialyl derivatives. Binding of the sialylated oligosaccharides to an SNA-agarose column as well as methylation analysis revealed that the oligosaccharides have only alpha-2,6-linked sialic acid residues. The neutral and sialidase-treated acidic oligosaccharides were fractionated by serial lectin column chromatography followed by Bio-Gel P-4 column chromatography. Structural studies of each oligosaccharide by sequential exo- and endoglycosidase digestion and by methylation analysis revealed that CD45 contains mainly bi-, tri-, and tetraantennary complex-type sugar chains. About 46% of the tetraantennary complex-type sugar chains had the poly(N-acetyllactosamine) groups and 18% of the 2,4-branched triantennary complex-type sugar chains had the fucosyl N-acetyllactosamine group. A portion of the bi- and 2,4-branched triantennary complex-type sugar chains were bisected. In addition to these sugar chains, a small amount of high mannose-type and hybrid-type sugar chains were detected.

Asparagine↗
PubMed1993

A leukocyte integrin binding peptide from intercellular adhesion molecule-2 stimulates T cell adhesion and natural killer cell activity.

Adhesion is of pivotal importance for a number of leukocyte functions. Little is known about the binding between leukocyte integrins and the intercellular adhesion molecules (ICAMs). Normally integrins are nonadhesive, and require a stimulus to become active. We have now identified a peptide from ICAM-2, which binds to leukocyte integrins and activates adhesion. Furthermore, the peptide strongly increased the binding and cytotoxicity of natural killer cells. These findings show that adhesion-dependent leukocyte functions can be activated by ligand-derived peptides, and therefore provide evidence that the avidity of leukocyte integrins is up-regulated by integrin-ligand interactions.

Alkaloids↗
PubMed1993

A peptide from ICAM-2 binds to the leukocyte integrin CD11a/CD18 and inhibits endothelial cell adhesion.

Numerous leukocyte functions depend on adhesive intercellular interactions. The leukocyte-specific integrins CD11a/CD18 (lymphocyte function-associated antigen-1 (LFA-1)) and CD11b/CD18 (complement type 3 receptor (Mac-1)), which bind to the intercellular adhesion molecules ICAM-1 and ICAM-2, play a key role in adhesion. Little is known about the binding in molecular detail. We have now defined a peptide region from the first immunoglobulin domain of ICAM-2 that is specifically involved in binding to CD11a/CD18. A synthetic peptide from this part of ICAM-2, covering residues 21-42, bound to purified CD11a/CD18 and inhibited the adhesion of endothelial cells to this integrin. It also inhibited the binding of B lymphoblastoid cells to endothelial cells. Leukocytes bound to the peptide coated on plastic. Several shorter peptides from the same region showed less or no activity.

Amino Acid Sequence↗
PubMed1993

The leukocyte surface antigens CD11b and CD18 mediate the oxidative burst activation of human peritoneal macrophages induced by type 1 fimbriated Escherichia coli.

Analysis by immunofluorescence-activated cell sorting of human peritoneal macrophages from patients undergoing intermittent peritoneal dialysis revealed that they express the CD11/CD18 surface antigens, with CD11b and CD18 as the predominant ones. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of immunoprecipitates obtained from lysates of 125I-labeled macrophages with rabbit polyclonal antibodies against the CD11a-c/CD18 complex or against CD18, revealed four radioactive bands corresponding to CD11a, CD11b, CD11c, and CD18. Monoclonal antibodies against CD11b and CD18 inhibited by 80 and 90%, respectively, the oxidative burst activation of the macrophages by type 1 fimbriated Escherichia coli, whereas monoclonal antibodies against CD11a, CD11c, and CD43 were without effect. Our results suggest that CD11b and CD18 (receptors for C3bi) serve also as receptors for mannose-specific E. coli on human peritoneal macrophages and may be involved in the lectinophagocytosis of the bacteria by these cells.

Antibodies, Monoclonal↗
PubMed1993

The vascular E-selectin binds to the leukocyte integrins CD11/CD18.

Leukocyte adhesion involves at least three molecular families of adhesion proteins: the leukocyte integrins CD11/CD18, the intercellular adhesion molecules (ICAMs) and the carbohydrate-binding L-, E- and P-selectins. The intercellular adhesion molecules are well-known ligands for the CD11/CD18 integrins. We now show that E-selectin specifically binds to the sialyl Lex carbohydrate epitopes of leukocyte integrins. Thus, the different families of leukocyte adhesion molecules form an integrated adhesion network.

Antigens, CD↗
PubMed1992

Regulated Ca2+ signalling through leukocyte CD11b/CD18 integrin.

General mechanisms of adhesion in the immune response are coordinated by the leukocyte integrins CD11/CD18. The possible participation of these differentiation molecules in early events of transmembrane signalling was investigated. Monoclonal antibody (mAb) cross-linking of CD18, the integrin beta subunit ubiquitously expressed by all leukocytes, increased the cytosolic free Ca2+ concentration ([Ca2+]i) by 2-3-fold in monocyte THP-1 cells. Digitalized imaging in single adherent cells showed that this Ca2+ response is temporally biphasic, involves both release of Ca2+ from the intracellular stores as well as Ca2+ influx from the external compartment, and is dramatically down-modulated by terminal differentiation of THP-1 cells to a mature monocyte phenotype. Similarly, only a minor subset of 20-30% of peripheral blood monocytes heterogeneously maintain the CD18-mediated Ca(2+)-signalling properties. Cross-linking of CD18 also increased cytosolic free [Ca2+]i in a subset of approx. 15-20% of resting T lymphocytes, in a Ca2+ response that was completely abrogated during T-cell mitogenic activation with lectins or alloreactive antigen. While cross-linking of CD11a or CD11c was without effect, occupancy of CD11b increased cytosolic free [Ca2+]i in monocytic cells. This response was functionally coupled with a transient activation state of CD11b/CD18, which was reflected in its increased avidity to bind the complementary ligand fibrinogen. These results suggest that occupancy of CD18 transduces a stimulatory Ca2+ signal that is critically regulated by the state of cell activation/differentiation and by the association with the unique alpha-subunit CD11b. These intrinsic signalling properties may directly participate in regulating the oligospecific ligand recognition of leukocyte integrins.

Antibodies, Monoclonal↗
PubMed1992

Migration of recombinant IL-2-activated T and natural killer cells in the intercellular space of human H-2 glioma spheroids in vitro. A study on adhesion molecules involved.

The migration of rIL-2-activated T and NK cells into the intercellular space of glioma tissue was studied using multicellular spheroids grown from the human H-2 glioblastoma cell line as targets. Lymphocytes of all analyzed subtypes migrated into the spheroids, but CD56+ cells were particularly migratory. Lymphocytes and the H-2 tissue expressed adhesion molecule subunits for the following potential cell-cell or cell-matrix interactions: alpha 3 beta 1 (VLA-3) to fibronectin, laminin, and collagen; alpha 4 beta 1 (VLA-4) and alpha 5 beta 1 (VLA-5) to fibronectin; alpha 6 beta 1 (VLA-6) to laminin; alpha 4 beta 1 to VCAM-1; alpha L beta 2 (Leu-CAMa/LFA-1) to CD54 (ICAM-1); CD44 to fibronectin, collagen, laminin, hyaluronate; CD2 to CD58 (LFA-3); and CD56 (N-CAM) to CD56. In the H-2 tissue, CD54 and VCAM-1 were expressed as a gradient. The expression of CD54 was weak in the peripheral zone and the expression was stronger in the quiescent deeper zone, whereas the distribution of VCAM-1 showed an inversed pattern. The low expression of CD54 was up-regulated along the frontier of migrating lymphocytes. The migration was almost totally prevented by the anti-CD18 (beta 2) mAb IB4 and TS1/18, and also strongly inhibited by the anti-CD54 mAb LB-2. Instead, mAb known to inhibit the binding of beta 1 integrins to fibronectin were not significantly inhibitory. However, a combination of the GPEILDVPST and GRGDS peptides, which compete for the binding of alpha 4 beta 1 and alpha 5 beta 1 to fibronectin and may also affect other adhesion systems, partially prevented migration.

Amino Acid Sequence↗
PubMed1992

Regulation of the p59fyn protein tyrosine kinase by the CD45 phosphotyrosine phosphatase.

Triggering of the T cell antigen receptor/CD3 (TcR/CD3) complex leads to rapid tyrosine phosphorylation of regulatory proteins that participate in initiating T cell activation and proliferation. This signal transduction event requires the presence of the TcR/CD3-associated protein tyrosine kinase p59fyn. There is also evidence that the CD45 phosphotyrosine phosphatase is involved in TcR/CD3 signalling. We show here by capping experiments using double indirect immunofluorescence techniques that the receptor phosphotyrosine phosphatase CD45 and the intracellular protein tyrosine kinase p59fyn specifically co-distribute in functional T lymphocytes. Furthermore, we provide evidence that isolated p59fyn is a substrate for CD45 as indicated by the rapid dephosphorylation of the regulatory Tyr531 of p59fyn by CD45. This dephosphorylation is accompanied by a severalfold increase in the catalytic activity of p59fyn as measured by its autophosphorylation and phosphorylation of an exogenous substrate. We also demonstrate that CD45-mediated dephosphorylation and activation of p59fyn apparently occurs at a slow basal rate in resting T cells. This represents the first identification of a physiologic regulator of p59fyn and implies a mechanism for the role of CD45 in TcR/CD3 signal transduction.

Amino Acid Sequence↗