PubMed Health⌕ Search

Biomedical subjects

T Eskeland

Publications and source records attributed to T Eskeland.

At least 19 recordsLinked to original sources
PubMed1995

Interferon-gamma inhibits endocytosis of soluble animated beta-1,3-D-glucan and neutral red in mouse peritoneal macrophages.

We previously demonstrated that soluble animated beta-1,3-D-glucan (AG) is internalized after binding to a specific beta-glucan receptor on macrophages. Internalization, but not binding, of AG is reduced when the macrophages are treated with IFN-gamma. Because our data indicated that AG is taken up by macrophages through beta-glucan receptor-mediated endocytosis, we wanted to characterize further the inhibitory effect of IFN-gamma on endocytosis. We compared the internalization of AG and neutral red (NR). NR is internalized by macrophages through fluid-phase endocytosis. AG and NR showed a similar influx/efflux pattern. The initial rate of accumulation was much larger for AG than for NR, however, probably because of the involvement of the beta-glucan receptor in the uptake of AG. Internalized AG was associated with membranes of the endocytic vesicles and formed characteristic rings on confocal laser scanning microscopy (CLSM) images. Both the influx and efflux of AG and NR was inhibited by treatment of macrophages with IFN-gamma. Phorbol myristate acetate (PMA) added to the cell cultures increased the accumulation of AG and NR and reversed the inhibitory effect of IFN-gamma. The effect of PMA was dependent on functionally intact microfilaments and microtubules. CLSM showed that the accumulated AG was localized mostly in small vesicles (size < 2 microns) in IFN-gamma-treated cells, in large and small vesicles in untreated cells, and mostly in large vesicles (size > 2 microns) in PMA-treated cells. In conclusion, IFN-gamma inhibits both the beta-glucan receptor-mediated endocytosis of AG and the fluid-phase endocytosis of NR, probably by inhibiting the formation of large vesicles.

Amination↗
PubMed1994

A novel immunomodulator soluble aminated beta-1,3-D-glucan: binding characteristics to mouse peritoneal macrophages.

We have previously reported that soluble aminated beta-1,3-D-glucan (AG), a potent immunomodulator, specifically inhibited binding and internalization of AG-coated microbeads (GDM) in mouse peritoneal macrophages. The present study was undertaken to determine parameters of AG binding to macrophages. For this purpose, AG was conjugated with tyraminyl cellobiose (TC), which can be radioiodinated. With this method the immunomodulator was labelled with a very high specific radioactivity, allowing sensitive measurements of binding. Maximal binding capacity was 0.33 micrograms [125I]TC-AG/10(6) cells. Binding was inhibited by TC-AG and AG, but not by mannose and mannan, showing that the receptor different from the mannose receptor was involved. Binding was reversible, with an initial association rate of 120 cpm/min, and a much faster initial dissociation rate of 680 cpm/min. Bound [125I]TC-AG was internalized. These findings suggest that both AG and GDM are bound and internalized via the same beta-glucan receptor in mouse peritoneal macrophages.

Adjuvants, Immunologic↗
PubMed1994

IFN-gamma inhibits internalization of soluble aminated beta-1,3-D-glucan by macrophages and thereby down-regulates the glucan induced release of TNF-alpha and IL-1 beta.

We have previously shown that soluble animated beta-1,3-D-glucan (AG) and glucan-derivatized microbeads (GDM) bind to the specific beta-glucan receptor on mouse peritoneal macrophages. Phagocytosis of GDM by macrophages is mediated through the beta-glucan receptor. IFN-gamma which increases macrophage phagocytic capacity, also increased the phagocytosis of GDM. In the present study we show that IFN-gamma inhibits internalization of AG in macrophages in a dose- and time-dependent manner. The inhibitory effect of IFN-gamma was neutralized by treatment of the macrophages with cycloheximide. These results were confirmed by confocal laser scanning microscopy which showed that IFN-gamma treated cells incorporated less fluorescein-labelled AG than did untreated cells. IFN-gamma did not change the macrophage-binding capacity for AG showing that the inhibitory effect of IFN-gamma is not caused by decreased number of beta-glucan receptors on the cells. The stimulatory effect of AG on IL-1 beta and TNF-alpha release from macrophages was reduced by pretreatment of the cells with IFN-gamma. We conclude that the uptake of AG and GDM in macrophages, both mediated through the beta-glucan receptor, are differently regulated by IFN-gamma. The reduced internalization of AG after IFN-gamma treatment of macrophages, is probably responsible for the down-regulation of IL-1 and TNF-alpha secretion.

Animals↗
PubMed1993

Cytokines and PGE2 modulate the phagocytic function of the beta-glucan receptor in macrophages.

Under serum-free conditions the beta-glucan receptor of mouse macrophages mediates phagocytosis of beta-1,3-D-glucan-coated microbeads (diameter 2 microns). IFN-gamma increases the phagocytic function of the beta-glucan receptor in a dose-dependent manner, giving the plateau level at 100 U/ml. Maximum activity appears 9 h after addition of IFN-gamma to the cells. The effect disappears within 24 h. The effect of IFN-gamma may be a result of augmented receptor synthesis since treatment with cycloheximide reduces the phagocytosis. IL-1 also increases the phagocytic function of the beta-glucan receptor giving a dose-dependent response and with the plateau level reached at 10 U/ml. Maximum activity is found 4 h after addition of IL-1 to macrophages. The effect disappears within 24 h. TNF does not alter the phagocytic function of the beta-glucan receptor, but TNF together with IL-1 prolongs the effect of IL-1. PGE2 reduces the phagocytic function of the beta-glucan receptor. Maximum reduction is achieved with 8 ng/ml. Time-course studies show the lowest phagocytic activity 9 h after addition of PGE2 to the cells.

Animals↗
PubMed1991

Phagocytosis of beta-1,3-D-glucan-derivatized microbeads by mouse peritoneal macrophages involves three different receptors.

Intraperitoneal injection of beta-1,3-D-glucan coupled to the surface of monodisperse methacrylate microbeads improves the resistance against bacterial infections in mice, while methacrylate microbeads alone do not. The effect of the glucan-derivatized microbeads (GDM) is considered to be mediated through peritoneal macrophages. We show that both GDM and the underivatized methacrylate microbeads (UDM) treated with normal serum were rapidly bound and phagocytized by mouse peritoneal macrophages in vitro. We found that both complement and fibronectin opsonized the beads and were responsible for the uptake. Treatment of microbeads with serum lacking fibronectin and complement activity still gave some uptake of GDM, but not uptake of UDM. The uptake of GDM was similar to the uptake of untreated GDM and was inhibited by pretreatment of macrophages with soluble beta-1,3-D-glucan. Our conclusion is that GDM and UDM intraperitoneally bind fibronectin and C3 through activation of the alternative pathway of complement. This leads to their phagocytosis by macrophages through fibronectin and complement receptors. GDM are also internalized via beta-glucan receptors. We present the hypothesis that the beta-glucan receptors on peritoneal macrophages account for the protective effect of GDM in intraperitoneal bacterial infections.

Animals↗
PubMed1989

Modulation of the expression of chondroitin sulfate proteoglycan in stimulated human monocytes.

Proteoglycan biosynthesis was studied in human monocytes and monocyte-derived macrophages (MDM) after exposure to typical activators of the monocyte/macrophage system: interferon-gamma (IFN-gamma), lipopolysaccharide (LPS), and phorbol 12-myristate 13-acetate (PMA). By morphological examination, both monocytes and MDM were stimulated by these activators. Treatment with IFN-gamma resulted in a slight decrease in the expression of [35S]chondroitin sulfate proteoglycan (CSPG) in both monocytes and MDM, whereas LPS treatment increased the [35S]CSPG expression 1.8 and 2.2 times, respectively. PMA, in contrast, decreased the CSPG expression 0.4 times in monocytes, whereas MDM were stimulated to increase the biosynthesis 1.9 times. An increase in the sulfate density of the chondroitin sulfate chains was evident following differentiation of monocytes into MDM due to the expression of disulfated disaccharide units of the chondroitin sulfate E type (CS-E). However, monocytes exposed to PMA did also express disaccharides of the chondroitin sulfate E type. Furthermore, the expression of CS-E in MDM was increased 2 times following PMA treatment. An inactive phorbol ester, phorbol 12,13-diacetate, did not affect the expression of CS-E in either monocytes or MDM when compared with control cultures, suggesting that protein kinase C-dependent signal pathways may be involved in the regulation of sulfation of CSPG. Exposure to LPS or IFN-gamma did not lead to any changes in the sulfation of the chondroitin sulfate chains.

Cells, Cultured↗
PubMed1987

Formation of the membrane attack complex of complement (MAC) on erythrocytes from monocyte-produced terminal complement components.

By using antibodies against C5, C6, C7, C8, and C9, we found that terminal complement components were deposited on IgM-coated sheep erythrocytes (EIgM) kept in serum-free endotoxin-stimulated monocyte cultures for 24 or 48 h. Monoclonal antibodies revealed C9 neoantigens on the EIgM. There was no specific binding of an anti-S protein antibody, which reacts with the SC5b-9 complex, to the EIgM. Controls were native sheep erythrocytes (E) treated similarly which, in contrast to EIgM, do not activate the classical pathway of complement. Cycloheximide (1.0 microgram/ml) in the cell cultures resulted in no specific binding of the anti-C9 antibodies to EIgM. A fraction of the EIgM was lysed during incubation with the monocytes. We conclude that the monocytes secrete C5, C6, C7, C8, and C9, which form the membrane attack complex of complement (C5b-9) on the EIgM.

Animals↗
PubMed1987

Mouse peritoneal macrophages cultured serum-free deposit complement on IgM-coated sheep erythrocytes in vitro.

We conclude that the macrophages during cultivation produce the complement components of the classical pathway of complement, deposit complement components on EIgM and then phagocytose these cells via complement receptors. The conclusion is based on the following: EIgM, an activator of the classical pathway, are ingested when cultured serum-free with mouse peritoneal macrophages. We found a significantly higher binding of labelled protein to EIgM than to E kept in macrophage cultures in the presence of tritiated leucine, showing that de novo synthesis of macrophage-derived protein with affinity to EIgM takes place. A fraction of the bound protein is C3b and iC3b, since anti-mouse C3 antibodies bound to the co-cultured EIgM. Cycloheximide or anti-Mac-1 in the cultures inhibited macrophage attachment and uptake of EIgM. The phagocyte uptake of EIgM coated with complement by serum pretreatment was not inhibited by cycloheximide. This shows that the phagocytosis of the EIgM is dependent on erythrocyte-bound complement proteins made by the macrophage.

Animals↗
PubMed1987

Human alveolar macrophages and monocytes generate the functional classical pathway of complement in vitro.

Binding of labelled protein to EIgM kept with macrophage or monocyte cultures with 3H-leucine under serum-free conditions, shows that de novo synthesis of protein with affinity to EIgM takes place. We find that monoclonal anti-C3c and anti-C3g antibodies and polyclonal anti-C4 and anti-C5 antibodies bind to such erythrocytes. This demonstrates that C4b, C3b and iC3b are deposited on the EIgM. Additional evidence for complement synthesis is the increase in binding of anti-C4 antibodies to EIgM when the incubation time was increased from 48 to 96 hours. Stimulation of the mononuclear phagocyte cultures with ET was necessary to obtain significant amounts of erythrocyte-bound complement proteins. From these results we conclude that the functional classical pathway of complement is produced in vitro by the monocytes and macrophages.

Cells, Cultured↗
PubMed1986

In vitro interactions of murine peritoneal macrophages and sarcoma cells. II. Induction of DNA synthesis in macrophages.

Unstimulated peritoneal macrophages grown in vitro for 5 days do not incorporate thymidine. Sarcoma (AA) cells introduced to the culture on the fifth day die and disintegrate whereas the macrophages become markedly stimulated and autoradiography shows that they are triggered to synthesize DNA. In syngeneic cocultures up to 28% of the macrophages incorporate thymidine after 5 days of coculture.

Animals↗
PubMed1986

Formation of the functional alternative pathway of complement by human monocytes in vitro as demonstrated by phagocytosis of agarose beads.

Native agarose beads (diameter 5-10 micron), activators of the alternative complement pathway, are slowly phagocytosed when incubated with human monocytes cultured under serum-free conditions. Agarose beads preincubated with monocyte cultures and then transferred to new cultures are more easily phagocytosed than native beads. These results indicate that the phagocytosis of agarose beads depends on opsonization of the beads by one or several substances of monocyte origin. By using antihuman C3 antibodies, trypsin treatment, and sodium dodecyl sulphate washing, we were able to demonstrate C3b and iC3b on the agarose beads. The molecules were covalently bound to the surface of the beads. We conclude that in vitro human monocytes produce and secrete the essential factors for activation and propagation of the alternative complement pathway (factors C3, B, D, H and I), which becomes evident with an external activator like agarose beads in the cultures. The activation of complement by agarose beads results in the attachment of C3b and iC3b to the surface of the beads, which are then phagocytosed by means of C3b and iC3b receptors on the monocytes.

Cell Communication↗
PubMed1986

Synthesis of complement components C5, C6, C7, C8 and C9 in vitro by human monocytes and assembly of the terminal complement complex.

Monocytes cultured under serum-free conditions secreted protein which bound covalently and non-covalently to agarose beads, an activator of the alternative pathway of complement. There was a significantly binding of monoclonal anti-C3c antibodies, polyclonal anti-C5, anti-C6, anti-C7, anti-C8, and anti-C9 antibodies, and of a monoclonal antibody against a neoantigen of polymerized C9 to agarose beads incubated with the monocytes for 24, 48, 72 or 96 h. From these results, we conclude that monocytes produce C5, C6, C7, C8 and C9 that assemble as the terminal complement complex on the surface of the agarose beads. Activation by agarose of the alternative pathway with generation of particle bound C3 and C5 convertases is a prerequisite for the subsequent formation of the terminal complement complex. Whether SC5b-9 or the membrane attack of complement (C5b-9) is formed on the beads will be examined.

Antibodies, Monoclonal↗
PubMed1986

Human cord blood monocytes: binding and ingesting capacity mediated by Fc and C3b receptors.

Human cord blood monocytes have been compared to monocytes from adults. Our results show that unstimulated cord blood monocytes are as effective as monocytes from adults with regard to their binding and ingesting capacity mediated by Fc and C3b receptors. Furthermore, when stimulated with PMA (phorbol-myristate-acetate), the C3b receptor-mediated phagocytosis in cord blood monocytes was enhanced to the same extent as in monocytes from adults. These findings show that Fc and C3b receptor functions in human monocytes are fully developed at birth and cannot explain the increased susceptibility to infections seen in the neonate.

Adult↗
PubMed1985

Fibronectin binds to complement-coated agarose beads and increases their association to mouse macrophages.

We have studied the binding of fibronectin to complement (C3b, C3bi, C3d)-coated agarose beads and its effect on cell association of such beads to mouse macrophages. Fibronectin bound to agarose beads preincubated in human serum, whereas no binding occurred after preincubation of the beads with complement-inactivated (50 degrees C for 20 min or ethylenediaminetetraacetic acid) sera. The binding of iodine-labelled fibronectin to beads preincubated in fibronectin-depleted serum (HS-FIB) was about twice that of beads preincubated in normal serum. Unlabelled fibronectin inhibited the following binding of labelled fibronectin to beads pretreated in HS-FIB. A similar amount of fibronectin bound to agarose beads coated with equimolar amounts of C3b, C3bi, or C3d, suggesting that the common domain C3d carries the main binding site(s) for fibronectin. Preincubation of serum-treated and trypsinized agarose beads with fibronectin led to an increased association (22%) of such beads to mouse macrophages. The results indicate that fibronectin promotes binding of complement-coated agarose beads to mouse macrophages, whereas the ingestion of the beads is mediated via complement C3 receptors.

Animals↗
PubMed1984

Phagocytosis by human monocytes of particles activating the alternative pathway of complement.

The phagocytosis of particles activating the alternative pathway of complement by human monocytes cultured under serum-free conditions was studied. In contrast to native zymosan particles, which were easily ingested, rabbit erythrocytes and agarose beads had to be coated with C3b or C3bi to be engulfed by the monocytes. The binding and ingestion by monocytes of particles coated with C3bi were greater than for the same particles coated with the equivalent amount of C3b. The binding and uptake of rabbit erythrocytes and agarose beads were proportional to the amount of C3b or C3bi on the particles. In contrast to the complement activator particles, C3b- and C3bi-coated sheep erythrocytes, which are non-activators, were not ingested by the monocytes, although attachment to the monocytes took place. The presence of methylamine or cobra venom factor, which are complement inhibitors, strongly reduced the ingestion of native zymosan by the monocytes, whereas the uptake of C3b- or C3bi-coated zymosan particles were only weakly affected. This suggests that the binding of native zymosan to monocytes is sensitive to interference from a cell-derived alternative pathway C3 convertase (C3bBb). Binding and uptake of activators by human monocytes via complement receptor(s) are discussed.

Animals↗
PubMed1983

Complement (C3) receptor-mediated attachment of agarose beads to mouse peritoneal macrophages and human monocytes.

We have determined the receptors on human monocytes and mouse peritoneal macrophages producing agarose binding. By using isolated human complement factors C3, B and D, agarose beads were coated with C3b. In some experiments C3b was converted to C3bi by using human serum diluted 1:20. Agarose beads coated with C3b or C3bi bound strongly to monocytes. Only agarose beads coated with C3bi were attached to mouse macrophages. Trypsinization of agarose beads coated with C3bi abolished the attachment of the beads to macrophages and monocytes, probably because of conversion of C3bi to C3d. Endocytosis by macrophages of agarose preincubated in human serum or in C5-deficient AKR mouse serum reached the same levels, indicating that the amount of C5 present in serum during preincubation is not important for the degree of endocytosis. It is concluded that internalization of agarose by macrophages is mediated via the C3bi receptor.

Animals↗