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1.
In this article, we describe a-chicken T helper-activated cell line, III-C5, which is non-antigen-specific but IL-2-dependent. By virtue of its absolute requirement for IL-2, the III-C5 cell line is a useful tool for quantifying IL-2 production by any chicken-activated T lymphocytes in culture. The III-C5 line will be used to quantify IL-2 production in mixed lymphocyte reaction in vitro, in order to study the functional activity of T lymphocytes from avian chimeras constructed in our laboratory and particularly for studying their state of tolerance. 相似文献
2.
O J Semmes M S Sztein J M Bailey W D Merritt 《International journal of immunopharmacology》1992,14(4):583-593
Murine interleukin-2-dependent T-lymphocytes (CT6) were treated with tunicamycin, an inhibitor of both glycoprotein and ganglioside synthesis, to study the involvement of glycosylation in the IL-2 proliferative response. Tunicamycin inhibited proliferation in a dose-dependent manner at concentrations which did not inhibit protein synthesis (10-50 ng/ml). Swainsonine, a glycoprotein processing inhibitor, had no effect on proliferation. Inhibition of proliferation by tunicamycin was accompanied by an inhibition of binding of 125I-IL-2 to its high-affinity receptor. Scatchard analysis showed that receptor number was decreased by tunicamycin treatment. On the other hand, tunicamycin did not affect either the binding of the monoclonal antibody 7D4, specific for the 55 kDa low-affinity protein subunit of the IL-2 receptor, or the recycling of the IL-2 receptor. To determine the specific effects of tunicamycin on the biosynthesis of particular CT6 glycoconjugates, cells were radiolabeled with 3H-glucosamine and incorporation into ganglioside, neutral glycolipid and glycoprotein fractions was measured. Low doses of tunicamycin inhibited ganglioside synthesis and glycoprotein glycosylation to the same extent, whereas no effect on neutral glycolipid synthesis was observed. These results suggest that glycosylation of glycoprotein and/or gangliosides might play an important role in the formation of a functional high-affinity IL-2 receptor complex in CT6 cells. 相似文献
3.
Since interleukin-2 (IL-2) and IL-4 act in concert to support the development of cytotoxic T lymphocytes (CTL) and the generation of antigen-specific tumour infiltrating lymphocytes (TIL), we investigated the interaction of these cytokines with an established TIL line. TIL proliferated in an additive fashion in response to suboptimal concentrations of IL-2 and various concentrations of IL-4. TIL possessed high-affinity IL-4 receptors whether cultured in recombinant IL-2 (rIL-2) or rIL-4, but cells cultured in rIL-2 had higher numbers of IL-4 receptors than cells cultured in rIL-4. When TIL were cultured in increasing concentrations of rIL-2, a dose-dependent enhancement in IL-4 receptor number was observed. The maximum induction of IL-4 receptor expression was achieved by 4 hr of incubation with rIL-2 and was completely blocked by cycloheximide. Other cytokines, such as rIL-1, recombinant tumour necrosis factor (rTNF), recombinant interferon-alpha (rIFN-alpha) and rIFN-gamma, had no effect on IL-4 receptor number. rIL-2 also up-regulated IL-4 receptors on CTLL-2, a murine CTL line. These data indicate that high-affinity IL-4 receptors exist on murine TIL and they can be up-regulated by IL-2. Our observation that IL-2 up-regulates IL-4 receptor may help explain the additive effects of these lymphokines on the proliferation of TIL and other cell lines. It may also help explain their co-operative effects on the generation of antigen-specific TIL and the differentiation of CTL. 相似文献
4.
Role of C3b receptors in the enhancement of interleukin-2-dependent T-cell proliferation 总被引:4,自引:0,他引:4
Anna Erdei E. Spaeth J. Alsenz E. Rüde T. Schulz J. Gergely M.P. Dierich 《Molecular immunology》1984,21(12):1215-1221
The mechanism by which the complement system influences immune responses to T-cell-dependent antigens has not yet been clarified. That is why we studied the effect of the third complement component (C3) on different T-cell-dependent processes using well-defined mouse T-cell lines. While C3 did not influence the interleukin-2 (IL-2) production of the ST2/K-9 helper T-cells, the IL-2-dependent proliferation of the ST1 line was shown to be dose-dependently enhanced by C3. It is proved that neither the haemolytic activity of C3 nor the C3a fragment had any role in the process. The effect of C3 on the IL-2-dependent T-cell growth is even more enhanced (up to five-fold) when using polymerised C3. When the ST1 cell line is cultured in the presence of the cross-linked ligand, T-cells formed 80% less rosettes with red blood cells coated with antibody and mouse or human C3b. It is strongly suggested that C3—particularly when aggregated—exerts its enhancing effect on the growth of the IL-2-dependent cell lines by binding to C3b receptors present on such T-cells. 相似文献
5.
6.
Martin Gullberg 《Molecular immunology》1987,24(12):1365-1371
Activated T cells express at least two affinity classes of interleukin-2 (IL-2) receptors. The number of low-affinity receptors is normally 10–30 times greater than that of the high-affinity receptors. In this report, normal human T cells are used in a cellular system in which the number of low-affinity receptors can be manipulated. The resulting receptor composition, which has been characterized in a previous report, contain such decreased levels of low-affinity IL-2 receptors that almost half of the surface pool of anti-IL-2 receptor antibody (anti-Tac) binding sites is associated with high-affinity receptors. By using such cells the dynamics and functions of high-affinity IL-2 receptors were studied and compared with receptors on a cell population expressing the normal 10–30-fold excess of anti-Tac binding sites over high-affinity IL-2 receptors. The results reveal that the rapid turnover of high-affinity IL-2 receptors is independent of the quantitative level of Tac antigen expression. The rapid kinetic of IL-2 internalization results in a 80–90% reduction of the steady-state levels of high-affinity receptors in the presence of IL-2. Most importantly, by using a cell population that expresses very low levels of Tac antigens, it became evident that IL-2 internalization is associated with an immediate substantial decrease of the surface level of anti-Tac binding sites. The Tac antigen thus appeared to be internalized together with the high-affinity IL-2 receptor complex but nevertheless the normal 10–30-fold excess of Tac antigens, over high-affinity IL-2 receptors, seems not to influence the process of internalization. 相似文献
7.
8.
Bovine T lymphocytes. I. Generation and maintenance of an interleukin-2-dependent, cytotoxic T-lymphocyte cell line. 总被引:3,自引:0,他引:3
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Primary and secondary bovine allogeneic mixed leucocyte cultures were examined for the generation of antigen-specific cytotoxic leucocytes. While optimal generation of murine and human cytotoxic T lymphocytes typically requires 4-8 days, alloantigen-specific cytotoxic bovine leucocytes were demonstrated consistently only after prolonged incubation periods, optimally found to be about 15 days. Restimulation of long-term bovine mixed leucocyte cultures with the original stimulator population revealed responder cells demonstrating augmented alloantigen-specific lytic activity. When placed into human recombinant interleukin-2, responder cells expanded and required passaging every 3-4 days. The same was not true of cells placed into interleukin-2-free medium. Cells cultured in interleukin-2-containing medium retained alloantigen specificity after 10 weeks of culture. Moreover, they continued to display total dependence on human, simian or bovine interleukin-2 for growth. 相似文献
9.
Y Shibata P G McCaffrey J Minowada A Volkman Y Oghiso 《Journal of leukocyte biology》1992,51(1):32-38
An interleukin 3 (IL-3)-dependent macrophage-like cell line, 11-1-B3, was newly established from CBA/J mouse bone marrow cell cultures. Assay of eicosanoids in the culture supernatants of the intact and [3H]arachidonic acid (AA)-prelabeled cells showed that, after stimulation with the Ca2+ ionophore A23187, the 11-1-B3 cells synthesized and released relatively large amounts of prostaglandin E2 (PGE2) and leukotriene B4 (LTB4) but not LTC4. In addition, 11-1-B3 cells showed Ca(2+)-dependent and alkaline pH-optimal phospholipase A2 (PLA2) activity that preferentially hydrolyzed cleavage of sn-2-arachidonyl- but not sn-2-oleoylphosphatidylcholine. The cellular enzyme was distributed with 90% of the activity in the cytosol and 10% in the membrane fraction. Treatment of cells with A23187 for 5-10 min resulted in five- to sevenfold increases in the membrane-associated PLA2 but activity in the cytosol was unchanged. This increase in membrane-associated enzyme activity was transient, returning to the pretreatment distribution after 30 min. In sharp contrast, phorbol myristate acetate (PMA) stimulation failed to induce either eicosanoid release or PLA2 activation, although PMA induced translocation of protein kinase C (PKC) to the membrane fraction within 10 min. The data suggest that increases in cellular Ca2+ directly activate membrane-associated PLA2 and consequently initiate AA metabolism; PKC activation by PMA requires additional steps to activate PLA2, a mechanism that is apparently deficient in the IL-3-dependent M phi-like cells. 相似文献
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12.
《Journal of immunological methods》1987,99(2):271-275
Biological interleukin-2 (IL-2) assays require cells that proliferate only in the presence of IL-2. The most often used human cells for this purpose have been phytohaemagglutinin (PHA)-stimulated buffy coat lymphocytes made dependent on IL-2. The results obtained by this T-blast method have not been easily reproducible and/or comparable due to differences in individual buffy coat batches. Murine cytotoxic T-cell lines (CTLL and CTLL-2) have been widely used in human IL-2 assays, but there remains a need for a human cell line. We have developed and IL-2 bioassay, based on the use of human HTLV-I-infected Th-cell line, ATH8. These cells express IL-2 receptor and are totally dependent on extrinsic IL-2 for proliferation. The results of the ATH8 IL-2 assay were reproducible and comparable to those obtained with successful T-blast assays. ATH8 cells do not require a prolonged culture period before they are suitable for the IL-2 assay as do T-blasts. 相似文献
13.
Regulatory effect of interleukin-4 (IL-4) on the expression and function of lymphocyte adhesion receptors involved in IL-2-induced cell aggregation.
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Human recombinant interleukin-4 (rIL-4) was studied for its capacity to inhibit rIL-2-induced lymphoid cell aggregation. In contrast to rIL-2, rIL-4 was unable to induce cluster formation by itself. However, when added simultaneously with rIL-2 to cultures of freshly isolated peripheral blood lymphocytes (PBL), rIL-4 inhibited cell aggregation in a dose-dependent way. In contrast, PBL, preactivated by a 4-day culture in the presence of 500 U/ml rIL-2, were not inhibited in their adhesive capacity by rIL-4. Inhibition of cell aggregation was most prominent at 24 hr and virtually lost after 72 hr of culture. Phenotypical analysis revealed that rIL-4, with similar kinetics, decreased the rIL-2-mediated up-regulation of the CD2, CD54 and CD49e adhesion molecules. In addition, it was observed that up-regulation of the activation epitope on CD11a recognized by the mAb NKI-L16, was prevented. During 24hr of culture rIL-4 itself did not alter the expression of these antigens. Blocking experiments with mAb directed against adhesion structures did not reveal a direct role for CD49e, but obviously demonstrated involvement of CD11a/CD18-CD54 and CD2-CD58 interactions in the rIL-2-induced adhesion. Therefore, rIL-4 appears to inhibit the early phase of rIL-2-induced aggregation by preventing the up-regulation of CD54 and CD2 antigens and by inhibiting the generation of the activated state of the CD11a/CD18 receptor. 相似文献
14.
F Akasu L Poplonski K M Snow R Volpe E C Keystone 《Clinical immunology and immunopathology》1992,63(2):142-147
We examined the expression of high-affinity interleukin (IL)-2 receptors (IL-2R) as well as Tac and HLA-DR antigens on peripheral blood (PB) T cells from 11 rheumatoid arthritis (RA) patients and 8 healthy controls induced in the autologous mixed lymphocyte reaction (AMLR). The proportion of HLA-DR- and Tac-bearing T cells and expression of these activation antigens were higher in patients relative to controls (P less than 0.01) in freshly isolated unstimulated PB mononuclear cells. AMLR stimulation of RA T cells failed to induce an increase in the proportion of HLA-DR and Tac-bearing T cells which was observed in health controls. After AMLR stimulation the number of high-affinity IL-2R were significantly lower in RA patients compared with controls (P less than 0.01). The number of high-affinity IL-2R on patient T cells correlated strongly with AMLR reactivity as measured by [3H]thymidine incorporation (r = 0.821, P = 0.002). The results suggest that the AMLR defect in RA may result from impaired generation of high-affinity IL-2R. 相似文献
15.
IL-2 receptors on rabbit T-cell lines and their transfectants expressing the human IL-2 receptor alpha chain.
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Low-affinity (dissociation constant: Kd = 7 nM) and high-affinity (Kd = 27 pM) interleukin-2 receptors (IL-2R) were detected on rabbit T-cell lines by IL-2 binding studies. Chemical cross-linking studies using 125I-labelled IL-2 showed that rabbit low-affinity IL-2R was singly expressed alpha-chain (MW 55,000) and that high-affinity IL-2R was composed of at least alpha- and beta- (MW 75,000) chains, similar to the human and murine counterparts. The existence of an additional chain (MW 25,000) was suggested in the rabbit IL-2R. Rabbit T-cell transfectant lines were established by human IL-2R alpha-chain (IL-2R alpha) cDNA transfection. These transfectant lines possessed not only extremely large numbers of human IL-2R alpha (over 10 times more than endogenous rabbit alpha-chain) but also twice as many high-affinity sites as their parental lines. The number of high-affinity sites on the transfectants significantly decreased when human alpha-chains were blocked, indicating that these transfectants expressed high-affinity receptor consisting of the exogenous human alpha-chain and rabbit beta-chain. This was confirmed by cross-linking experiments. The observation that expression of extremely large numbers of exogenous alpha-chains lead to an increase of the total number of high-affinity sites in the apparent absence of an increase of beta-chain expression raises the possibility that not only the beta-chain but also the alpha-chain may play an important role in regulating the number of high-affinity receptors. 相似文献
16.
The role of IL-4 and IL-2 on normal human T-cell activation and proliferation was studied. Both IL-2 and IL-4 were unable to induce proliferation of resting T cells. Therefore, we investigated their effect and the regulation of the T-cell proliferative response in competent T cells. T cells were rendered competent following incubation with PDB/ionomycin for 30 min or suboptimal concentrations of PHA for 60 min. Cells were then washed and recultured with PDB, IL-2, or IL-4 in the second or progression phase of the culture. Cells cultured in medium alone in this phase did not proliferate. IL-2 and IL-4 independently promoted competent T cells to proliferate to a similar degree as the response to PDB and the combination of IL-2 and IL-4 was not additive. The induction of competence and subsequent responsiveness to IL-2 and IL-4 could be maintained for about 24 hr after which time they become gradually less responsive to the interleukin in the progression phase. Addition of anti-IL-2R mAb or anti-IL-2 mAb resulted in selective inhibition of IL-2-mediated proliferation only. Similarly, addition of anti-IL-4 mAb resulted only in inhibition of IL-4-mediated proliferation. Addition of IL-2 during the progression phase led to an enhancement of IL-2R (TAC) expression while IL-4 did not affect IL-2R expression. The production of IL-2 and IL-4 by competent T cells could not be enhanced by the noncorresponding lymphokine. These results on the protein level were confirmed at the mRNA level as well and demonstrated that only PDB and IL-2 could induce IL-2 mRNA and PDB and IL-4 enhanced IL-4 mRNA. The immunosuppressive drug, cyclosporin A, failed to inhibit progression triggered by PDB, IL-2 or IL-4 in competent T cells. These findings suggest that IL-2 and IL-4 trigger T-cell proliferation through symmetrical, but independent pathways. 相似文献
17.
Expression of high-affinity IL-4 receptors on human melanoma,ovarian and breast carcinoma cells
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N. I. OBIRI J. P. SIEGEL F. VARRICCHIO R. K. PURI 《Clinical and experimental immunology》1994,95(1):148-155
It has previously been shown that murine sarcoma cells express high-affinity IL-4 receptors (1L-4R) which are internalized after binding to the ligand (Puri et al., Cancer Res 1991; 51:3011-7). We have also reported that human renal cell carcinoma cells express high-affinity IL-4R. and IL-4 inhibits tumour growth in vitro (Obiri et al, J Clin Invest 1993; 91:88). In this study we investigated the expression and function of IL-4R on other human solid tumours. Human melanoma, ovarian carcinoma and breast carcinoma cell lines were assessed for the cell surface expression of IL-4R by radio-ligand receptor binding and for IL-4R gene expression by Northern blot analysis. Primary cultures of mesothelioma and neurofibrosarcoma cells were similarly investigated. Human melanoma, ovarian carcinoma and breast carcinoma cell lines expressed IL-4R on their cell surface with a dissociation constant (Kd) of 140-549 pM. These tumour lines expressed a single 4 kb species of mRNA for IL-4R. Similarly, primary cultures of mesothelioma and neurofibrosarcoma cells were positive for the IL-4R mRNA by Northern blot analysis. Fresh, non-cultured mesothelioma and neurofibrosarcoma tumour sections were also positive for the presence of IL-4R as determined by immunohistochemistry of frozen sections using anti-IL-4R antibody. In order to study possible functions of IL-4R, we evaluated the effects of 11.-4 on cell growth and its effect on MHC antigen expression in the presence or absence of interferon-gamma (IFN-γ)- In tissue culture, IL-4 reduced the growth of tumour cell lines and primary cell cultures studied. IL-4 had very title effect on MHC class I antigen expression on ovarian, breast and melanoma cell lines; however. MHC class I (HLA-DR) expression was enhanced on melanoma and breast carcinoma cells. IL-4 also enhanced the IFN-γ-induced class II expression on melanoma and breast carcinoma cells. Taken together, our observations indicate that IL-4R are expressed on a variety of human solid tumours and these receptors may be functional. IL-4 alone and in combination with IFN-γ may play a role in host immune response against cancers. 相似文献
18.
Gangliosides interact with interleukin-4 and inhibit interleukin-4-stimulated helper T-cell proliferation. 总被引:2,自引:0,他引:2
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Gangliosides are potent immunosuppressive agents in vitro, and gangliosides shed from tumours in vivo may play an important role in the escape of tumours from immune destruction. We have investigated the effect of gangliosides on interleukin-4 (IL-4)-mediated processes in the murine helper T-cell line HT-2. Various gangliosides inhibited IL-4-stimulated DNA synthesis in HT-2 with IC50 values in the range 26-60 micrograms/ml. However, the proliferation of four lymphokine-independent cell lines was unaffected by 500 micrograms/ml gangliosides, as was the IL-1-stimulated secretion of IL-2 by EL-4 NOB-1 cells. Gangliosides were highly effective inhibitors when added to G0-G1-synchronized HT-2 cells during the first 6 hr after IL-4 stimulation, indicating that they act early in the IL-4 signalling pathway. High levels of exogenous IL-4 completely reversed inhibition of proliferation by gangliosides, which suggests that gangliosides compete with cellular IL-4 receptors for available lymphokine. Receptor-binding experiments confirmed that gangliosides blocked binding of [125I]IL-4 to receptors on intact HT-2 cells in a dose-dependent fashion. Gel-filtration fast protein liquid chromatography (FPLC) demonstrated that [125I]IL-4 co-eluted with ganglioside micelles after co-incubation before chromatography, and an overlay technique showed that IL-4 bound efficiently to gangliosides on thin-layer chromatography plates. Taken together, these results indicate that gangliosides act as potent suppressors of IL-4-dependent processes in lymphocytes, and that their mechanism of action involves direct interaction with IL-4, thus preventing IL-4 binding to high-affinity IL-4 receptors. This information helps to explain the diverse immunosuppressive actions reported for gangliosides, both in vitro and in vivo. 相似文献
19.
Goleva E Li LB Leung DY 《American journal of respiratory cell and molecular biology》2009,40(2):223-230
Corticosteroids are the most common therapeutic approach for control of tissue inflammation. Combination IL-2/IL-4 is known to induce T-cell steroid resistance. This can be reversed with IFN-gamma; however, the mechanism by which this occurs is unknown. In the current study, we found that treatment of peripheral blood mononuclear cells with combination IL-2/IL-4 for 48 hours, but not with IL-2 or IL-4 alone, abrogated dexamethasone (DEX)-induced glucocorticoid receptor (GCR)-alpha nuclear translocation in both CD4(+) and CD8(+) T cells. The presence of IL-4 significantly down-regulated IFN-gamma production by IL-2-stimulated cells. Importantly, addition of IFN-gamma to the IL-2/IL-4 combination restored GCRalpha nuclear translocation in response to DEX. Furthermore, DEX-induced mitogen-activated protein kinase (MAPK) phosphatase-1 induction, used as a readout for corticosteroid-induced transactivation, was significantly greater (P < 0.05) in media and IL-2/IL-4/IFN-gamma-treated conditions compared with IL-2/IL-4-treated cells. The combination of IL-2/IL-4 induced p38 MAPK activation in CD3(+) cells (30.5 +/- 5.7% cells expressed phospho-p38 MAPK versus no phospho-p38 MAPK expression after media treatment). The presence of the p38 MAPK inhibitor, SB203580, or IFN-gamma inhibited p38 MAPK phosphorylation and enhanced GCRalpha nuclear translocation in response to DEX. These data indicate that combination IL-2/IL-4 inhibits GCRalpha nuclear translocation in human T cells, and this effect is reversed by IFN-gamma via inhibition of p38 MAPK activation. 相似文献
20.
Role for T cells, IL-2 and IL-6 in the IL-4-dependent in vitro human IgE synthesis. 总被引:8,自引:0,他引:8
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E Maggi G F Del Prete P Parronchi A Tiri D Macchia P Biswas C Simonelli M Ricci S Romagnani 《Immunology》1989,68(3):300-306
The role of T cells and monocytes, as well as that of cytokines, such as IL-1, IL-2 and IL-6, on the IL-4-dependent in vitro human IgE synthesis was investigated. Recombinant IL-4, IL-4-containing T-cell clone supernatants and different combinations of recombinant cytokines failed to induce highly purified B cells to synthesize IgE. IL-4-dependent IgE synthesis was restored by addition to purified B cells of either untreated or mitomycin C-treated autologous T lymphocytes. Addition to purified B cells of autologous monocytes did not restore the IgE response, but usually it exerted a potentiating effect on the synthesis of IgE induced by IL-4 in the presence of suboptimal concentrations of T cells. The activity of T cells apparently preceded that of IL-4 and required a physical contact with B cells. The presence in culture of IL-2 also appeared to be necessary for the T-cell and IL-4-dependent IgE synthesis. Even though not essential, IL-6 was able to potentiate IgE synthesis in most experiments, whereas IL-1 did not display any modulatory effect. 相似文献