共查询到20条相似文献,搜索用时 15 毫秒
1.
K Abdi M L Reed S J Mentzer S H Herrmann 《Journal of immunology (Baltimore, Md. : 1950)》1989,142(9):2971-2980
Antibody reactive with the CD3 complex on the surface of T lymphocytes can either: inhibit CTL lysis of target cells expressing Ag; or redirect CTL to lyse target cells expressing FcR in the absence of Ag expression. To investigate these phenomena we examined the effect of anti-CD3 mAb on two indicators of CTL activation, the release of esterase and target cell lysis. Esterase release by long term allo-reactive human CTL in response to target cells (JY or HLA transfected K562 cells) was found to be Ag specific and correlate with target cell lysis. Addition of anti-CD3 to either JY targets or K562 cells expressing FcR resulted in a high level of esterase release. Triggering of esterase release was found with both soluble intact and Fab fragment of anti-CD3 in the absence of cells expressing measurable FcR. This apparent FcR-independent triggering of esterase release occurred at 37 degrees C but not at 24 degrees C. In contrast esterase activity was released from CTL at both 24 and 37 degrees C in response to intact target cells, JY or K562 cells plus intact anti-CD3 mAb. Addition of anti-CD3 mAb, at a level capable of blocking target cell lysis by greater than 50%, resulted in an initial velocity of esterase release almost twice that found in response to JY target cells. With a low level of anti-CD3 mAb, able to block JY lysis by approximately 10%, the initial rate of esterase release was much slower than that found in response to target cells. In contrast when FcR+ cells, K562, were added along with a low level of anti-CD3 the initial velocity of esterase release was about twofold more than the velocity of esterase release triggered by soluble anti-CD3 alone. These results indicate that soluble antibody can trigger long term active CTL and the velocity of this triggering correlates with anti-CD3-mediated inhibition as well as redirected lysis. 相似文献
2.
Early steps in specific tumor cell lysis by sensitized mouse T lymphocytes. I. Resolution and characterization. 总被引:4,自引:0,他引:4
E Martz 《Journal of immunology (Baltimore, Md. : 1950)》1975,115(1):261-267
Addition of high molecular weight dextran to culture medium prevents the initiation of T lymphocyte-mediated killing by holding the cytolytic T lymphocytes (CTL) and target cells in suspension and preventing intercellular contact. Suspension in 10% dextran was used to interrupt the ongoing formation of adhesions between CTL and target cells already in contact in a centrifuged pellet. The results demonstrate that 1) firm adhesions form between CTL and target cells within 1 min at 37 degrees C; 2) once formed, these adhesions are stable at low temperature and are resistant to mechanical shearing forces; 3) these adhesions can be disrupted by EDTA; 4) immediately after the adhesions form, separation of the CTL from the target cells prevents lysis of the latter; 5) after incubation of targets adhering to CTL for an additional 6 min at 37 degrees C, removal of the CTL no longer prevents target cell lysis. Thus, target cells become "programmed" for subsequent lysis within a few minutes after contact with CTL, after which lysis occurs during the next several hours without further participation of the effector cell. At 15 degrees C, adhesions form 1/17 as fast as at 37 degrees C. Programming of target cells for lysis occurs 1/76 as fast at 15 degrees C as at 37 degrees C. Thus, the programming for lysis step is about 4-fold more temperature dependent than the adhesion step. In addition to being detected by subsequent target cell lysis in 10% dextran, the adhering cell clusters can be counted with low power microscopy. This permitted verification that EDTA separates the clusters after programming for lysis is complete. Moreover, the great majority of the clusters seen at 37 degrees C are antigen-specific. Knowledge of the cluster size distribution and the subsequent level of lysis permits the deduction that not less than 6% of the sensitized peritoneal cell populations used were CTL. 相似文献
3.
A Müllbacher R Wallich R W Moyer M M Simon 《Journal of immunology (Baltimore, Md. : 1950)》1999,162(12):7315-7321
Previous observations that the highly conserved poxvirus-encoded serpins inhibit cytotoxic activities of alloreactive CTL via granule and/or Fas-mediated pathways was taken to indicate their involvement in immune evasion by poxviruses. We now show that interference with 51Cr release from target cells by ectromelia and cowpoxvirus is limited to alloreactive but not MHC-restricted CTL. The data are in support of the paramount importance of CTL and its effector molecule perforin in the recovery from primary ectromelia virus infection and question the role of serpins in the evasion of poxviruses from killing by CTL. Further analysis of poxvirus interference with target cell lysis by alloreactive CTL revealed that suppression primarily affects the Fas-mediated, and to a lesser extent, the granule exocytosis pathway. Serpin-2 is the main contributor to suppression for both killing pathways. In addition, inhibition of lysis was shown to be both target cell type- and MHC allotype-dependent. We hypothesize that differences in TCR affinities and/or state of activation between alloreactive and MHC-restricted CTL as well as the quality (origin) of target cells are responsible for the observed phenomenon. 相似文献
4.
D T Harris A H Hale L Lefrancois 《Journal of immunology (Baltimore, Md. : 1950)》1981,126(5):1914-1918
We have analyzed the requirement for the expression of the major surface glycoprotein (G protein) of vesicular stomatitis virus (VSV) on target cells for recognition and lysis by anti-VSV cytotoxic T lymphocytes (CTL). In addition, we have attempted to determine if the carbohydrate moieties on the G protein are required for recognition and lysis by anti-VSV CTL. When VSV (Orsay) is grown at 30 degrees C in the presence of tunicamycin (TM), glycosylation of G protein is inhibited; however, nonglycosylated G protein is found on the surface of the cell and active virus particles are produced. In contrast, VSV (Orsay) grown at 39 degrees C in the presence of TM produces low titers of virus and the presence of G protein on the surface of cells is not detectable. The susceptibility of these target cells to lysis by anti-VSV CTL was analyzed. The results suggest that expression of the G protein is required for target cell lysis by anti-VSV CTL. However, the presence of the carbohydrate moieties on the G protein are nt an absolute requirement for recognition by anti-VSV CTL. VSV-infected target cells incubated in the presence of TM were lysed by anti-VSV CTL up to 50 to 80% of the infected target cell control. This result suggests either that some clones of anti-VSV CTL recognize carbohydrate moieties or that carbohydrate moieties play some as yet undefined nonantigenic role in the recognition of the target antigen by the CTL receptor. 相似文献
5.
T lymphocyte-mediated cytolysis: dissociation of the binding and lytic mechanisms of the effector cell. 总被引:3,自引:0,他引:3
To determine functional relationships between the cytotoxic T lymphocyte (CTL) receptor for target binding and the lytic mechanism, we have studied the reaction between two immunized CTL populations (AalphaB and BalphaA), both at the population and the single-cell level. When studied at the population level, the reaction of AalphaB with BalphaA (bidirectional system) resulted in formation of AalphaB/BalphaA conjugates and bidirectional cytolysis. However, when the viability of cells in individual AalphaB/BalphaA conjugates was analyzed, unidirectional instead of bidirectional lysis occurred. These results indicate that under conditions that are conducive to lysis, binding of a potentially lytic cell to its target does not necessarily result in target lysis. Short heat treatment of CTL (44 degrees C, 10 min) totally abolished their lytic activity, without affecting their capacity to bind specifically, thus dissociating the binding from the lytic activity of the CTL. The cytolytic activity is probably associated with, or triggered by the CTL-binding unit. The binding unit, on the other hand, appears to be a functional receptor of the CTL, which is involved in but not sufficient to bring about lysis. 相似文献
6.
Nicotinamide in concentrations of 5 mM and greater protected fibroblast target cells from lysis by lymphokine-activated killer cells (LAK cells). Protection was concentration dependent and was exerted at the level of the target cell. Nicotinamide did not interfere with effector-target cell conjugate formation or with the calcium dependent triggering step of the lytic process. Target cell lysis in cultures without nicotinamide was accompanied by fragmentation of target cell DNA. The DNA of target cells cultured with LAK cells in the presence of nicotinamide remained intact. 3-Aminobenzamide which, like nicotinamide, inhibits poly(ADP-ribose) synthetase but is not a precursor of NAD, was an effective inhibitor of target cell lysis while nicotinic acid, an alternative precursor of NAD in cells, was not. The data point to a central role for poly(ADP-ribose) synthetase in the events leading up to DNA fragmentation and the release of 51Cr from target cells damaged by lymphokine-activated killer cells. 相似文献
7.
A K Abbas 《Journal of immunology (Baltimore, Md. : 1950)》1979,123(5):2011-2018
Alloreactive cells generated by in vitro stimulation of C57BL/6 (H-2b) spleen lymphocytes with irradiated MOPC 315 or MOPC 104E(H-2d) cells were shown to lyse 51Cr-labeled myeloma targets at high effector:target ratios under conditions of inefficient cell contact, the alloreactive cells cause variable and frequently minimal lysis of myeloma targets but markedly suppress antibody secretion even by viable myeloma cells. The suppressor cells are radioresistant T cells lacking I-J subregion-encoded surface determinants; their precursors are insensitive to cyclophosphamide; suppression is H-2 specific and not mediated by secreted factors; and the suppression is blocked by Cytochalasin B, a known inhibitor of T cell-mediated cytolysis. These properties are typical of cytolytic T lymphocytes (CTL) and not of defined suppressor T cells, suggesting that inhibition of myeloma function probably represents a pre-lytic effect of the alloreactive CTL, although a CTL-like suppressor cell effect cannot be definitively excluded. These results are discussed with reference to the possible relationships between suppressor and cytolytic T lymphocytes. 相似文献
8.
Lack of DNA degradation in target cells lysed by granules derived from cytolytic T lymphocytes 总被引:7,自引:0,他引:7
S H Gromkowski T C Brown D Masson J Tschopp 《Journal of immunology (Baltimore, Md. : 1950)》1988,141(3):774-778
It has been shown previously that fragmentation of target cell DNA is an early event in lysis mediated by cytolytic T lymphocytes (CTL). In this study, we have investigated whether CTL-derived granules that exhibit lytic activity also induce DNA fragmentation in murine target cells. Cytolytic granules isolated from three different alloreactive CTL clones were tested for the induction of DNA fragmentation in P815 and EL4 target cells, by using a Triton X-100-facilitated, radiolabeled DNA release assay. In contrast to the CTL clones from which they were derived, the cytolytic granules did not induce DNA fragmentation. Agarose gel electrophoretic analysis of DNA confirmed the lack of discrete DNA fragments in target cells lysed by CTL-derived granules. Possible explanations for the difference in the ability of CTL and CTL-derived granules to trigger DNA fragmentation are discussed. 相似文献
9.
Alex Matter Laurée Müller 《Virchows Archiv. B, Cell pathology including molecular pathology》1981,37(1):11-28
Heterologous (rabbit) antibodies were raised against murine P-815 mastocytoma cells of DBA/2 origin. Antisera and IgG preparations were highly cytotoxic, whereas Fab fragments thereof lost all activity. Fab fragments also showed a much lower avidity than IgG, both for tumor and normal DBA/2 and C57 spleen cells as measured by the release of iodinated Fab and IgG. Both preparations bound specifically to P-815 cells since they were capable of inhibiting T cell-mediated target cell lysis. The binding of IgG and monovalent Fab fragments was studied by fluorescence. Rhodamine-coupled IgG bound homogeneously in the cold and quickly formed patches upon warming but did not form caps even after prolonged incubation at 37 degrees C. Rhodamine-coupled Fab fragments also bound homogeneously. Their distribution was unaltered after incubation at 37 degrees C even when tumor cells formed uropod-like tails. Fab fragments, however, could be induced to cap with a second and third antibody layer. P-815 cells labeled with rhodamine-coupled Fab fragments were incubated with cytolytic T cells (CTL). The conjugates formed between CTL and fluorescent target cells were observed. No gross redistribution of surface antigens on target cells was observed even at late stages of the lytic process. CTL, therefore, do not seem to operate via a redistribution of surface antigens. 相似文献
10.
Free fatty acids are produced in and secreted from target cells very early in cytotoxic T lymphocyte-mediated killing 总被引:1,自引:0,他引:1
Conjugation of CTL with their cognate targets elicits a number of early changes within the target cell that are thought to play an important role in the lytic mechanism. We now report that at times earlier than 5 min after conjugation with allospecific CTL, free fatty acids (FFA) are produced in and then secreted from alloantigen-bearing target cells. Using murine CTL clones with different alloantigen specificities, stimulation of FFA production from target cells was found to be Ag specific. FFA production does not appear to be specific for any particular FFA species. Indeed, a wide spectrum of cis unsaturated as well as saturated FFA are produced. FFA production is well correlated with, and specific for, CTL-mediated target cell lysis. Other means of perturbing or lysing target cells, including freeze/thaw disruption, detergent solubilization, or increasing membrane permeabilization with ionomycin, do not stimulate FFA production. In particular, FFA production is not stimulated by treatment with pore-forming granules under conditions that cause more than 90% target cell lysis. These results suggest that FFA production plays an important role in CTL-mediated lysis because stimulation of FFA release specifically requires an event that is CTL induced, occurs very early after conjugation, and is strongly correlated with CTL-mediated lysis. 相似文献
11.
Beal AM Anikeeva N Varma R Cameron TO Norris PJ Dustin ML Sykulev Y 《Journal of immunology (Baltimore, Md. : 1950)》2008,181(7):4815-4824
Destruction of virus-infected cells by CTL is an extremely sensitive and efficient process. Our previous data suggest that LFA-1-ICAM-1 interactions in the peripheral supramolecular activation cluster (pSMAC) of the immunological synapse mediate formation of a tight adhesion junction that might contribute to the sensitivity of target cell lysis by CTL. Herein, we compared more (CD8(+)) and less (CD4(+)) effective CTL to understand the molecular events that promote efficient target cell lysis. We found that abrogation of the pSMAC formation significantly impaired the ability of CD8(+) but not CD4(+) CTL to lyse target cells despite having no effect of the amount of released granules by both CD8(+) and CD4(+) CTL. Consistent with this, CD4(+) CTL break their synapses more often than do CD8(+) CTL, which leads to the escape of the cytolytic molecules from the interface. CD4(+) CTL treatment with a protein kinase Ctheta inhibitor increases synapse stability and sensitivity of specific target cell lysis. Thus, formation of a stable pSMAC, which is partially controlled by protein kinase Ctheta, functions to confine the released lytic molecules at the synaptic interface and to enhance the effectiveness of target cell lysis. 相似文献
12.
A S Perelson C A Macken E A Grimm L S Roos B Bonavida 《Journal of immunology (Baltimore, Md. : 1950)》1984,132(5):2190-2198
We measured the effects of having multiple cytotoxic T lymphocytes (CTL) bound to one target cell by using the single-cell cytotoxicity in agarose assay. We found that even though there is variability in the time at which individual target cells are lysed, we can identify a general trend: the mean rate of lysis increases with the number of CTL bound per target cell, reaching a maximum when the CTL-target cell ratio is three. Combining a quantitative model for the rate of lethal hitting in multicellular conjugates with a multi-event model for the rate of target cell disintegration, we developed a new multistage kinetic model for predicting the rate of target cell lysis in multiple lymphocyte-target cell conjugates. The variability in the time at which target cells are hit and the variability in the time until they disintegrate are incorporated into the model. By analyzing our measured data in the context of the multistage kinetic model, we were able to estimate via nonlinear least squares regression the target cell disintegration rate, but not the lethal hitting rate. Lethal hitting appeared to be too fast, when compared with disintegration, to significantly affect the time of target cell lysis. By using previously determined values of the lethal hitting rate for single lymphocyte-target cell conjugates and by postulating that lymphocytes act independently of each other in delivering lethal hits, we were able to estimate the rate at which target cells are hit in multiple-lymphocyte single target cell conjugates. By using this estimate of the lethal hitting rate and the regression estimate of the disintegration rate, the multistage kinetic model gave a quantitative fit to our data. From this analysis, we found that the rate at which a target cell disintegrates after being lethally hit increases with the number of CTL per conjugate. This result is quite surprising, because once the first hit has been received, a target cell can disintegrate in a killer cell-independent manner. Under the conditions of our experiment, it appears as if target cell disintegration is not killer cell-independent. Furthermore, our analysis of the time course of target cell disintegration suggests that the process is not governed by simple first order kinetics, but rather by a more complex multistep mechanism. 相似文献
13.
Inappropriate activation of p34cdc2 kinase has been shown to occur during apoptosis induced by cytotoxic T-cell derived perforin and fragmentin. We analysed the effect of two inhibitors of p34cdc2 kinase on alloreactive Tc-cell-mediated lysis and DNA fragmentation of P815 and L1210 target cells. Olomoucine, a specific inhibitor of cyclin dependent kinases, did not affect DNA fragmentation in the target cells. Lysis of olomoucine-treated target cells as assessed by 51Cr release over a typical 8-h period was also unaffected. We also examined the effects of thapsigargin on target cell death. This toxin causes increased intracellular calcium rises that then result in irreversible inhibition of cyclin dependent kinases, including p34cdc2 kinase. The same extent of specific cell lysis was induced by cytotoxic T cells from perforin(-/-), granzyme B(-/-), granzyme A(-/-), perforin(-/-) X granzymeB(-/-) X granzymeA(-/-) KO mice or normal mice in untreated target cells or target cells treated with either olomoucine or thapsigargin. Similarly DNA fragmentation measured by release of tritiated DNA was also unaffected. Thus inhibition of p34cdc2 kinase affects neither the Fas nor the perforin/granzyme pathways of alloreactive cytotoxic T-cell killing as measured by DNA fragmentation or chromium release. P815 cells treated with olomoucine were arrested in the cell cycle after 12-16 h exposure to the toxin. After cell cycle arrest, target cells now showed enhanced 51Cr release induced by effector cytotoxic T cells (CTL) derived from perforin(-/-) mice compared to untreated cells. This lysis was accompanied by an increase in cell surface Fas expression. Olomoucine induced cell cycle arrest and expression of Fas was reversible and when cells re-entered the cell cycle, surface expression of Fas was lost. 相似文献
14.
O,S,S,-Trimethyl phosphorodithioate (OSS-TMP), an organophosphate esterase inhibitor, has been shown to block the effector phase of the cytolytic reaction mediated by murine and human cytotoxic T lymphocytes (CTL) and human natural killer cells. The murine interleukin 2-dependent CTLL-1 (anti-Iad) clone was used to determine the phase of the cytolytic pathway inhibited by OSS-TMP. Pretreatment of the CTL or target cell with OSS-TMP was not effective at blocking lysis; however, inhibition of lysis was achieved if the reaction was carried out in the continuous presence of OSS-TMP (IC50 = 55 microM) or when CTL-target conjugates were performed and incubated with OSS-TMP (IC50 = 640 microM). Two structural analogues of OSS-TMP were unable to inhibit CTL-mediated lysis. In contrast to OSS-TMP, N-alpha-p-tosyl-L-lysine chloromethylketone required only a 5-min preincubation with the CTL to inhibit lysis. OSS-TMP did not block recognition-adhesion step(s) of the reaction since the ability to form conjugates was not impaired; however, the lytic efficiency of individual CTL-target pairs were blocked. OSS-TMP did not appear to be an inhibitor of the major granule-associated protease that cleaves the substrate, N-alpha-benzyloxycarbonyl-L-lysine thiobenzylester. Ca2+ pulse and kinetic experiments indicated that the OSS-TMP-sensitive site was at a pre-Ca2+-dependent phase but after recognition-adhesion. Human CTL and natural killer cell activity was also inhibited by OSS-TMP, suggesting the presence of a common site of action among these cytolytic systems. The results indicate that OSS-TMP may be a useful reagent in characterizing the early post-recognition events in the cytolytic pathway of CTL and natural killer effector cells. 相似文献
15.
Kojima Y Kawasaki-Koyanagi A Sueyoshi N Kanai A Yagita H Okumura K 《Biochemical and biophysical research communications》2002,296(2):328-336
Fas ligand (FasL) has been implicated in cytotoxic T lymphocyte (CTL)- and natural killer (NK) cell-mediated cytotoxicity. In the present study, we investigated the localization of FasL in murine CTL and NK cells. Immunocytochemical staining showed that FasL was stored in cytoplasmic granules of CD8+ CTL clones and in vivo activated CTL and NK cells, where perforin and granzyme A also resided. Immunoelectron microscopy revealed that FasL was localized on outer membrane of the cytoplasmic granules, while perforin was localized in internal vesicles. Western blot analysis showed that the membrane-type FasL of 40 kDa was stored in CD8+ CTL clones but not in CD4+ CTL clones. By utilizing a granule exocytosis inhibitor (TN16), we demonstrated that FasL translocated onto cell surface upon degranulation of anti-CD3-stimulated CD8+ CTL clones. Moreover, TN16 markedly inhibited the FasL-mediated cytotoxicity by CD8+ T cell clones and NK cells. These results suggested a substantial contribution of FasL to granule exocytosis-mediated target cell lysis by CD8+ CTL and NK cells. 相似文献
16.
R F Todd 《Cellular immunology》1975,20(2):257-268
Target tumor cells pretreated with high concentrations of papain or Pronase were resistant to lysis by cytotoxic T lymphocytes (CTL), whereas treatment with trypsin or neuraminidase had no protective effect. Parallel determinations of the H-2 content of target cells following enzyme treatment showed that approximately 80% of surface H-2 was removed by papain or Pronase, 40% by trypsin, and virtually none by neuraminidase treatment. Both susceptibility to lysis by CTL and content of surface H-2 after papain treatment were fully restored by 6 hr at 37 °C in nutrient medium. These findings suggest that lymphocyte-mediated cytolysis (LMC) determinants (target cell antigens bound by CTL) are sensitive to degradation by papain and Pronase but are resistant to the enzymatic action of trypsin and neuraminidase. That a similar pattern of enzyme sensitivity is shown by serologically defined H-2 antigens indicates that both functional classes, LMC and H-2, may have a structural association. 相似文献
17.
E Martz 《Journal of immunology (Baltimore, Md. : 1950)》1976,117(3):1023-1027
In a previous study of the mechanism of specific target cell lysis by alloimmune cytolytic T lymphocytes (CTL), we established that the target cell becomes irreversibly programmed to lyse within a few minutes after contact with the CTL. We here show that at each point in time, the level of specific release of the potassium analog, 86Rb equals the percentage of target cells which have been programmed to lyse. It is also shown that specific release of 86Rb is more rapid than that of a small metabolite of similar weight, 14C-nicotinamide, which in turn is specifically released more rapidly than 51Cr. Thus, an electrolyte-permeable lesion is produced in the target cell membrane within minutes of contact with the CTL. Since measurements of 86Rb release, unlike measurements of programming for lysis, do not involve exposure of the cells to EDTA and vigorous shearing forces, the present observations corroborate and extend, by an independent and gentler method, our previous conclusion that the CTL effects crucial and irreversible changes in the target cell within minutes after contact. The present results are consistent with the possibility that the first, and perhaps the only damage administered directly by the CTL is a membrane lesion permeable to electrolytes and possibly to small molecules. 相似文献
18.
Recent investigations examining mononuclear cell antibody-dependent cell-mediated cytotoxicity against tumor cell lines suggest that K lymphocytes and not monocytes are active in this cytotoxic reaction. We have found, however, that in an allogeneic assay system, human monocyte monolayers as well as lymphocytes mediate substantial lysis of 51Cr-labeled antibody-coated CEM lymphoblast tumor cells. This cytotoxicity is temperature-dependent and rapid, with most 51Cr release occurring in the first 4 hr of co-incubation. Interaction between target cell-bound antibody and the monocyte Fc receptor is necessary as demonstrated by the marked fall in antibody-dependent cell-mediated cytotoxicity (ADCC) produced by staphylococcal protein A, high concentrations of nonspecific immunoglobulin, and dilution of the target cell antiserum. Morphologic and functional characteristics of the monocyte-monolayer preparations establish their relative purity (greater than 95%) and indicate that monocytes and not contaminating lymphocytes are responsible for tumor cell lysis. Furthermore, preincubation of monocyte and lymphocyte preparations with latex particles or low concentrations of immunoglobulin distinguished monocyte from lymphocyte ADCC. Thus, normal human monocytes have the capacity to carry out antibody-dependent cytotoxicity against nucleated malignant target cells. 相似文献
19.
P Perez J A Bluestone D A Stephany D M Segal 《Journal of immunology (Baltimore, Md. : 1950)》1985,134(1):478-485
The formation of conjugates between cloned anti-H-2Kb and Dd cytotoxic T lymphocytes (CTL) and splenic target cells has been studied by dual parameter flow cytometry. By varying effector-target combinations and by blocking with anti-MHC class I monoclonal antibodies, we found that the specificity of conjugate formation, in general, paralleled that expected from cytotoxicity studies; however, a significant number of "nonspecific" conjugates was always observed. As expected from previous studies, conjugate formation did not occur below 10 degrees C and was inhibited by cytochalasin B, EDTA, and anti-Lyt-2 antibodies. Conjugate formation followed first-order kinetics. The rate of formation of conjugates increased with temperature from 24 degrees to 37 degrees C; at 37 degrees C, the half-time was 1.4 min. After a 6-min lag period, lysis of target cells could be detected at 37 degrees C but not at 30 degrees C or below. Because target cell lysis proceeded during a period of time when the number of conjugates remained constant, considerable effector cell recycling must have occurred. Comparisons of the fluorescence emissions from conjugated effector or target cells with those from unconjugated cells demonstrated that nearly all conjugates contained one effector cell and one target cell, independent of the ratio of the two cell types in the original mix. Once formed, anti-H-2Kb conjugates were stable when diluted into medium alone, but rapidly disaggregated in medium containing either anti-Lyt-2 or anti-Kb monoclonal antibodies, both of which blocked conjugate formation. This finding suggests that conjugates are normally stabilized by intercellular bonds that are constantly breaking and reforming at the cell:cell interface, and that the antibodies disrupt the conjugates by preventing the reformation of broken bonds. 相似文献
20.
I S Misko T B Sculley C Schmidt D J Moss T Soszynski K Burman 《Cellular immunology》1991,132(2):295-307
We have approached the challenge of generating a primary T cell response to Epstein-Barr virus (EBV) in vitro by stimulating naive T cells with the autologous EBV-transformed lymphoblastoid cell line (LCL), a rich source of EBV-associated cytotoxic T lymphocyte (CTL) epitopes. Responsive T cells from three EBV-seronegative donors were cloned in agarose, phenotyped for T cell markers by flow cytometry, and their cytotoxic properties analyzed in the 51Cr release assay. Most clones (greater than 95%) expressed the CD4 phenotype and 59% of these clones showed cytotoxic properties. The dominant CTL response was specific for FCS-associated epitopes presented by FCS-grown autologous LCL target cells and was restricted by class II HLA antigens. Other clonal components included: (i) an EBV-specific response by HLA-restricted CD4 CTL clones that did not discriminate between A- and B-type EBV transformants; (ii) an EBV-specific response by an HLA-restricted CD4 CTL clone that discriminated between A- and B-type transformants, and (iii) a nonspecific cytotoxic response by CD3+,4+,8-, CD3+,4-,8-, and CD3-,4-,8- clones that were broadly allotypic or restricted to the lysis of K562 target cells. The EBV-specific CTL clones did not lyse the autologous EBV-negative B or T cell blasts and their specificity patterns of lysis were supported by the cold target competition data. These studies highlight the role of CD4 CTL in the establishment in vitro of a primary immune response to a human virus. 相似文献