Effect of cyclosporin A on human lymphocyte responses in vitro. III. CsA inhibits the production of T lymphocyte growth factors in secondary mixed lymphocyte responses but does not inhibit the response of primed lymphocytes to TCGF

The mechanism whereby Cyclosporin A (CsA) inhibits secondary mixed lymphocyte responses was assessed. CsA added to secondary MLR cultures inhibited proliferation and induction of cytolytic lymphocyte activity. This inhibition was found to be associated with the inhibition of T lymphocyte stimulating growth factor(s) (TCGF) production in the supernatants of secondary MLR cultures. As little as 1.0 micrograms/ml of CsA added to secondary MLR cultures resulted in no measurable TCGF activity. In contrast, moderate doses of CsA (1.0, 2.5 micrograms/ml), which completely inhibited the secondary MLR response to alloantigen, did not inhibit the proliferative and CML response of alloantigen-primed lymphocytes to these stimulating growth factors. Even at high doses of CsA (20 micrograms/ml), substantial levels of proliferation (50% of control response) and CML induction (60% of control response) were observed when the primed cells were exposed to secondary MLR supernatants containing TCGF activity. It was concluded that inhibition of secondary mixed lymphocyte responses by CsA may be due in part to the inhibition of TCGF production rather than the inhibition of the effect of TCGF on mature cytotoxic T lymphocytes.     

Selective effects of interferon on distinct sites of the T lymphocyte triggering process

Lectin- and antigen-induced proliferation of murine T cells consists of two major events, namely, a rapid induction of susceptibility to growth factors and a later-occurring, accessory cell-dependent production of T cell growth factors (TCGF). The mechanism by which interferon (IFN) inhibits T cell responses was studied accordingly. A decrease of Con A-induced proliferation was observed in the presence of increasing amounts of IFN. The reduced proliferative response in such cultures was found to be due to an accumulation of cells in the G0/G1 phase of the cell cycle. Furthermore, the results show that IFN did not inhibit the early events in T cell triggering, because the acquisition of responsiveness of resting T cells to TCGF was unaltered in the presence of IFN, nor did it interfere with production of TCGF. In contrast, IFN was found to interfere with the TCGF-dependent T cell blast growth. Cytofluorometric analysis of the proliferative phase revealed that IFN exerts its effect on T cells, which have entered the proliferative cycle, by a postmitotic accumulation in G0/G1, thus reducing the proliferating population. The results demonstrate that IFN primarily affects the later phase of proliferative activity after T cell triggering, leaving the helper cell functions untouched.     

An anticomplementary agent, K-76 monocarboxylic acid: its site and mechanism of inhibition of the complement activation cascade.

A monocarboxylic acid derivative (K-76 COOH) of K-76, purified from the culture filtrate of Stachybotrys complement I nov. sp. K-76, inhibits complement (C) activity. Its inhibitory action is mainly on C5 step. It strongly inhibits the generation of EAC1,4b,2a,3b,5b from C5 and EAC1,4b,2a,3b, and accelerates the decay of EAC1,4b,2a,3b,5b. It also causes some inhibition of the reactions of the reactions of C2,C3,C6,C7 and C9 with their respective preceding intermediate cells. It has no effect on the generation of EAC1,4b from C4 and EAC1, or of EAC-8 from C8 and EAC-7, and apparently increases the generation of EAC1,4b from C1 and EAC4b probably by inhibiting transfer or turnover of C1. It does not affect the rate of decay of EAC1,4b,2a or the T max of generation of EAC1,4b,2a, and it inhibits immune adherence only at high concentration. K-76 COOH also strongly inhibits hemolysis through the alternative pathway of C activation by cobra venom factor, but it does not seem to inhibit the early steps of the alternative pathway, because it has little affect on the consumption of C3 or the conversion of beta 1C to beta 1A on treatment of C serum with zymosan. K-76 COOH probably combines with C5 molecules, forming the inactive complexes, or it causes the structural alteration of C5.     

A Complement Inhibitor Produced by Stachybotrys complementi,nov. sp. K-76, a New Species of Fungi Imperfecti

A complement inhibitor, K-76, was isolated and purified from the culture supernatant of a fungus, Stachybotrys complementi, nov. sp. K-76, isolated from soil of Ishigaki Island, Okinawa. K-76 is a sesquiterpene compound and it can be oxidized to a monocarboxylic derivative (K-76 COOH), the sodium salt of which is very soluble and much less toxic than K-76. K-76 and K-76 COOH both inhibited complement activation by either the classical or alternative pathway. They inhibited generation of the factor chemotactic to human polymorphonuclear leukocytes from human serum by aggregated immunoglobulin. When sensitized erythrocytes were treated with complement in the presence of K-76 COOH, the resulting unlysed cells were found to be in the state of EAC1, 4b, 2a, 3b. Thus K-76 COOH is considered to block mainly the C5 intermediate step. K-76 COOH did not inhibit any proteases or esterases tested, except when tested at high concentration.     

Anti-idiotypic antiserum to monoclonal anti-Sm inhibits the autoantigen-induced proliferative response

Anti-idiotypic sera were produced in BALB/c mice against three established monoclonal anti-Sm antibodies. Inhibition assays showed that the anti-idiotypic antibodies recognized determinants that were present on all three monoclonal antibodies but not on normal mouse IgG from unimmunized BALB/c mice or myeloma proteins. Normal (+/+) and autoimmune (lpr/lpr) MRL/MpJ or C3H/HeJ mice were immunized with Sm in complete Freund's adjuvant. Immune T cells from the draining lymph nodes proliferated in response to the addition of Sm in vitro. Anti-idiotypic serum added to these cultures inhibited the proliferative response by 50 to 70%, whereas normal BALB/c serum had no effect. This inhibition of proliferation was antigen specific, because the anti-idiotypic serum did not inhibit the T cell proliferative response to an irrelevant antigen, TNP-KLH, or ovalbumin. Kinetic studies showed that the anti-idiotypic serum inhibited an early event in antigen-induced proliferative response, because the addition of serum late in culture did not cause any significant reduction in proliferation. The reduced proliferative response was due to direct action of the anti-idiotypic serum on the Lyt-1+, 2- T cell population.     

Mechanism of T cell activation. II. Antigen- and lectin-dependent acquisition of responsiveness to TCGF is a nonmitogenic, active response of resting T cells

Small resting T cells, which do not respond to T cell growth factor (TCGF), acquire responsiveness upon a short (4-hr) pulse of specific ligands by presenting growth receptors for TCGF. The results demonstrate that the same mechanisms operate in the specific induction of primary MLR in that a 5-hr MLR is sufficient to render the responder cells reactive to TCGF. Furthermore, the results demonstrate that an active "response" by the resting T cells is required for expression of functional growth receptors, as demonstrated by the fact that: 1) a 4-hr pulse of concanavalin A (Con A) at 4 degrees C did not result in gain of reactivity to TCGF, whereas a 4-hr pulse at 37 degrees C did; 2) this metabolic requirement for acquisition of responsiveness to TCGF was not due to a secondary requirement for cap-formation of Con A-binding membrane structures, as normal responses were observed in the presence of cytochalasin B (cyt B); 3) the process of Con A-induced acquisition of susceptibility to TCGF was puromycin sensitive.     

Regulation of T-cell activation and T-cell growth factor (TCGF) production by hydrogen peroxide

Activated macrophages are known to release a variety of immunoregulatory substances including the low-molecular-weight substances hydrogen peroxide and lactate. We report here that lactate but not hydrogen peroxide is capable of supporting a substantial production of T-cell growth factor (TCGF) in cultures of accessory cell-depleted splenic T-cell populations after stimulation with concanavalin A. Hydrogen peroxide and its biosynthetic precursor superoxide anion (O2-) mediate, however, a strong augmentation of the TCGF production by accessory cell-depleted T-cell populations in the presence of lactate. Lactate inhibits the incorporation of [3H]thymidine in short-term cultures (18-26 hr) of accessory cell-depleted T cells. This confirms the rule that (optimal) production of T-cell growth factor requires a growth inhibitory signal. Concentrations of hydrogen peroxide which augment TCGF production most effectively (i.e., 1 X 10(-5) M) do not inhibit the incorporation of [3H]thymidine; and higher concentrations (3 X 10(-5)-1 X 10(-4) M) of hydrogen peroxide inhibit both the production of TCGF and the incorporation of [3H]thymidine. In agreement with the augmenting effect of hydrogen peroxide on TCGF production, it was observed that the proliferative response in mixed lymphocyte cultures is suppressed by catalase and augmented by 1 X 10(-5) M H2O2. Proliferative and cytotoxic responses in mixed lymphocyte cultures with an external source of interleukin 2 (IL-2) in contrast, are not augmented by 1 X 10(-5) M H2O2. The relatively high concentration of 1 X 10(-4) M hydrogen peroxide was found to inhibit the proliferative responses in mixed lymphocyte cultures with or without external IL-2 but not the cytotoxic response in the presence of IL-2. This indicates that CTL precursor cells may be relatively resistant against H2O2.     

Bacteriolytic effect of cessation of glucosamine supply, induced by specific inhibition of glucosamine-6-phosphate synthetase

The antibiotic tetaine (bacilysin) and its C-terminal epoxyaminoacid--anticapsin--are powerful inhibitors of glucosamine-6-phosphate synthetase (EC 5.3.1.19.) in cell-free extracts of Escherichia coli K-12. Tetaine acts on growing cells as a bactericidal agent. This bactericidal action, measured from 10 to 160 muM concentration, is a consequence of the induction of lysis of growing cells. The induction of lysis by tetaine is compared with the lytic action of some beta-lactams. Hypertonic medium, destruction of the antibiotic, presence of chloramphenicol or the addition of N-acetylglucosamine protect E. coli K-12 cells against lysis induced by tetaine. These effects are compared with those observed in the presence of penicillin G. The results indicate that inhibition of early or late stages of peptidoglycan synthesis all result in more or less the same consequence, i.e. death via cell lysis.     

Inhibition of murine suppressor cell function by progesterone

The effects of progesterone on murine suppressor cell function generated in allogeneic MLCs were investigated. BALB/c splenic lymphocytes stimulated in vitro with C3H/He cells significantly suppressed the proliferative response of BALB/c lymphocytes in a secondary MLC. This suppression was highly specific for the sensitizing alloantigens since the suppressor cells had no effect on the proliferative response of BALB/c lymphocytes to third-party alloantigens. In addition, BALB/c lymphocytes stimulated with syngeneic cells were observed to nonspecifically suppress the MLC response to a lesser extent. One to 10 micrograms/ml progesterone added at initiation to suppressor cell generating cultures diminished the ability of both alloantigen specific and nonspecific suppressor cell populations to suppress the proliferative response of homologous lymphocytes to alloantigens. Experiments with pyrilamine, an antihistamine, which blocks cytotoxic T lymphocyte (CTL) generation, suggests that progesterone has a direct inhibitory effect on suppressor cell function independent of its ability to block CTL induction. The effects of progesterone on suppressor cells were not due to shifts in peak response time in MLC or induction of radiosensitive cells in progesterone-treated cultures. Estradiol at doses between 5 and 10 micrograms/ml, and cortisol at dose of 1 microgram/ml, also significantly inhibited suppressor cell function. These results suggest that the steroid hormone milieu within the placenta may effect the activity of allogeneic or nonspecific suppressor cell activity.     

Immunosuppressive properties of murine trophoblast

The modification of the immunological response by murine trophoblast cells of different sources was investigated using the mixed lymphocyte reaction (MLR) and the cell mediated lympholysis (CML) test. MLR between C57BL (H-2b) stimulator splenocytes (mitomycin C treated in the unidirectional MLR) and BALB/c (H-2d) responder lymph node cells were markedly suppressed by trophoblast of ectoplacental cone (EPC) and placental origin. The same in vitro effect was observed with supernatants (SN) of trophoblast cells and with supernatants of blastocysts. Addition of anti-progesterone serum (APS), anti-testosterone serum (RAT), and anti-immunoglobulin serum (RAHIg) in serial dilutions to the trophoblast-MLR system revealed that the immunosuppressive effect of trophoblast giant cells and trophoblast giant cell culture supernatants can be abolished with APS. Identical results were obtained with APS added to immunosuppressive doses of progesterone. CML between C57BL responder lymph node cells and mitomycin C-treated BALB/c stimulator spleen cells was also markedly suppressed when trophoblast of EPC origin was added. A similar suppression of cytotoxic T-cell induction was seen when progesterone was added to the system. The immunosuppressive action of trophoblast as detected in vitro is likely to play an important role in the maintainance of pregnancy by protecting the semiallogeneic conceptus against immune aggression by the maternal immune system.     

The PERK-eIF2α phosphorylation arm is a pro-survival pathway of BCR-ABL signaling and confers resistance to imatinib treatment in chronic myeloid leukemia cells

Activation of adaptive mechanisms plays a crucial role in cancer progression and drug resistance by allowing cell survival under stressful conditions. Therefore, inhibition of the adaptive response is considered as a prospective therapeutic strategy. The PERK-eIF2α phosphorylation pathway is an important arm of the unfolded protein response (UPR), which is induced under conditions of endoplasmic reticulum (ER) stress. Our previous work showed that ER stress is induced in chronic myeloid leukemia (CML) cells. Herein, we demonstrate that the PERK-eIF2α phosphorylation pathway is upregulated in CML cell lines and CD34⁺ cells from CML patients and is associated with CML progression and imatinib resistance. We also show that induction of apoptosis by imatinib results in the downregulation of the PERK-eIF2α phosphorylation arm. Furthermore, we demonstrate that inactivation of the PERK-eIF2α phosphorylation arm decreases the clonogenic and proliferative capacities of CML cells and sensitizes them to death by imatinib. These findings provide evidence for a pro-survival role of PERK-eIF2α phosphorylation arm that contributes to CML progression and development of imatinib resistance. Thus, the PERK-eIF2α phosphorylation arm may represent a suitable target for therapeutic intervention for CML disease.     

Constitutive and mitogen-induced production of T cell growth factor by stable T cell hybridoma lines

Stable T cell growth factor- (TCGF; IL 2) producing cloned T cell hybridoma lines were constructed by fusing murine alloantigen-activated T cells with the 8-azaguanine-resistant lymphoma line, BW5147. Many, but not all, clones of one of these hybridomas, i.e., hybridoma 24, secreted TCGF constitutively, but production was markedly enhanced by stimulation with T cell mitogens. Large numbers of TCGF-secreting hybridoma cells in a stable functional state could be obtained from histocompatible mice inoculated with cloned T cell hybridomas. Moreover, such in vivo-derived hybridoma cells could be stimulated sequentially with mitogen at least twice to secrete their biologically-active product, resulting in larger TCGF yields from the same cells. The secreted product of these T cell hybridoma lines resembled TCGF isolated from other cellular sources in that it: a) supported the growth of a TCGF-dependent T cell line; b) provided help for the induction of alloantigen-reactive cytotoxic T lymphocytes from thymocyte precursors; c) facilitated concanavalin A-induced mitogenic responses of low thymocyte numbers; d) had an apparent m.w. of 30,000 to 40,000 by gel filtration chromatography; and e) was eluted from DEAE-Sephacel ion-exchange chromatography columns by salt concentrations of 30 to 150 mM NaCl. The ability of these T cell hybridomas to grow in vivo and retain their functional characteristics in a stable form should prove useful in terms of providing large numbers of TCGF-secreting cells and studying in vivo aspects of the production of TCGF as well as other immunoregulatory mediators.     

Temperature requirements and kinetics of T cell signaling. Velocity of triggering correlates with functional effects of anti-CD3 mAb

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.     

Blockade of physiologically secreted IFN-gamma inhibits human T lymphocyte and natural killer cell activation

The role of physiologically secreted human IFN-gamma in T lymphocyte and NK cell activation has been probed with a panel of mouse mAb directed against various epitopes of the human IFN-gamma molecule, or human IFN-gamma R. Addition to the culture medium of those mAb that neutralize the antiviral activity of IFN-gamma or interact with its receptor inhibited proliferative and cytotoxic responses elicited in PBL by HLA alloantigens, anti-CD3 mAb, and IL-2, but not the proliferative response to PHA. The IFN-gamma blockade also inhibited IFN-gamma, IL-2, and TNF-alpha release during MLC. Kinetic experiments showed that reduction of proliferative and cytotoxic responses to HLA alloantigens is maximal when IFN-gamma is blocked within the first 48 h. Exogenous rIFN-gamma restored the proliferative response only when added at the beginning. Moreover, when IFN-gamma was blocked, T lymphocytes recovered from 6-day MLC displayed a profound decrease in their expression of p55 and p75 chains of the IL-2R, as well as in the number of high-affinity IL-2 binding sites. These findings strongly suggest that IFN-gamma is required in the early phases of induction of the oligo- and polyclonal proliferative and cytotoxic responses of lymphocytes.     

IL-10 inhibits mitogen-induced T cell proliferation by selectively inhibiting macrophage costimulatory function.

IL-10, a newly designated cytokine primarily produced by the Th2 subset of CD4+ T lymphocytes and Ly-1+ B lymphocytes, has recently been hypothesized to inhibit cytokine production by Th1 T cell clones by blocking accessory cell- (AC) dependent costimulatory function. To evaluate the effect of IL-10 on Con A-induced proliferative responses of resting murine T cells, purified T cells were cultured with different types of AC. The addition of IL-10 produced a 70 to 90% inhibition of resting T lymphocyte proliferation when purified populations of macrophages were used as AC, but had no effect on the AC function of T-depleted spleen cells, activated B cells, dendritic cells, or L cells. The inhibitory effects of IL-10 were inversely related to the concentration of mitogen and could be reversed by the addition of the neutralizing anti-IL-10 mAb, SXC1. The inhibition of macrophage AC function was not related to the induction of a suppressor cytokine as stimulation by mixtures of macrophages and limiting numbers of dendritic cells was not inhibited. The decrease in proliferative responses was primarily secondary to inhibition of IL-2 production although the failure of exogenous IL-2 to completely reconstitute the response suggested that IL-10 may also exert inhibitory effects on the induction of expression of a functional IL-2R. These results are most consistent with a model in which IL-10 inhibits the induction of expression on macrophages of a critical costimulatory molecule that may be constitutively expressed on other types of AC.     

The xenogeneic effect. III. Induction of cell-mediated cytotoxicity by alloantigen-stimulated thymocytes in the presence of xenogeneic reconstitution factor.

A supernatant from human mixed leukocyte cultures (MLC), Xenogeneic Reconstitution Factor (XRF), was added to one-way murine MLC. This supernatant greatly enhances the generation of cytotoxic cells from C57BL/6 or DBA/2 responder thymocytes, as assayed by a standard 51Cr-release assay. This enhancement is shown 1) to be dependent upon the presence of BDF1 semi-allogeneic stimulator cells and 2) to result in the generation of cytotoxic cells that are specific for the H-2 type of the stimulating alloantigen. In some experiments, alloantigen-stimulated murine spleen cells, when cultured with XRF showed increased induction of cell-mediated cytotoxicity. These experiments show that XRF contains a cytotoxic-cell activating factor(s) that is functionally similar to those found in murine-derived supernatants, and that this factor can participate in the generation of cytotoxic cells from precursors present in the murine thymus.     

TGF-beta inhibits growth factor-induced DNA synthesis in hamster fibroblasts without affecting the early mitogenic events

Transforming growth factor beta (TGF-beta) was found to inhibit (IC50 = 0.1 ng/ml) alpha-thrombin or FGF-induced mitogenicity in G0-arrested Chinese hamster lung fibroblasts. Growth factor-stimulated cells became rapidly insensitive to TGF-beta addition during their progression through G0/G1 suggesting that an early step of the mitogenic response was the target of TGF-beta action. Surprisingly, none of the well characterized early mitogenic events commonly triggered by growth factors was found to be affected by TGF-beta addition. These responses included: phosphoinositide breakdown, activation of protein kinase C as determined by EGF receptor down-modulation, subsequent rises in pHi, c-fos, and c-myc mRNA levels, ribosomal protein S6 phosphorylation, the increase in RNA and protein synthesis, induction of ornithine decarboxylase. Only the induction of thymidine kinase, a marker of entry in the S phase, was found to be repressed by TGF-beta, with maximal inhibition when TGF-beta was added early in G1. These results indicate that the inhibitory action of TGF-beta does not affect the growth factors signalling pathways but touches an early event different from those so far analyzed.     

Serum-induced inhibition of isotonic fluid absorption by the kidney proximal tubule. III. Further evidence that complement-mediated cell lysis is involved.

The results of our previous studies have suggested that serum-induced inhibition of proximal tubular fluid absorption is due to complement-induced lysis of the tubular cells. The present study provides further evidence in support of this idea as well as other information pertinent to the mechanism of complement activation in vivo. 1. The electrical resistance of the luminal brush border membrane is reduced drastically concomitantly with a drop in cell potential difference when serum is perfused intraluminally. 2. Human C1 inhibitor (30-50 units/ml) does not significantly affect the inhibitory activity of human serum on fluid absorption, suggesting the non-involvement of the classical pathway. 3. Reactive lysis reagents (C56, C7, C8 + C9) partially inhibit fluid absorption when infused into the lumen. 4. In contrast to our previous report (Sato, K. and Ullrich, K.J. (1974) Biochim. Biophys. Acta 354, 182-187), very fresh serum, 10-times diluted can inhibit fluid absorption if perfused for 10 min. 5. Both mouse and guinea pig serum, which are normally inactive, are activated to attack the tubular cells if 1/100 volume rat or rabbit serum is added to them No such activation occurs by mixing guinea pig serum and mouse serum. The available data suggest that the presence of the later complement components but not the heat-labile factor (Factor B) or C3PA or C1 in the added serum is a prerequisite for mouse and guinea pig sera to be activated to inhibit fluid absorption.     

The mechanism of cell-mediated cytotoxicity. III. Protease-specific inhibitors preferentially block later events in cytotoxic T lymphocyte-mediated lysis than do inhibitors of methylation or thiol-reactive agents

Highly active mouse cytotoxic T lymphocytes (CTL) generated in secondary mixed-lymphocyte responses were used to examine the manner in which adenosine derivatives, thiol-specific reagents, or protease-specific probes affected CTL-mediated lysis (CML). The adenosine deaminase inhibitor deoxycoformycin (dCF) enhanced inhibition by adenosine (AR) or by deoxyadenosine (AdR), but not by 7-deazaadenosine (tubercidin). L-Homocysteinethiolactone (L-Hcy) acted synergistically with AR, but not with AdR or tubercidin, to block CML. Thus, AR derivatives may act both by affecting cellular methylation reactions, as demonstrated by the synergism between AR and L-Hcy, and by inhibiting other events required for CML. Conditions were then established to determine whether these reagents preferentially affected either the Ca2+-independent initial stage of cytolysis or the subsequent Ca2+-dependent events. Methylation inhibitors blocked lysis most effectively if added before effector-target binding. Similarly, the nonpenetrating thiol-specific reagent quaternary ammonium monobromobimane (qBBr) was more inhibitory when added prior to the Ca2+-dependent stage. Protease inhibitors such as alpha-1-antichymotrypsin and protease substrates such as acetyltyrosine ethyl ester (ATEE) or tyrosine ethyl ester (TEE) also inhibited CML. But, in contrast to qBBr or methylation inhibitors, neither TEE nor ATEE was more effective when added prior to the initial effector-target interaction. Furthermore, TEE did not appreciably affect CTL binding to target cells at concentrations that nearly abrogated CML. Thus, the implicated protease step is unique in that it does not appear to participate in recognition or binding.     

Activation of antigen-enriched B cells. II. Role of linked recognition in B cell proliferation to thymus-dependent antigens

The responsiveness to T-dependent (TD) and T-independent (TI) TNP-antigens of murine splenic B cells previously enriched for antigen-binding cells (ABC) was examined. TNP-TI antigens induced B cell proliferation. TNP-TD antigens did not induce a proliferative response regardless of the physical form or nature of the TNP-TD antigen (e.g., soluble vs particulate, low or high haptenation of carrier, TNP on various insoluble matrices, etc.). TNP-TD antigens were effective in enhancing the response of the TNP-ABC to all concentrations of lipopolysaccharide (LPS) tested, indicating that binding of antigen to surface immunoglobulin alters the LPS responsiveness of the cell. Irradiated, keyhole limpet hemocyanin- (KLH) primed T cells induced a threefold to fourfold greater B cell proliferative response with TNP-KLH than with fluoresceinated KLH (FLU-KLH) or FLU-KLH together with TNP-human serum albumin (TNP-HSA). Therefore, linked recognition appears essential for optimal T cell-mediated B cell proliferation, whereas the induction of B cell proliferation via nonlinked, carrier-activated T cells is a minor component of the response.