Treatment Of Spleen Cells Research Articles

Cellular origins and target cells of immunoregulatory factors in mice with disseminated histoplasmosis.

Disseminated infection with Histoplasma capsulatum stimulates the production of a suppressor factor (SF-H) by spleen cells from C3H/HeJ mice and a helper factor (HF-H) by spleen cells from C57BL/6 mice. In the present study these disparate immunoregulatory factors were analyzed in detail with regard to: a) the surface phenotype of the cells that produce SF-H and HF-H; b) the role of accessory cells in the production of these factors; and c) the surface phenotype of the target cells activated by SF-H and HF-H. Treatment of spleen cells from Histoplasma-infected C3H/HeJ mice with anti-Thy-1.2 plus complement (C) or with anti-Ly-2 plus C or with anti-I-Jk antiserum plus C abolished production of SF-H. Conversely, generation of HF-H by spleen cells from C57BL/6 mice was abrogated by treatment with either anti-Thy-1.2 plus C or with anti-Ly-1 plus C. Thus, a Thy-1.2+, Ly-2+, I-J+ T cell releases SF-H, and a Thy-1.2+, Ly-1+ T cell secretes HF-H. Production of SF-H and HF-H by splenic T cells was reduced markedly by depletion of macrophages (M phi); readdition of 1% syngeneic, plastic-adherent splenocytes from normal or infected mice to M phi-depleted, splenic T cell cultures of either strain restored the capacity to generate immunoregulatory factors. Furthermore, adherent splenocytes from normal or infected mice liberated a factor or factors that enhanced production of both SF-H and HF-H. Kinetic studies demonstrated that activation of normal spleen cells required at least 48 hr of exposure to SF-H or HF-H. Both factors failed to activate splenocytes pretreated with anti-Thy-1.2 plus C. Spleen cells from C3H/HeJ mice depleted of Ly-1+, Ly-2+, or I-J+ cells and exposed to SF-H did not demonstrate suppressor activity, whereas Ly-1-depleted splenocytes from C57BL/6 mice exposed to HF-H failed to exert helper activity. Therefore, the target of SF-H is a Thy-1.2+, Ly-1+2+, I-J+ T cell, and the target of HF-H is a Thy-1.2+, Ly-1+ T cell.

Isolation of human splenic macrophages and lymphocytes by countercurrent centrifugal elutriation

Certain tissues, such as the spleen, are rich sources of mononuclear phagocytes (MP); however, separating the phagocytes from tissues and removing the contaminating cells have been difficult. We report here a method for the extraction and purification of human splenic MP that employs gentle homogenization of splenic fragments with a Tenbroeck tissue homogenizer, controlled digestion with purified collagenase to free MP from splenic stroma, incubation with DNase to dissociate cell clumps and purification by countercurrent centrifugal elutriation (CCE). With homogenization and enzymatic digestion most of the splenic nonspecific-esterase-positive cells are freed into suspension as determined by morphometric analysis of 2 μm sections from plastic embedded spleen stained for α-naphthyl butyrate esterase (ANB). Overall cell recovery after homogenization and enzyme treatment is 56±7%; no selective cell loss occurs as determined by differential cell counts at each purification step. CCE of up to 3×10 9 treated spleen cells results in recovery of 63±6% of the elutriated cells and separates nearly 50% of the recovered MP into enriched fractions. These MP are morphologically intact as determined by light and electron microscopy and are actively phagocytic. Highly purified (96%) autologous splenic lymphocytes are a useful by-product of this separation technique.

Demonstration of long-lived memory T suppressor cells in the IgE response.

Administration to mice of repeated small doses of glutaraldehyde-polymerized ovalbumin (OA-POL) results in drastic suppression of the IgE antibody response. The high anamnestic IgE antibody responses usually obtained after 2 injections of native OA in alum were completely abrogated when OA-POL was administered between the 2 immunizations. Treatment with OA-POL prior to a primary immunization with 2 micrograms DNP-OA in alum induced a profound suppression of both anti-DNP and anticarrier IgE responses. The suppression, which was detectable for at least 7 months, was antigen-specific, selective for the IgE class and was sensitive to treatment of spleen cells with anti-Thy-1.2 antibody and complement. Adoptive transfer of spleen cells from donors treated with OA-POL 200 days, 16 days or 200 and 16 days prior to transfer, resulted in different kinetics of suppression in the recipient. The results are consistent with the presence of long-lived IgE-selective T suppressor cell memory. The possibility that separate Ts and Ts memory cell populations are generated is discussed.

Activation of murine B lymphocytes by Neisseria meningitidis and isolated meningococcal surface antigens.

Heat-killed Neisseria meningitidis was found to be a potent mitogen for mouse splenic lymphocytes. Results obtained with different cell separation techniques indicated that the bacteria acted to selectively induce proliferation of B lymphocytes. First, partial or total depletion of T lymphocytes by treatment with various anti-T-cell antisera plus complement did not affect the ability of the remaining spleen cells to proliferate in response to N. meningitidis. Second, T lymphocytes purified by affinity chromatography through an immunoglobulin-antiimmunoglobulin-coated glass bead column were unresponsive to meningococcal stimulation, even when provided with a source of macrophages (irradiated or mitomycin C-treated spleen cells). Finally, treatment of spleen cells with soy bean agglutinin showed that, whereas the soy bean agglutinin-positive population (B-enriched lymphocytes) was highly responsive to stimulation by N. meningitidis, the soy bean agglutinin-negative population (T-enriched lymphocytes) displayed only a background level of proliferation when exposed to the bacteria. Isolated meningococcal surface antigens such as lipopolysaccharide (LPS) and outer membranes also possessed mitogenic activity and induced proliferation of B lymphocytes in a dose-dependent manner. Both LPS and non-LPS components contributed to the mitogenicity of outer membranes since the addition of outer membrane preparations to spleen cells from the low LPS responder C3H/HeJ mouse strain gave rise to a high level of proliferative activity.

Modulation of granulomatous hypersensitivity. VI. T lymphocyte subsets influence mast cell density in liver granulomas of Schistosoma mansoni-infected mice.

Abstract In murine schistosomiasis mansoni, a complex series of cell-cell interactions involving T cell subclasses regulates the intensity of the inflammatory granulomatous response. Recent evidence suggests that granuloma mast cells also participate in the regulatory process by the release of histamine. The current study was performed to determine factors that affect the number of granuloma mast cells. More mast cells were detected in liver granulomas from chronically (20-wk) as opposed to acutely (8-wk) infected mice. Adoptive transfer of spleen cells from 20-wk-infected donors into acutely (6-wk) infected recipients increased granuloma mast cell density. Treatment of spleen cells with anti-Thy-1.2 or anti-Lyt-1.1 antiserum and complement, but not anti-Lyt-2.1 or normal mouse serum, abrogated adoptive transfer-induced, augmentation of granuloma mast cell density. Treatment of acutely infected animals with cyclophosphamide or cimetidine (H2 antagonist enhanced granuloma mast cell density. These data suggest that granuloma mast cell density is dependent upon subsets of T lymphocytes.

Effect of Mitogenic Stimulation of Murine Splenocytes on PEG-Induced Cell Fusion

A cell fusion assay system was devised as a means of measuring cell activation. Polyethylene glycol (PEG) was used to induce fusion between spleen cells of various mouse strains and the BW5147 thymoma cell line. A fusion index (FI) was calculated by determining the ratio of the number of nuclei in fused cells to the number of nuclei in all cells and multiplying by 100 (the FI could range from 0 to 100). Spleen cells from BALB/c mice were compared in PEG-induced fusion assays. BALB/c spleen cells stimulated with phytohemagglutinin, leucoagglutinin, concanavalin A, pokeweed mitogen and LPS showed FI two- to three-fold higher than those found in unstimulated cultures, indicating that stimulated cells fuse at much higher rates. This response is mitogen dose-dependent and parallels DNA synthesis as measured by 3H-TdR incorporation. Treatment of spleen cells with cycloheximide 12 h prior to fusion had no effect on FI. In vivo and in vitro stimulation of BALB/c, C57BL/6 and NZW mice with LPS resulted in enhanced FI. This was not the case in low-responder DBA/2 and nonresponder C3H/HeJ animals.

Stimulation of mouse lymphocytes by a mitogen derived from Mycoplasma arthritidis. III. Ir gene control of lymphocyte transformation correlates with binding of the mitogen to specific Ia-bearing cells.

The supernatant from Mycoplasma arthritidis broth cultures (MAS) contains a T cell mitogen that is under Ir gene control. Responsiveness to this mitogen is dictated by the I-E/I-C subregion of the major histocompatibility complex and is dependent upon adherent radioresistant Ia+ accessory cells from responding haplotype animals. In this study, we established that MAS could be removed from culture supernatants by absorption with spleen cells from mice that themselves are responsive to the mitogen (k and d haplotypes), but activity is not removed by spleen cells from mouse strains that are nonresponsive to the mitogen (b, q, and s haplotypes). Absorption studies with lymphoid cells from congenic and recombinant strain mice established that absorption of the mitogen was itself linked to the I-E/I-C subregion of the major histocompatibility complex. Thymocytes from responding haplotype strains were incapable of removing MAS activity, and spleen cells devoid of Thy-1-positive cells retained their full absorbing capacity. The ability to effectively absorb MAS activity was abrogated by the pretreatment of spleen cells with anti-Ia antiserum and complement. Furthermore, the ability of spleen cells from responding haplotype strains to respond to MAS was blocked by the addition of anti-Ia serum to the cell cultures. Whereas the latter treatment resulted in an almost complete elimination of MAS responsiveness, the ability of similarly treated spleen cells to respond to the mitogens PHA and Con A was only minimally depressed. These results are consistent with our hypothesis that the mitogenic moiety of MAS actually binds to I-E/I-C-coded Ia antigens.

Activity and dexamethasone sensitivity of natural cytotoxic cell subpopulations

Natural cytotoxic (NC) activity is the natural cell-mediated cytotoxic activity of normal spleen cells directed at fibroblast targets. Treatment of spleen cells with dexamethasone, a synthetic glucocorticosteroid, reveals two populations of NC effectors: one population whose cytolytic activity is inhibited by dexamethasone (DEX S) and another which is resistant to inhibition by dexamethasone (DEX R). The short-term culturing of spleen cells results in a shift in the population of splenic NC effectors from predominantly DEX S to predominantly DEX R. This shift also occurs when spleen cells are treated with conditioned medium from which the short-term cultured DEX R populations were derived.

Inhibition of antibody synthesis by histamine in concanavalin A-treated mice: the possible role of glucocorticosteroids.

Administration of histamine (50 mg/kg) to BALB/C mice injected with concanavalin A (Con A) (100 micrograms, i.v.) 24 hr previously, results in a marked decrease in antibody synthesis to sheep red blood cells (SRBC) injected 2 hr later. This phenomenon occurs with nonimmunosuppressive doses of Con A and is strain-specific. It does not take place in the response to the T-independent antigen polyvinylpyrrolidone (PVP) or if histamine is administered after the antigen. Adoptive transfer of normal syngeneic cells at the same time as antigen does not reverse this effect. Excess suppressor cell generation was excluded by co-cultivation of treated spleen cells with normal cells in vitro and by determining their antibody response to SRBC 5 days later. 2-Methylhistamine, a histamine type 1 (H1) receptor agonist, mimicks the effect of histamine whereas dimaprit, a histamine type 2 (H2) receptor agonist, does not. Because histamine interaction with H1 receptors causes the release of adrenocorticotropic hormone (ACTH), we examined the effects of ACTH and corticosterone in this system and found that both could mimick the effect of histamine. These results suggest that the interaction of histamine with H1 receptors causes the release of glucocorticosteroids that may interfere with either Con A-activated T helper cell function or macrophage processing of T-dependent antigen.

Enhanced production of murine interferon gamma by T cells generated in response to bacterial infection.

Spleen cell cultures derived from animals infected 6 d earlier with Listeria monocytogenes produced 10-20-fold more murine interferon gamma (MuIFN gamma) than spleen cells from nonimmune mice in response to stimulation with T cell mitogens. A striking temporal association was found between the enhanced synthesis of MuIFN gamma and the development of anti-Listeria immunity in that both the potential for increased MuIFN gamma production and the generation of Listeria-protective T cells developed and then decayed in unison. Treatment of spleen cells with monoclonal anti-Thy-1.2 plus complement virtually abolished the ability of cells from Listeria-immune mice to synthesize MuIFN gamma. The T cells producing MuIFN gamma were found to be more susceptible to complement-mediated lysis with monoclonal anti-Lyt-1.2 than with monoclonal anti-Lyt-2.2. The production of MuIFN gamma was not affected by treating spleen cells with anti-IgG antisera or with a monoclonal antibody directed against I-A specificities. MuIFN gamma was detected 4 h after the beginning of mitogenic stimulation of spleen cell cultures, and peak levels of MuIFN gamma were reached by 18 h. The IFN synthesized by mitogen-induced spleen cells derived from Listeria-immune mice were relatively labile at pH 2.0 and neutralized by a rabbit anti-MuIFN gamma serum but not by an antiserum having specificities for MuIFN alpha and MuIFN beta. The apparent molecular weight of the MuIFN gamma, as estimated by molecular sieving on a Bio-gel P-60 column, was estimated to be 38,000, and the isoelectric point as determined by chromatofocusing was extremely heterogeneous, ranging between pH 5.0 and pH 7.0.

Activation of a suppressor T-cell pathway by interferon.

In addition to antiviral activities, murine fibroblast (type I) interferon (IFN-beta) suppresses immune responses. The mechanism(s) by which IFN-beta suppresses antibody responses by murine spleen cells to sheep erythrocytes in vitro has been investigated. IFN-beta-mediated suppression is partially or completely prevented by catalase, 2-mercaptoethanol, and certain peroxidase substrates (ascorbic acid, potassium iodide, and tyrosine). These same reagents also block suppression by mediators from concanavalin A-activated murine suppressor T cells, soluble immune response suppressor (SIRS)/macrophage-derived suppressor factor (Mphi-SF), and act by inactivating Mphi-SF or preventing formation of Mphi-SF from SIRS. Therefore, these experiments suggested that IFN-beta may act by inducing production of a molecule that has properties of SIRS. Treatment of spleen cells with IFN-beta leads to generation of a population of Lyt2+ suppressor T cells that acts by elaborating a soluble factor. This IFN-beta-induced suppressor T-cell factor (IFN-TsF) has properties in common with SIRS. First, both SIRS and IFN-TsF suppress antibody responses with the same characteristic kinetic pattern; responses initiate normally but prematurely terminate after day 4 of culture. Second, IFN-TsF and SIRS are of comparable size (45,000-55,000 daltons) and are converted to Mphi-SF by low (1 microM) concentrations of H2O2 or by macrophages. Third, Mphi-SF obtained from IFN-TsF or SIRS is inactivated by similar concentrations of reagents such as ascorbic acid, potassium iodide, and 2-mercaptoethanol. These data show that the immunosuppressive properties of IFN-beta are due, at least in part, to its ability to activate suppressor T cells that produce mediators that appear to be analogous to those in the SIRS/Mphi-SF pathway of immunosuppression.

Cellular regulation of gamma interferon production: Lyt phenotype of the suppressor cell.

Suppressor cell activity for in vitro gamma interferon (IFNgamma) production can be induced in C57BL/6 mice with the T-cell mitogens staphylococcal enterotoxin A and concanavalin A. The suppressor cell lacked mitogen specificity in that injection of mice with either staphylococcal enterotoxin A or concanavalin A induced potent suppressor cell activity that inhibited the in vitro induction of IFNgamma by either mitogen. The suppressor cell is a T cell with a relatively high density of Thy 1 antigen. The IFNgamma-producing cell has a low density of Thy 1 and appears to have evolved from cells with a high density of Thy 1.2 or to have required such cells as helpers in the production of IFNgamma. Monoclonal anti-Lyt 2.2 antibody completely eliminated the suppressor cell activity, whereas anti-Lyt 1.2 partially, but significantly, reduced the suppressor cell activity. The suppressor cell, then, may be or may require a Lyt 1,2,3(+) cell with a relatively low density of the Lyt 1 antigen for activity. The Lyt phenotype of the IFNgamma- producing cells was examined for both virgin and suppressor cell populations. Treatment of cells with either anti-Lyt 1.2 or anti-Lyt 2.2 completely eliminated IFNgamma production in both virgin and suppressor cell cultures. Cocultivation of Lyt 1(+) and Lyt 2(+) cells in both virgin and suppressor cell cultures significantly restored the ability to produce IFNgamma when cells were stimulated with staphylococcal enterotoxin A. For the virgin cells, this seems to indicate clearly that both Lyt 1(+) and Lyt 2(+) cells are required for IFN production, but that Lyt 1,2,3(+) cells are not required. For the suppressor cell population this type of treatment resulted in IFNgamma production comparable to that of the virgin cell population. The treatment eliminated or depleted Lyt 1,2,3(+) cells from the suppressor cell population, thus providing additional evidence that a Lyt 1,2,3(+) cell is required for suppressor cell activity. Treatment of spleen cells with anti-Lyt antibodies at the time that they were involved in ongoing IFNgamma production showed that the IFNgamma-producing cell is Lyt 2(+). At least a 1:1 ratio of virgin to suppressor cells was required for optimal suppression of IFNgamma production by virgin spleen cells. This suggests that suppression involved cell-to-cell contact. Preliminary data showed that the suppressor cell activity could also be induced in in vitro cultures incubated with mitogen for 4 days. This is evidence that the suppressor cell is inducible and not necessarily recruited to the spleen from other lymphoid organs.

Concanavalin A-induced alteration of surface marker expression on murine T cells

The activation of suppressor T cells by Concanavalin A (Con A) was characterized by changes in expression of T cell surface markers. Treatment of normal mouse spleen cells with monoclonal antibody to Ly-1 prior to activation with Con A consistently reduced the resulting suppressor cell activity as assayed by mixed culture with primed spleen cells responding to sheep erythrocytes. Addition of indomethacin to anti-Ly-1 depleted spleen cells together with Con A had no effect on the resulting suppressor cell activity, whereas its addition to normal spleen cells together with Con A significantly abrogated suppressor T cell induction. In contrast, treatment of normal spleen cells with anti-Ly-1 following 24 h culture with Con A had no effect on resulting suppressor cell activity. Treatment of spleen cells either prior to or following culture with Con A with monoclonal antibody to Ly-2 completely abrogated suppressor T cell activity. Suppressor T cells activated by Con A were depleted significantly by fractionation on columns containing insolubilized histamine. In contrast, fractionation of normal spleen cells on histamine columns followed by culture with Con A had no effect on subsequent suppressor T cell induction. Furthermore, direct addition of histamine (5 × 10 −4M−5 × 10 −8M) to Con A preculture had no effect on resultant suppressor T cell generation, suggesting that possible immunodulatory actions of histamine may be expressed at the effector rather than during induction stages of Con A-induced suppressor T cell activation.

Taenia solium: mitogenic effect of larval extracts on murine B lymphocytes.

The effect of an extract (CE) obtained from Cysticercus cellulosae on the proliferation of lymphocytes was studied in cultures of murine spleen cells. The addition of CE to the cultures resulted in the highly significant uptake and incorporation of tritiated thymidine (3H-TdR) into DNA. This phenomena was dose-dependent, with doses lower and higher than the optimal concentration causing less marked effects. The kinetic peak of this response was found to occur on day 2 of culture. CE evoked a proliferative response in cultures of spleen cells from congenitally athymic (nu/nu) BALB/c mice. Cultures of bone marrow-derived (B) lymphocytes, generated by treatment of spleen cells with rabbit antithymocyte serum and complement, incorporated 3H-TdR to a degree similar to that of normal spleen cell cultures. CE did not induce the proliferation of thymocytes. To eliminate the possibility that the mitogenic effect of CE was due to LPS, we carried out experiments using Polymyxin B (PB). CE was mitogenic after treatment with PB which inactivated the LPD effect. In addition, CE elicited 3H-TdR uptake in spleen cells from the LPS nonresponsive C3H/HeJ mouse strain.

Lymphocytes express Ia antigens of foreign haplotype following treatment with neuraminidase.

Evidence is presented which indicates that neuraminidase (NA) treatment of spleen cells both destroys old Ia antigens and reveals new Ia specificities which are not normally expressed by splenocytes. It was found that NA treatment unmasked alien I-Ak-like specificities on A.TH (Is) spleen cells, and Is-like antigens on A.TL (Ik) spleen cells. These conclusions were based on direct testing of NA-treated targets with a range of alloantisera and on cell-absorption experiments. Furthermore, the cellular distribution of NA-exposed antigens resembled that of convential Ia antigens, the new antigens being expressed on more than 90 percent of splenic B cells and a subpopulation of splenic T cells. However, although some of the antigens exposed by NA on A.TH cells appeared to resemble the Ia. 3 and 15 specificities, additional antigens were involved which did not correlate with any previously described Ia antigens.

Induction of parasite-specific helper T lymphocytes during Trypanosoma cruzi infections in mice.

Development of parasite-specific T helper cells was examined in mice infected with Trypanosoma cruzi. At various times during the course of infection mice were challenged with TNP conjugated to fixed culture forms of T. cruzi (TNP-TC), and the resultant splenic plaque-forming cells (PFC) against TNP were determined. By day 10 post-infection significant responses against TNP-TC were observed but not against TNP-BSA. Infected mice that were not challenged with TNP-TC did not produce anti-TNP PFC, which demonstrated that the TNP-TC response was not the result of nonspecific B cell activation. Treatment of spleen cells from infected mice with anti-theta antiserum plus C ablated the anti-TNP-TC response when these cells were transferred to normal mice that were subsequently challenged with TNP-TC, whereas treatment of the cells with anti-Ig plus C prior to transfer had no effect on the TNP-TC response. These results demonstrate enhancement of parasite-specific Th activity of mice infected with T. cruzi and that cell-cell interaction in development of responses to neoantigens is fully functional when sensitized Th are present, even though the animals are unresponsive to heterologous antigens.

Specific unresponsiveness to skin allografts in anti-lymphocyte serum-treated, marrow-injected mice: participation of donor marrow-derived suppressor T cells.

Sequential changes of cell-mediated immune reactivities were examined in anti-lymphocyte serum-(ALS) treated, C3H/He (C3H; H-2k) bone marrow-injected (C57BL/6 X A)F1 (B6AF1; H-2b/k.d) mice bearing enhanced C3H skin grafts. Spleen cells of these mice exhibited marked suppression of the proliferative response to phytohemagglutinin and concanavalin A. When the spleen cells were assayed for the direct lymphocyte-mediated cytotoxicity against H-2k targets, their lytic activity remained low until the time of graft rejection, in contrast to the increasingly high cytotoxic activity exhibited by spleen cells of control B6AF1 mice given only ALS and C3H skin grafts. When spleen cells of marrow-injected B6AF1 mice were cultured with mitomycin-C treated C3H spleen cells, the proliferative response was significantly suppressed the throughout the course, despite the early appearance of high "secondary-type" cytotoxic activity. Co-culture experiments demonstrated the presence of C3H antigen-specific suppressor cells in the ALS-treated, marrow-injected mice bearing intact allografts. Treatment of spleen cells with anti-H-2, anti-Thy 1 and anti-I-J sera and C revealed that the suppressor cells present late in the marrow-injected mice were T cells of donor C3H bone marrow cell origin.

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Lethal growth of a syngeneic transplanted tumor (KMT-17) was inhibited in inbred WKA rats pretreated with the antileukemia drug busulfan (BU). However, the lethal growth of KMT-17 was not inhibited by pretreatment with cyclophosphamide, adriamycin, or ftorafur. With the Winn assay, spleen cells from BU-pretreated KMT-17-bearing rats (TBR) inhibited the growth of admixed KMT-17 cells more strongly than did spleen cells from BU-untreated TBR. The augmented tumor inhibitory activity of spleen cells was KMT-17-specific, and this activity was abrogated by in vitro treatment of spleen cells with anti-T serum and guinea pig complement. Augmentation of the immune response to KMT-17-associated antigen(s) in BU-pretreated TBR was also demonstrated in lymphocyte-mediated cytotoxicity, as detected by a 51Cr release assay and by a delayed-type hypersensitivity with a radioisotope footpad assay. Tumor regression in BU-pretreated rats was demonstrated to be mediated by the augmentation of T-cell-mediated immune responses to tumor-associated antigens. The tumor inhibitory effect of BU was abrogated by adoptive transfer with thymus cells from normal rats but not with those from BU-pretreated rats 1 day before tumor inoculation. The augmentation oif the antitumor immune responses by pretreatment with BU was suggested to be due to the fact that BU selectively inhibited the suppressor cells of their precursors.

Studies of the mechanism of action of diketocoriolin B to enhance antibody formation.

Diketocoriolin B (DKC), at a very low dose (0.1 micrograms/mouse) or in a low concentration (0.01 ng/culture) augmented antibody formation to sheep red blood cells (SRBC) in vivo and in vitro. The addition of DKC to spleen cell cultures 24 approximately 48 hours after the start of the culture showed the strongest effect. The stimulatory effect was not influenced by the elimination of macrophages. DKC also augmented antibody formation in T cell-depleted spleen cell cultures and in cultures of spleen cells of athymic mice. The treatment of spleen cells with DKC at 37 degrees C or 4 degrees C for 30 minutes before the start of the culture enhanced antibody formation to SRBC. both treatments of macrophage-rich and lymphocyte-rich cells with DKC enhanced antibody formation.

Concanavalin A-induced resistance to Listeria monocytogenes and activation of macrophages: defect in mice treated with 89Sr.

89Sr-treated mice injected with concanavalin A (Con A) 24 h prior to infection with Listeria monocytogenes (LM) could not enhance the clearance of LM from the spleen. Adoptive transfer of normal syngeneic spleen cells together with Con A rendered these animals more resistant. Spleen cells of 89Sr-treated or age-matched control mice were stimulated with con A for 24 h, and supernatant fluids were assessed for macrophage-activating factor (MAF), i.e. the ability to activate resident peritoneal macrophages to kill LM intracellularly in vitro. A defective MAF production by spleen cells was observed in 89Sr-treated, 2 week-old, and athymic nude mice. Also, treatment of spleen cells with anti-Thy-1.2 antiserum plus complement inhibited MAF production. Synergism between spleen cells from 89Sr-treated and nude mice did not occur. The cells required for MAF production were relatively resistant to gamma irradiation. Nylon wool filtration did not modify the ability of spleen cells to make MAF. 89Sr-treated mice possess macrophages responsive to MAF derived from normal spleen cells. The data suggest that the failure of 89Sr-treated mice to develop an anti-LM response observed in thi system could be due to a defective capacity to produce protective humoral factors and/or cells in response to Con A.