Number Of Cells In Lymph Nodes Research Articles

Effects of the color additive caramel color III and 2-acetyl-4(5)-tetrahydroxybutylimidazole (THI) on the immune system of rats

Administration of ammonia caramel color (AC) to rats may decrease blood lymphocyte counts, specifically in rats fed a diet low in vitamin B 6. This effect is associated with 2-acetyl-4(5)-(1,2,3,4-tetrahydroxybutyl)imidazole (THI). To characterize and compare the effects of AC and THI and to study the influence of dietary pyridoxine, two studies in rats were conducted. Weanling rats fed a diet containing 2–3 ppm pyridoxine and exposed to 4% AC or 5.72 ppm THI in drinking water for 4 weeks showed reduced cell numbers in spleen and popliteal lymph nodes, as well as in the blood. Flow cytometric analyses demonstrated a comparable reduction in B and T lymphocytes. In blood, spleen, and popliteal lymph nodes, CD4 + lymphocytes were more reduced than CD8 + cells. The number of bone marrow cells was not affected. Although thymus weight and cell number were not affected either, a decreased cortex over medulla area ratio and an increase in medullary cell density largely due to an increase in CD4 + thymocytes was observed. Decreased numbers of ED2 + macrophages were observed in the thymic cortex and in the spleen red pulp. All effects observed were either less pronounced or absent in a study with rats fed a diet containing 11–12 ppm pyridoxine. The effects of AC and THI on the immune system were similar. Whereas AC exposure was associated with changes in vitamin B 6 status, THI did not induce obvious effects on vitamin B 6 parameters. It is proposed that the effects of AC and THI on the immune system are not caused by a disturbance of vitamin B 6 metabolism, but may in fact result from a disturbance of a specific PLP-dependent process.

Prethymic Nylon Wool‐Passed Bone Marrow Cells, Substituting for Helper T Cells, Can Augment the Generation of Cytotoxic T Lymphocytes from Their Precursors

Nylon wool-passed bone marrow (NW-BM) cells treated with anti-Thy.1 monoclonal antibody and complement were added to a mixed lymphocyte culture which contained a limiting number of lymph node cells, as responder cells, and a sufficient number of mitomycin-c-treated allogeneic spleen cells as stimulator cells. NW-BM cells of the same MHC haplotype as responder cells enhanced the generation of allo-specific cytotoxic T lymphocytes (CTL) not only at a relatively high dose (3 x 10(3) cells/well) of responder cells, but also at an extremely dilute dose (1 x 10(3) cells/well). NW-BM cells which had a third-party MHC haplotype, a haplotype different from both responder and stimulator cells, also enhanced the generation of CTL at relatively high doses, but not at low doses, of responder cells. NW-BM cells which had MHC haplotypes identical with those of responder cells induced CTL from helper T cell-depleted responder cells, but NW-BM cells which had the third-party haplotype did not. These results showed that the enhancing effects of NW-BM cells of the same MHC haplotype as responder cells might be due to a specific helper effect and the enhancing effect of NW-BM cells of the third-party haplotype might be due to a nonspecific filler effect, which only conditioned the cultured cells. It was also found that, to exhibit the helper effect, NW-BM cells had to possess MHC class II, but not MHC class I, molecules in common with CTL precursors. This study showed that in the induction of CTL, prethymic NW-BM cells had a capability comparable to that of mature helper T cells.

Lymphocyte Subsets in the Lymph Nodes of Rats With Autoimmune Orchitis

We determined temporal variations of cell subpopulations in the immunization draining lymph nodes during the development of an experimental autoimmune orchitis (EAO) induced in Wistar rats. A phenotypic characterization of T cells and their subsets (CD4+ and CD8+), B, and Ia+ cells was performed by immunofluorescent technique. At the end of the immunization period (30 days), rats injected with testicular homogenate plus adjuvants presented a considerable increase in absolute cell number but normal lymphocyte subset percentages. Testicular damage became evident at 50 days after the first immunization and increased its severity at 80 days: animals that developed EAO presented a lower number of CD8+ cells as compared with undamaged rats. This latter group showed a low CD4/CD8 ratio due to the high proportion of CD8+ cells, which could probably have a suppressor function. At 80 days massive testicular infiltration and decreased absolute cell number in lymph nodes suggest the possible migration of specific lymphocytes to the target organ.

2,3,7,8-tetrachlorodibenzo- P-dioxin (TCDD) induced suppression of the local immune response

TCDD suppressed the normal immune response in popliteal and inguinal lymph nodes, when administered i.p. (50 μg/kg) to C57BL/6 mice, 4 days before immunization with the T-dependent antigen ovalbumin (10 μg/pad) in the hind foot pads. A hampered increase in lymph node cell number and a reduced frequency of antigen-specific B-cells were observed, despite the fact that cell proliferation in vivo was normal. While the restimulation of lymph node cells in vitro with ConA or LPS was normal, suggesting that the APC function was largely unaffected, the OVA-induced proliferation was greatly reduced. The anti-OVA antibody (ab) concentration both in serum and in supernatants of cultured lymph node cells was lower than in controls. In contrast, the production of anti-BSA ab upon LPS stimulation was normal. This indicates that the ability of the B-cells to produce ab and to proliferate was not disturbed. The DTH assay clearly showed an impaired T-cell function in TCDD-treated animals. Since APC or B-cells have appeared normal in their functions tested in this study, we propose that TCDD disturbed T-cell functions, leading to an impaired activation of B-cells.

Flow cytometric analysis of protein kinase C in lymph node cells during the induction phase of contact hypersensitivity reaction.

On the fourth day after a single painting of 2,4,6-trinitro-chlorobenzene on the abdominal skin, inguinal lymph nodes were removed, and a single-cell suspension was prepared. The cells were analyzed flow cytometrically, using monoclonal anti-protein kinase C antibody. It was found that the number of lymph node cells in which protein kinase C was detected on the cell surface was significantly increased over that in non-treated mice (p less than 0.01). On the basis of our results and discussions in the literature, it is thought that protein kinase C is related to the initiation of the contact hypersensitivity reaction.

Suppression of antigen-specific interleukin 2 production in the MRL/MpJ-lpr/lpr Mouse

Several murine strains with spontaneously occurring systemic lupus erythematosus-like disease demonstrate defects in immunoregulation. The MRL/MpJ-lpr/lpr (MRL-1) strain develops a severe age-progressive defect in interleukin 2 (IL 2) production in response to mitogen or antigen. In this study, we demonstrate in vitro the presence of suppressor cells in the lymph nodes of naive mice of the MRL background. Suppression by MRL-1 lymph node cells was partially reversed by treatment with anti-Lyt-1.2 monoclonal antibody and complement and was moderately radiosensitive. Suppression by lymph node cells from the congenic MRL/MpJ-+/+ (MRL-+) mouse was somewhat more resistant to treatment with anti-Lyt-1.2 and complement, or radiation. Lymph node cells from the H-2-syngeneic mouse strain, C3H/HeJ, failed to suppress. Thus, lymph nodes from mice of the MRL background contain cells capable of suppressing in vitro IL 2 responses. We next performed cell transfers to determine whether suppressor cells contribute in vivo to the IL 2 defect. Lymph node cells, but not spleen cells, from MRL-1 mice by 5 to 6 mo of age suppressed antigen-specific IL 2, CTL, and DTH responses when transferred into young MRL-+ recipients. Transfer of identical numbers of lymph node cells from age-matched MRL-+ mice failed to suppress IL 2 production. Transfer of suppression was sensitive to treatment with monoclonal anti-Lyt-2.1 and complement, and to 250 rad of radiation. Thus, this study suggests a role for active suppression of IL 2 production in the establishment of the IL 2 defect in the MRL-1 mouse. Further, suppression may involve phenotypically distinct T lymphocyte subpopulations.

Suppression of antigen-specific interleukin 2 production in the MRL/MpJ-lpr/lpr mouse.

Several murine strains with spontaneously occurring systemic lupus erythematosus-like disease demonstrate defects in immunoregulation. The MRL/MpJ-lpr/lpr (MRL-1) strain develops a severe age-progressive defect in interleukin 2 (IL 2) production in response to mitogen or antigen. In this study, we demonstrate in vitro the presence of suppressor cells in the lymph nodes of naive mice of the MRL background. Suppression by MRL-1 lymph node cells was partially reversed by treatment with anti-Lyt-1.2 monoclonal antibody and complement and was moderately radiosensitive. Suppression by lymph node cells from the congenic MRL/MpJ-+/+ (MRL-+) mouse was somewhat more resistant to treatment with anti-Lyt-1.2 and complement, or radiation. Lymph node cells from the H-2-syngeneic mouse strain, C3H/HeJ, failed to suppress. Thus, lymph nodes from mice of the MRL background contain cells capable of suppressing in vitro IL 2 responses. We next performed cell transfers to determine whether suppressor cells contribute in vivo to the IL 2 defect. Lymph node cells, but not spleen cells, from MRL-1 mice by 5 to 6 mo of age suppressed antigen-specific IL 2, CTL, and DTH responses when transferred into young MRL-+ recipients. Transfer of identical numbers of lymph node cells from age-matched MRL-+ mice failed to suppress IL 2 production. Transfer of suppression was sensitive to treatment with monoclonal anti-Lyt-2.1 and complement, and to 250 rad of radiation. Thus, this study suggests a role for active suppression of IL 2 production in the establishment of the IL 2 defect in the MRL-1 mouse. Further, suppression may involve phenotypically distinct T lymphocyte subpopulations.

Immunität der Haut — Infektion und perkutane Immunisierung von Kaninchen mit E. coli ATCC 13676

After intracutaneous infection of rabbits with a suspension of E. coli which was followed by a transient local inflammation, the local and systemic immune responses were determined using the lymphocyte stimulation test (LTT) and the hemolysis plaque assay (HPA). Lymphocytes of the lymphatic system draining the infected skin area and blood lymphocytes were used. With lymphocytes derived from the local lymph nodes, a substantial increase of specific stimulation in the LTT was detected beginning at day 3 after infection and lasting up to the termination of the experiment (3 weeks). Blood lymphocytes were stimulated at a lower level: The activity showed a peak at day 4 and an elevated level only during a 10-day period. After the intracutaneous infection with E. coli, increasing numbers of antibody-releasing lymph node cells were detected in the HPA. The antibody-secreting cells of the IgM and IgG classes clearly showed an increasing specificity for E. coli lipopolysaccharide coupled to sheep red blood cells. As with the LTT, the highest activities (values of specificity and number of plaque-forming lymphocytes) were observed at the end of the experimental period. An emulsified preparation of a heat-inactivated E. coli culture ( E. coli-BKS) which had been applied locally onto the artificially altered skin evoked a similar immunological response after a 2 or 3-weeks treatment. In such animals an increased activity of lymph node cells could be registered by LTT and HPA as compared to reactions from placebo-treated control animals. However, the topical immunization with nonviable E. coli stimulated not only lymphocytes which produced antibodies directed specifically against E. coli lipopolysaccharide as demonstrated by the HPA. An increased number of lymphocytes reacted even with native sheep red blood cells. This observations is discussed in respect of a polyclonal B-cell activation by lipopolysaccharide of the E. coli-BKS.

The arrest of T cell maturation in spontaneously hypertensive rats with a deficient production of thymic factors.

Spontaneously hypertensive rats (SHR) which regularly develop hypertension and periarteritis nodosa showed a progressive loss of T cell numbers and functions early in life. When six months old and compared with W rats, the original strain of SHR, they were found to possess a reduced number of thymocytes that formed rosettes with guinea pig erythrocytes and that reacted to anti-Thy1. 1 and anti-T (W3/13) sera. The number of spleen and lymph node cells which reacted to anti-T (W3/13) and anti-helper T (W3/25) sera also decreased. The antibody response of 3-month-old SHR spleen cells to SRBC was about one-fifth that of Wistar rats and progressively declined with age. Subcutaneous grafting of 3-month-old SHR thymus tissues failed to promote differentiation and functions of T cells in the spleen of nude mice, whereas 3-month-old W thymus tissues showed significant recovery of T cell generation and functions. The T cell functions of old SHR was completely restored by grafting adult W thymus tissues but was not restored by grafting adult SHR thymus tissues. Similar recovery was also obtained by the injection of thymus extracts from W thymus tissues. These results suggest that thymic epithelial cell products regulating T cell maturation may decrease in SHR with increasing age.

The cellular IgE response of rodents to infection with Nippostrongylus brasiliensis, Trichinella spiralis and Schistosoma mansoni

The IgE response at the cellular level to helminthic infection was studied in BALB/c mice inoculated with the infective larvae of the nematodes Nippostronglylus brasiliensis (Nb) or Trichinella spiralis (Ts) or with the cercariae of the trematode Schistosoma mansoni (Sm). Changes in mesenteric lymph node (MLN) cell number, cell surface(s) IgD, IgM, IgE and Thy-1.2 and intracytoplasmic (c) IgE were recorded. In addition, a comparable study was conducted in rats infected with Nb. At 11 days after infection (DAI) of mice with Nb or Ts, or rats with Nb, there was a 3-fold increase in cell number in the MLN. There was a marked increase in cell number in the MLN of mice infected with Sm at 7 weeks after infection (WAI) and in the spleens of Sm-infected mice at 4 WAI. The percentage of cIgE + cells increased from undectectable levels in uninfected mice and rats to as high as 0.5–1.3% in the MLN of helminth-infected mice and rats. Analysis of cell surface molecules with a fluorescence activated cell sorter (FACS) showed that Nb and Ts infection induced slight increases in the percentages of B cells and slight decreases in the percentage of T cells. More remarkably, the percentage of sIgE + cells in the MLN of both Nb- and Ts-infected mice rose from undetectable levels in uninfected mice to 33 and 27%, respectively, at 15 DAI. This rise was stimulated in Ts-infected mice predominantly by adult Ts. In the MLN of Nb-infected rats, the percentage of cells that were sIgE + was greater than 50% at 15 DAI. However, there was no detectable increase in sIgE + cells in the spleen and MLN of Sm-infected mice until 5 WAI; peak levels of approximately 20% sIgE + cells were reached at 8 WAI. Treatment of MLN cells, from mice infected with Nb, Ts or Sm and rats infected with Nb, with pH 4.0 acetate buffer for 1 min (acid treatment) removed all detectable sIgE from greater than 90% of the sIgE + cells, but did not remove sIgD or sIgM from cells with these surface isotypes. The effect of acid treatment on sIgE was similar even after a secondary infection of mice or rats with nematode larvae. These data show that helminthic infection, in general, is a potent stimulator of the IgE system at the cellular level and that almost all of the sIgE + cells that arise have acquired cytophilic sIgE.

Allogeneic reactivity of maternal lymphoid cells during the course of gestation. Modifications and sex differences in a local GVH assay

The immune reactivity of lymphoid cells from pregnant mice was studied during the course of pregnancy in primiparous and multiparous animals either “isopregnant” (male and female of same strain) or “allopregnant” (male and female differing at H-2), using a local GVH assay (CBA lymphoid cells injected into (CBA × A/J)F1 recipients). The findings were as follows: (1) The lymphoid cell number in the para-aortic lymph nodes (PALN) was increased at all stages of gestation. The peak occurred in the 2nd week in primiparity and as early as 60 h after fertilization in multiparity. (2) PALN cell alloreactivity was weak at the beginning and higher than normal in the third week of pregnancy. (3) Spleen cell alloreactivity was increased in the second week and decreased in the third week in primiparous compared with multiparous animals. (4). Anti-paternal alloreactivity exhibited by spleen cells of allogestation was decreased (as compared to cells of isogestation) especially in primiparous mice, particularly in the third week. At this time, the anti-paternal alloreactivity of PALN cells was increased. (5) The influence of the recipient's sex on GVHR intensity was reversed when the cells were obtained from a pregnant donor, becoming stronger in male compared with female hybrids.

A modified ERG technique and the results obtained in X-linked retinitis pigmentosa.

An electroretinographic (ERG) technique is described in which the relationship between scotopic b wave amplitude and stimulus light intensity is determined. The relative amplitude of scotopic to photopic responses is assessed by means of red light and flicker. The method is applied to the detection of ERG abnormalities in heterozygotes for X-linked retinitis pigmentosa. These have been found in only a proportion of cases. The ERG results can be used to suggest the nature of the retinal abnormality.

Experimental allergic encephalomyelitis: Simple method for producing the localized form

In the past, the lesions of experimental allergic encephalomyelitis (EAE) have been induced to localize around brain tissue damaged by anoxia or direct physical or chemical attack. The procedure for producing the requisite antecedent brain injury has been simplified by use of a single subcutaneous injection of a neurotoxic chemical, thereby eliminating the need for surgery. The EAE lesions are concentrated in a very small area and can be obtained in one day after the passive transfer of a relatively small number of lymph node cells from immunized donors.

Role of neutralizing antibodies and T-cells in pathogenesis of herpes simplex virus infection in congenitally athymic mice

Congenitally athymic nude mice were infected with 10 4 p.f.u. herpes simplex type I (strain SC16). Following the passive transfer of neutralizing monoclonal antibodies (AP 7, AP 8 and AP 12) it was observed that AP 7 alone reduced the virus infectivity in the nervous system; AP 8 and AP 12 failed to protect mice probably due to poor in vivo binding to the neutralization site on the virus. Latent ganglionic infection could be established in nude mice following adoptive transfer of optimum number (2 × 10 7 cells/mouse) of immune lymph node cells from day 7 herpes virus-infected hairy immunocompetent donor mice. Moreover, in some of the immune lymph node cell protected nudes, latency could be maintained even in complete absence of neutralizing antibodies. Results of ear-ablation experiments revealed that removal of primary source of infection after day 5 of infection reduced the amount of virus in the ganglia and spinal cord. Acute neurological infection was not detected following transfer of protective anti-gp-D neutralizing antibody (LP 2) in combination with removal of infected pinna. These data suggest that continuous seeding of virus occurs in related ganglia via the axonal route from infected ear pinna. It appears that local T-cell-mediated immune mechanisms are involved in maintenance of latency.

Antigen presentation by peritoneal macrophages from young adult and old mice

Macrophages perform vital inductive and regulatory functions in immune processes and host defense mechanisms. However, macrophage function during senescence has not been extensively studied. Although antibody response is dramatically reduced in old animals, antigen presentation has never been directly assessed. Therefore, the antigen-presenting capabilities of purified peritoneal macrophages from young adult and old mice were studied by quantitatively measuring their ability to induce antigen specific proliferation of lymph node T lymphocytes. Increasing numbers (10 2 to 10 5) of macrophages from nonimmunized young adult (3 to 6 months) or aged (27 to 36 months) animals were cultured in the presence of antigen with a constant number (2 × 10 5) of column-separated popliteal lymph node cells from young adult mice. The latter had been immunized with the dinitrophenyl conjugate of bovine γ-globulin in complete Freund's adjuvant by footpad injection. Macrophages from old animals were equal to macrophages from young adult animals in stimulating T-lymphocyte proliferation, and the kinetics of incorporation was identical with increasing numbers of macrophages from either young adult or old animals. However, greater numbers of resident or induced peritoneal macrophages were always harvested from old animals. Differences in macrophage activity as assessed by different functional parameters may be reconciled by implicating subpopulations of macrophages that perform separate functions e.g., Ia-positive antigen presenter and Ia-negative scavenger macrophages.

Adoptive transfer of experimental autoimmune thyroiditis (EAT) with in vitro activated lymph node cells from thyroglobulin-sensitized guinea pigs: characterization of the cell that transfers EAT.

Relatively low numbers of lymph node cells (LNC) from Strain 2 or Strain 13 guinea pigs sensitized with guinea pig thyroglobulin (GPTG) could transfer experimental autoimmune thyroiditis (EAT) to normal syngeneic recipients after in vitro culture with GPTG. The EAT induced in recipients of cultured LNC was similar in incidence and severity to EAT induced by active immunization with GPTG in complete Freund's adjuvant. The cells that were effective in transferring EAT were shown to be immunoglobulin-negative, nylon wool nonadherent, and Ia-negative. The effector cells were sensitive to irradiation after in vitro activation, indicating that cell proliferation in the recipient is required for development of EAT. Recipient animals often developed moderate to severe lesions of EAT yet none had detectable delayed hypersensitivity to GPTG and the majority also had no detectable anti-GPTG antibody.

Immunosuppression in a murine B cell leukemia (BCL1): role of an adherent cell in the suppression of primary in vitro antibody responses.

The ability of lymph node cells from mice bearing the BCL1 tumor to respond in vitro to mitogens, allogeneic cells, and both TI and TD antigens was investigated. The lymph nodes of such mice are not invaded with tumor cells and contain normal numbers of T and B cells. Nevertheless, at the peak of tumor burden in the spleen and blood (approximately 8 to 12 wk after injection with tumor cells), the lymph node cells from the tumor-bearing mice display markedly decreased responsiveness both to allogeneic cells and to antigens. In addition, small numbers of lymph node cells from the tumor-bearing mice suppress primary antibody responses of normal lymph node cells. This nonspecific suppression of antibody responses is mediated by a G-10 Sephadex adherent, non-T, non-B cell present in the nodes of the tumor-bearing mice. Since the BCL1 tumor model is in many respects similar to the prolymphocytic type of human chronic lymphocytic leukemia, the present results may be helpful in elucidating the mechanisms underlying the in vivo immunosuppression associated with lymphocytic neoplasms in humans.

Autoimmune effector cells. I. Transfer of experimental encephalomyelitis with lymphoid cells cultured with antigen.

Relatively low numbers of spleen and lymph node cells from Lewis rats previously challenged with myelin basic protein efficiently transfer experimental allergic encephalomyelitis (EAE) to normal syngeneic recipients after in vitro culture with antigen. Moreover, cells obtained from rats that have recovered and are resistant to EAE can also transfer disease. Cell separation studies show that a nylon wool-adherent cell is responsible for transfer of EAE. Density gradient ultracentrifugation revealed that these effector cells are probably lymphoblasts.

T-T Cell Synergy in the in Vitro Generation of Secondary Cell-Mediated Cytotoxicity Against Syngeneic SV-40 Transformed Cells

Abstract The concept of T-T cell interaction that was first suggested during cell-mediated immune response to alloantigens was evaluated in vitro in a syngeneic tumor system. Combinations of lymph node and thymus cells from mice immune against SV-40 transformed cells were shown to exhibit synergism with respect to in vitro generation of secondary cytotoxic effector cells against the corresponding target cells. That is, the activity exhibited by combinations of lymph node and thymus cells was greater than that given by equal numbers of the two cell types cultured separately. With respect to the kinetics and magnitude of effector cell generation, the response occurring in synergizing cultures closely resembled that of optimal numbers of lymph node cells. In addition, the antigen specificity of effector cells generated by synergizing cultures was similar to that of effectors derived from cultures containing optimal numbers of responding lymph node cells. Pretreatment of either lymph node or thymus cells with mitomycin C or x-ray irradiation completely abolished their synergistic interaction. The lymph node and thymus cell populations participating in synergy were found to be thymus dependent. since we have previously shown that similar T-T cell interaction occurs in rejection of a syngeneic SV-40 induced sarcoma in mice, it is proposed that the in vitro generation of secondary cytotoxicity provides an in vitro model for the study of in vivo thymus cell interactions in response to tumor-associated antigens. These results suggest that we may be dealing with the same or similar T cell populations described before as displaying synergy in response to alloantigens in the graft-vs-host, mixed lymphocyte, and cell-mediated cytotoxicity reactions.

Regulation of B-lymphocyte clonal proliferation by stimulatory and inhibitory macrophage-derived factors

A functional subpopulation of murine B lymphocytes proliferate in semisolid agar culture in the presence of 2-mercaptoethanol to form colonies. The effects of diffusible macrophage-derived factors on this focal proliferation was investigated using a two-layer culture system which prevented macrophage-lymphocyte contact and permitted B-cell activation to be critically assessed under conditions of extremely low cell densities. Adherent peritoneal macrophages incorporated within underlayers of spleen or lymph node cell cultures potentiated both the number and size of developing B-cell colonies. These effects were most striking when low numbers of spleen or lymph node cells, or macrophage- depleted lymphoid cell suspensions were used. Thus, macrophage-depleted lymph node ceils gave rise to virtually no colonies, but colony-forming ability was restored by the presence of an optimal number of macrophages. When the number of macrophages exceeded that required for optimal stimulation, colony formation was suppressed; an effect which was largely prevented by indomethacin, an inhibitor of prostaglandin synthesis. Under these conditions, stimulation and inhibition of B-cell activation by macrophages could be dissociated, indicating that each signal is selectively controlled by individual molecules elaborated by the macrophage. With an appropriate number of macrophages required for B-cell activation, and sufficient indomethacin to inhibit the accumulation of macrophage-derived prostaglandin, B-lymphocyte clonal proliferation was a linear function of the number of B cells placed in culture. In the absence of macrophages, B-cell colony formation was potentiated by both lipopolysaccharide and intact sheep erythrocytes through a mechanism different from that of the macrophage-derived stimulatory factor. In addition to their direct stimulatory effect on B-cell proliferation, lipopolysaccharide and sheep erythrocytes were each capable of modulating the production and/or release of B-cell stimulatory and inhibitory factors by the macrophage. Parallel studies of conventional mitogen- stimulated lymphocyte cultures did not show a requirement for macrophages and confirm that the semisolid assay is uniquely suited to studies on the regulatory role of the macrophage in B-cell activation.