Plaque-forming Cell Assay Research Articles

Immunomodulating activities of water extract fromXanthium strumarium (II): Immunostimulating effects of the water layer after treated with chloroform

One of water and/or methanol extracts from 14 herbal drugs which were screened using murine splenocytes showed immunosuppressive activities previously. After water extract fromXanthium strumarium was treated with chloroform, 100μg/ml of water layer (XS-WC1) has very strong immunostimulating activities tested by3H-thymidine incorporation (control vs 100μg/ml, 4962 cpm vs 69515 cpm). MLR also appears to be stimulated strongly (control vs 100μg/ml, 26345 cpm vs 78688 cpm). When 100μg/ml of XS-WC1 and 0.8μg/ml of concanavalin A (ConA) were added, more3H-thymidine were incorporated significantly, compared with 0.8μg/ml of ConA only. In contrast with ConA, results from 5μg/ml of lipopolysaccharide (LPS) and 100μg/ml of XS-WC1 were not different, compared with 5μg/ml of LPS only. These results indicated that responses of XS-WC1 to B cell and T cell may be different. XS-WC1 was injected intraperitoneally (10mg/kg, 50mg/kg, 100mg/kg) for 4 days or 10 days and tested secretion of IgM or IgG by direct and indirect hemolytic plaque-forming cell assays, respectively. Numbers of hemolytic plaques for both IgM and IgG were increased significantly. Especially, secretion of IgGs was increased more than 10 times. After administration of XS-WC1 for 7 days (50mg/kg, 100mg/kg) splenomegaly due to graft vs host reaction was observed. Human lymphocytes separated from whole blood by Ficoll-Hypaque method were also proliferated after treatment of 10μg/ml and 50μg/ml of XS-WC1. As seen in murine lymphocytes, human lymphocyte proliferation was increased synergistically after treatment with both of XS-WC1 and phytohemagglutinin (PHA). It appears that XS-WC1 may have potential immunostimulating activities and that it remains to be purified further for isolation of active components.

Immunotoxic1ty of bis(tri‐n‐butyltin) oxide in the rat

Previous studies in the rat have shown that bis(tri-n-butyltin) oxide (TBTO), used as a biocide, was immunotoxic at dose levels that did not affect other organs. In order to determine a no-effect level, weanling rats were treated for at least 28 consecutive days with TBTO at 0, 0.5, 2, 5, or 50 mg/kg of diet. Studies on clinical chemistry, hematology, pathology, and immune function, that is, plaque-forming cell (PFC) assay, delayed-type hypersensitivity (DTH) response, and the splenic clearance of Listeria monocytogenes, were performed at the end of treatment. No treatment-related effects were noted on clinical chemistry and hematology parameters and on PFC and DTH response, whereas thymic atrophy and impaired clearance of L. monocytogenes were noted only at a dietary concentration of 50 mg/kg. These results confirm the thymus as a target organ of TBTO immunotoxicity. Under the conditions of these experiments the dietary concentration of 5 mg/kg, equivalent to a dose of 0.5 mg/kg body weight, represents a no observed effect level (NOEL) for immunotoxicity in the Sprague-Dawley rat.

Antibody responses of mice exposed to low‐power microwaves under combined, pulse‐and‐amplitude modulation

Irradiation by pulsed microwaves (9.4 GHz, 1 microsecond pulses at 1,000/s), both with and without concurrent amplitude modulation (AM) by a sinusoid at discrete frequencies between 14 and 41 MHz, was assessed for effects on the immune system of Balb/C mice. The mice were immunized either by sheep red blood cells (SRBC) or by glutaric-anhydride conjugated bovine serum albumin (GA-BSA), then exposed to the microwaves at a low rms power density (30 microW/cm2; whole-body-averaged SAR approximately 0.015 W/kg). Sham exposure or microwave irradiation took place during each of five contiguous days, 10 h/day. The antibody response was evaluated by the plaque-forming cell assay (SRBC experiment) or by the titration of IgM and IgG antibodies (GA-BSA experiment). In the absence of AM, the pulsed field did not greatly alter immune responsiveness. In contrast, exposure to the field under the combined-modulation condition resulted in significant, AM-frequency-dependent augmentation or weakening of immune responses.

Primary immune responses of selected small mammal species to heterologous erythrocytes

1. 1. We surveyed the primary humoral immune responsiveness of six small mammal species ( Peromyscus leucopus, Microtus pinetorum, Perognathus hispidus, Neotoma floridana, Onychomys leucogaster, Mus musculus) collected from wild populations in central Oklahoma using sheep red blood cells (SRBC) as the immunogen and a splenic plaque-forming cell (PFC) assay. 2. 2. Individuals within each wild species examined produced antibodies to a single intraperitoneal injection of SRBC, however, considerable interspecific and intraspecific variation in responsiveness was indicated. 3. 3. Overall, primary immune responsiveness varied from 0 to 5013 PFC/10 6 cells. Spleen weights, total splenic nucleated cell yields, PFC/spleen, PFC/mg spleen, and PFC/10 6 cells were significantly different among species. 4. 4. Mean cell yield in 0M. pinetorum was greater than in P. leucopus and CD-1 laboratory mice (included as positive controls). Number of PFC/10 60 cells was greater in CD-1 laboratory mice than P. hispidus and P. leucopus. The coefficient of variation for PFC/10 6 cells in CD-1 laboratory mice was 38% compared to 109, 129, and 56% for M. pinetorum, P. hispidus, and P. leucopus, respectively. 5. 5. Interspecific and intraspecific differences among wild species may be a reflection of disparate life histories and other environmental selection pressures.

Enhancement of Humoral Immunity by Heterologous Lipid Peroxidation Products Resulting from Burn Injury

Lipid peroxidation products (conjugated dienes) extracted from the plasma of scald-burned rats were injected intraperitoneally in mice before immunization of the mice with sheep erythrocytes. Five-day plaque-forming cell assays showed that mice receiving conjugated dienes isolated from the plasma of burned rats had enhanced specific immunoglobulin M antibody production compared to mice receiving dienes from normal rat plasma or sheep erythrocytes alone. These observations suggest that lipid peroxidation products in the plasma of burned animals may modulate the humoral immune response.

Alteration of Membrane Oligosaccharides by Castanospermine, an Alpha Glucosidase Inhibitor, Enhances Immunoglobulin Production in Staphylococcus aureus Cowan I‐Stimulated Lymphocyte Culture

Castanospermine (CSP) inhibits alpha-glucosidase, which is involved in the initial step of N-linked oligosaccharide processing of secretory and membrane glycoproteins. In Staphylococcus aureus Cowan I (SAC)-stimulated human lymphocyte culture, CSP at a dose of 20 micrograms/ml caused a twofold increase in immunoglobulin G (IgG) release after 7 days. An initial 48-h exposure to CSP sufficed for this enhancing effect. Plaque-forming cell assays on the seventh day disclosed that CSP caused an increase in the number of IgG-, IgA- and IgM-secreting cells. In cross-culture experiments, only a mixture of B cells pretreated with CSP and untreated T cells showed an increase in IgG production. Tritiated thymidine incorporation studies revealed that CSP enhanced B-cell responses to T cell-derived soluble factor (TSF). When incubated with CSP for 18 h, B cells showed an increased surface binding on [3H]concanavalin A (Con A). These results indicate that the alteration in B-cell membrane oligosaccharides enhances the response to TSF at an early stage of SAC culture, leading to an increase in Ig-secreting cell number at later stages. The present study provides evidence that cell-surface oligosaccharides of B cells play an important role in the responses of B cells to lymphokines.

Depression of prothymosin-α production in murine thymus correlates with staphylococcal enterotoxin-B-induced immunosuppression

Prothymosin-α (ProTα) and thymosin-β4 (Tβ4) were isolated from murine thymus and characterized by microsequence analysis. Murine Tβ4 has an identical sequence to bovine Tβ4, whereas murine ProTα is highly homologous to rat ProTα. Murine ProTα differs from rat ProTα at two positions, Glu100 and Asp108 of the rat sequence are substituted by aspartic and glutamic acid, respectively, in murine ProTα. The amount of ProTα in murine thymus was found to be reduced after in vivo treatment with staphylococcal enterotoxin-B (SEB), a superantigen which stimulates T cells bearing specific Vβ receptors. Results from the anti-SRBC (sheep erythrocyte) plaque-forming cell assay showed that the antibody response of the spleen cells from these animals was also suppressed. On the other hand, the amount of Tβ4 was not changed significantly. Our studies suggest that the suppression of SEB on antibody response correlates with the depression of ProTα production in the thymus.

Self-stimulation behavior: Consequences upon immunity?

A variety of behaviorial and emotional factors can affect the immune response by changing the brain immunoregulatory mechanisms, resulting in immunosuppression or immunopotentiation. This experiment deals with the effect of chronic self-stimulation behavior on the immune response and lymphoid tissue. Male rats were stereotaxically implanted with bipolar electrodes into the lateral hypothalamus and 7 days after surgery were screened for self-stimulation behavior. Lateral hypothalamus self-stimulating rats (LH-SS) were allowed to self-stimulate for 60 min/day for a period of 9 consecutive days: 5 days before and 4 days after immunization with 5 × 10 9 sheep red blood cells (SRBC). The animals were sacrificed and plaque-forming cell assay (PFC), microhaemmagglutination reaction with SRBC, and differential blood leucocyte counts were performed. The thymus, spleen, and inguinal lymph nodes were weighed and processed for histological examination. In the LH-SS group, an enhanced PFC response and increased anti-SRBC antibody titer were observed when compared to controls. The thymus and spleen of LH-SS rats were smaller in size in comparison with controls, but only with moderate changes in splenic cellular make-up. The relative number of lymphocytes was increased in peripheral blood of LH-SS rats when compared to intact animals. The results obtained suggest that chronic self-stimulation behavior can modulate some parameters of humoral immune response and affect the relative weight of lymphoid organs in the rat.

Dual effects of interleukin 4 on antigen‐activated human B cells: induction of proliferation and inhibition of interleukin 2‐dependent differentiation

We analyzed the effects of interleukin 2 (IL 2) and IL 4 isolated and in association on the specific response of human B cells triggered by trinitrophenylated polyacrylamide beads (TNP-PAA). IL 2 induced an increase (more than 10 times) in the number of hapten-binding cells [detected by a rosette-forming cell (RFC) assay] as well as the generation of antibody-producing cells [detected by a plaque-forming cell (PFC) assay]. IL 4 induced an isolated RFC response without PFC response. We verified that the IL 4 (as well as 12)-induced RFC were hapten specific and mediated through membrane IgM. Density fractionation experiments showed that IL 4-induced RFC were equally distributed between high-density and intermediate-density B cells. IL 2 appeared to drive more B cells into the intermediate density fraction. IL 2-induced PFC belonged to the RFC population and were intermediate-size B cells. IL 2 drove more RFC into an activated stage and it induced the differentiation of a number of them into antibody-producing cells. The evaluation of the proportion of RFC able to incorporate thymidine showed that both IL induced a substantial proliferation of antigen-activated B cells. However, IL 4 inhibited the IL 2-dependent PFC without affecting the number of RFC nor the proportion of proliferating RFC induced by this IL. These results directly demonstrate that human IL 4 triggers the expansion of antigen-activated B cells and selectively inhibits the IL 2-induced differentiation.

Immunotoxicity of the pyrrolizidine alkaloid monocrotaline following subchronic administration to [formula omitted] mice

Monocrotaline (MCT) is a member of a class of compounds known as pyrrolizidine alkaloids (PAs). PAs are found in the leaves and seeds of a variety of plant species. The potential intoxication of livestock and man through the ingestion of contaminated grains and other foods makes PAs a significant toxicological concern. MCT exposure in rats results in lesions to hepatic and cardiopulmonary tissues as well as alterations in lymphoid organ cellularity. However, no previous studies have investigated MCT-induced functional alterations in the immune system. In the present study, MCT was administered by gavage for 14 days at 0, 10, 25, 50, 75, and 150 mg/kg to female C57B1 6 mice. The antibody-mediated immune response to sheep red blood cells (SRBC) was assessed using the plaque-forming cell (PFC) assay and the hemolytic antibody isotope release assay (HAIR). Additionally, the cytotoxic T-lymphocyte (CTL) response to allogeneic P815 tumor cells was determined after MCT exposure. Although hepatic and pulmonary lesions are common sequelae to MCT exposure in rats, the C57B1 6 mouse appeared to be more resistant to these effects on the basis of histopathological examination. Furthermore, the overt toxicity of MCT was minimal with respect to organ weight changes in liver, kidney, and lung. In contrast, a dose-dependent suppression in the antibody response to SRBC was observed with a minimum significant dose of 25 mg/kg. At this dose the number of anti-SRBC PFC per 10 6 spleen cells and the amount of anti-SRBC antibody measured in the HAIR assay were 57 and 59% of control, respectively. The CTL response was significantly decreased (38% of control, 30:1 E:T), and the total number of CTL lytic units per spleen was 12% of control at 75 mg/kg. Antibody titers to P815 were also dose-dependently suppressed. Therefore, we conclude that the immune system in C57B1 6 mice is a sensitive target of MCT toxicity.

Quantitation of Antibody‐Secreting Cells in the Blood after Vaccination with aHaemophilus influenzaeType b Conjugate Vaccine

The human B-lymphocyte response to protein-conjugated polysaccharide antigens has not previously been studied at the cellular level. In order to do so, we developed and evaluated haemolytic plaque-forming cell assays detecting Haemophilus influenzae type b (Hib) capsular polysaccharide-specific antibody-secreting cells (AbSC) of the isotypes IgM, IgG, and IgA. The appearance of AbSC in the blood after vaccination of adults with diphtheria toxoid-conjugated Hib polysaccharide was investigated. AbSC were detected from post-vaccination day 5 to day 14. IgA was the predominant isotype among these cells. IgM AbSC peaked slightly earlier (median day 7) than IgG and IgA AbSC (both day 8). On post-vaccination day 8 the numbers of AbSC were: IgA, 1217/10(6) mononuclear cells (median); IgG, 211; and IgM, 30 (n = 11). Similar isotype distribution has earlier been found after vaccination with pure capsular polysaccharides from Hib and pneumococci. The predominance of IgA AbSC in response to both conjugate and pure polysaccharide vaccines is probably due to reactivation of the same clones of IgA-committed memory B cells originally primed at the mucosa by natural exposure to the polysaccharide or cross-reacting antigens.

Type 6 and 19 pneumococcal polysaccharides coupled to erythrocytes elicit pneumococcal cell wall‐specific primary IgM responses and capsular polysaccharide‐specific secondary IgG responses

Previous results have shown that the primary murine antibody responses to vaccine preparations of type 6 (S6; Danish type 6A) or type 19 (S19; Danish type 19F) pneumococcal capsular polysaccharides consist entirely of IgM antipneumococcal cell wall carbohydrate (PnC)-specific antibodies. No capsular polysaccharide-specific IgM antibodies were detectable by plaque-forming cell or enzyme-linked immunosorbent assay techniques. In this report, antibodies specific for S6 and S19 capsular polysaccharides were induced in secondary responses to chicken erythrocyte (CRBC) conjugates of S6 and S19. Essentially all detectable IgG produced in the secondary response was capsular polysaccharide specific and included all subclasses of IgG. In contrast, all detectable IgM produced in the primary response to S6-CRBC and S19-CRBC, and the IgM produced in the secondary response to S6-CRBC was not capsular polysaccharide specific since it reacted with PnC. Thus, there is a major change in the specificity of the primary IgM response compared to the secondary IgG response of mice immunized with S6-CRBC or S19-CRBC. Injection of PnC or any PnC-containing polysaccharide prior to immunization with S6-CRBC or S19-CRBC resulted in suppression of the primary IgM response. In contrast, only the capsular polysaccharide used in the immunizing polysaccharide-erythrocyte conjugate suppressed induction of the capsular polysaccharide-specific secondary IgG response. These results suggest that S6 and S19 capsular polysaccharide-specific IgG-producing memory B cells derive from capsular polysaccharide-specific precursors which do not produce detectable antibody after primary immunization.

The use of monoclonal antibodies in an enzyme immunospot assay to detect isotype-specific antibody-secreting cells in pigs and chickens

Monoclonal antibodies directed against porcine immunoglobulin isotypes G, G1, G2, M, and A and against chicken immunoglobulin isotopes G, M, and A were tested in an antigen-specific spotforming cell (SFC) assay based on the principle of the enzyme immunoassay. The SFC assay was used to quantitate ovalbumin (OA)-specific antibody-secreting cells (ASC) in pigs that had been primed and boosted with OA. The SFC assay was also used to quantitate trinitrophenyl (TNP)-specific ASC in chickens that had been primed with TNP-conjugated keyhole lympet haemocyanin (TNP-KLH). Although, the classical plaque-forming cell (PFC) assay cannot reliably detect isotope-specific ASC in pigs and chickens, it can detect these cells in mice. Therefore, we compared the OA- and TNP-specific SFC assays with PFC assays that were specific for these antigens in mice. The study demonstrated that the SFC assay is superior to the PFC assay in detecting both OA-specific ASC and TNP-specific ASC. The frequencies of OA-specific and TNP-specific SFC detected in mice were of the same order of magnitude as those detected in pigs and chickens. We concluded that the SFC assay is the better method for quantitating ASC in pigs, chickens, and probably all domestic animals for which isotype-specific monoclonal antibodies are available.

Effects of capsaicin treatment on immunoglobulin secretion in the rat: further evidence for involvement of tachykinin-containing afferent nerves

Neonatal capsaicin treatment has previously been shown to diminish the primary antibody response of adult rats to the subcutaneously administered T-dependent antigen, sheep red blood cells, as measured using a modification of the Cunningham plaque-forming cell assay technique. We have now studied the kinetics of this response in adult normal, neonatally capsaicin-pretreated and neonatally capsaicin-pretreated substance P-infused rats, and examined the effects of the tachykinin antagonist Spantide, on the plaque-forming cell response. Capsaicin pretreatment did not affect the antigen-specific plaque-forming cell response over the first 4 days following antigen injection. At days 5, 6 and 7 of the response, there was a statistically significant decrease in the number of plaque-forming cells secreting antigen-specific IgM, an effect not observed in capsaicin-pretreated rats which were given a subcutaneous infusion of substance P at the time of antigen injection. The tachykinin antagonist Spantide inhibited the plaque-forming cell response in normal rats after in vivo infusion at the time of antigen injection by more than 70%. This effect of Spandite was dose dependent, occured with maximal effect at 10 μM, and appeared to be independent of any histamine-mediated action. The results of this study provide further evidence for a receptor-mediated immunomodulatory role of tachykinin-containing primary afferent nerves.

Lack of high avidity IgM anti-ssDNA antibodies in cells from autoimmune-prone and autoimmune mice

Non-autoimmune prone CBA mice were compared with autoimmune prone NZB, NZW, and (NZB x NZW)F1 mice for the ability of their splenic cells to produce anti-ssDNA-forming cells spontaneously in vitro, measured in the plaque forming cell assay. The number of antibody forming cells was measured and the relative avidity of antibody produced determined using a plaque inhibition assay. Splenic lymphocytes from young animals of a non-autoimmune strain (CBA/J) were shown to be capable of generating anti-ssDNA IgM antibody-forming cells in culture which displayed a higher avidity for antigen than that from autoimmune-prone or frankly autoimmune mice. Since an increased switching from IgM to IgG autoantibody production and defects in Fc-mediated signalling by IgG antibody have been identified in autoimmunity, we suggest that the metabolic block, normally in force in non-autoimmune-prone animals, accounts for this elevated avidity of IgM autoantibody.

Host Response to Myeloma: Effect of Syngeneic Spleen Cells on the Growth and Functionof MOPC 104E Myeloma in Vitro

The effect of coculturing nonadherent and plastic adherent cells from the spleen with MOPC 104E KI81 for short (24 h) and long term (7 days) was investigated. In both culture systems, the effect of the spleen cells on the secretion of IgM by plaque-forming cells assay and growth of the plasmacytoma by cell counts and flow cytometry was measured. In these studies, a low effector:target (E:T) ratio which did not produce cytotoxicity to MOPC 104E cells was used. We observed that while nonadherent spleen cells from normal or MOPC 104E-primed mice inhibited secretion of IgM by the MOPC 104E cells, they stimulated the proliferation of MOPC 104E cells two times faster than MOPC 104E cells cultured alone. Plastic adherent cells from the spleens of normal mice or MOPC 104E-primed mice also inhibited secretion of IgM by the tumor cells as measured by plaque formation, and stimulated proliferation of MOPC 104E in 24 h coculture. Plastic adherent cells from normal nonprimed mice initially stimulated the myeloma to grow, but by 24 h, a large fraction of the population was in the G1 or possibly resting state. The effect of nonadherent and plastic adherent cells on the stem cell activity of MOPC 104E was also tested in 7-day colony-forming assays. Nonadherent cells had no effect on colony-forming units or plaque formation. Plastic adherent cells from normal spleen cells inhibited plaque formation by 68% but had no effect on colony formation. However, plastic adherent cells from spleens of mice primed in vivo with MOPC 104E tumor cells suppressed plaque formation by 98% and also reduced colony formation. The results showed that inhibition by macrophages of IgM production by MOPC 104E cells is independent of cell proliferation. The adherent macrophages from both normal and in vivo-primed spleen cells were Mac.1 positive after 7 days of coculture with MOPC 104E cells. However, the density of Mac.1 was greater on primed macrophages.

Immunotoxicity of the Pyrrolizidine Alkaloid Monocrotaline following Subchronic Administration to C57BI/6 Mice

Immunotoxicity of the Pyrrolizidine Alkaloid Monocrotaline following Subchronic Administration to C57B1/6 Mice. DEYO, J. A., AND KHRKVUET, N. I. (1990). Fundam. Appl. Toxicol14, 842–849. Monocrotaline (MCT) is a member of a class of compounds known as pyrrolizidine alkaloids (PAs). PAs are found in the leaves and seeds of a variety of plant species. The potential intoxication of livestock and man through the ingestion of contaminated grains and other foods makes PAs a significant toxicological concern. MCT exposure in rats results in lesions to hepatic and cardiopulmonary tissues as well as alterations in lymphoid organ cellularity. However, no previous studies have investigated MCT-induced functional alterations in the immune system. In the present study, MCT was administered by gavage for 14 days at 0, 10, 25, 50, 75, and 150 mg/kg to female C57B1/6 mice. The antibody-mediated immune response to sheep red blood cells (SRBC) was assessed using the plaque-forming cell (PFC) assay and the hemolytic antibody isotope release assay (HAIR). Additionally, the cytotoxic T-lymphocyte (CTL) response to allo-geneic P815 tumor cells was determined after MCT exposure. Although hepatic and pulmonary lesions are common sequelae to MCT exposure in rats, the C57B1/6 mouse appeared to be more resistant to these effects on the basis of histopathological examination. Furthermore, the overt toxicity of MCT was minimal with respect to organ weight changes in liver, kidney, and lung. In contrast, a dose-dependent suppression in the antibody response to SRBC was observed with a minimum significant dose of 25 mg/kg. At this dose the number of anti-SRBC PFC per 106 spleen cells and the amount of anti-SRBC antibody measured in the HAIR assay were 57 and 59% of control, respectively. The CTL response was significantly decreased (38% of control, 30: 1 E:T), and the total number of CTL lytic units per spleen was 12% of control at 75 mg/kg. Antibody titers to P815 were also dose-dependently suppressed. Therefore, we conclude that the immune system inC57Bl/6 mice is a sensitive target of MCT toxicity.

Immunological Consequences of Lesions of Nucleus Basalis in Rats

In this study we followed up the effects of acute (ten days) and chronic (two months) stereotaxic lesions of NB in rats on: 1.) cellularity of lymphatic organs, 2.) humoral and 3.) cellular immunity and 4.) in vitro functional reactivity of splenocytes to polyclonal mitogens. Experiments were done on (AoxDA) F1 male rats, aged 4 months and maintained in standard laboratory conditions. The coordinates for bilateral stereotaxic lesions were taken from atlas of Pellegrino. Electrolysis was performed unipolarly at the anode with 10 mA for 10s. Histological verification of the lesions was performed by the method of Wolf. For the evaluation of humoral or cell mediated immunity standard plaque forming cell assay to sheep red blood cells (SRBC) or transplantation of allogeneic skin were used. The data have shown that bilateral electrolytic lesions of NB cause the prolongation of rejection time to allogeneic skin, transplanted on tenth p.o. (15.8+-0.4 days versus 12.8+-0.5 in intact control, p<.001). Some prolongation of allograft survival was, however, seen also in group of rats in which electrodes were lowered in NB without turning off the electric current, indicating the consequences of mechanical lesions of NB on cell mediated immunity. Humoral immunity tested after sensitization of rats with SRBC, on tenth p.o. was found also to be slightly depressed and followed by depletion of thymus, but the values were on the border of statistical significance. Chronic electrolytic lesions of NB in rats which were not otherwise sensitized, revealed that in comparison to appropriate controls, the cellularity of thymus of NB lesioned rats is higher, while the cellularity of spleen is decreased. Although the tendency of hypoplasia of bone marrow seems to exist, the results are not statistically significant. The data imply that NB is not important only for several behavioural functions but also for mechanisms involved in the maintenance of normal immunological response and composition of lymphoid organs. It is obvious that NB might play an important role in the reestablishment of homeostasis after the initiation of an immune response.

Immunological functions in food-restricted rats: enhanced expression of high-affinity interleukin-2 receptors on splenic T cells

Several immunological functions of B and T cells including IL-2 receptor expression on T cells were measured in 12-month-old Fisher-344 male rats maintained from 6 weeks of age on an ad libitum (AL) or a 40% food-restricted (FR) diet. Direct anti-SRBC plaque-forming cell (PFC) assays revealed a higher response in FR rats than in AL rats when splenocytes were cultured with or without recombinant interleukin-2 (rIL-2). B cell functions were studied by using nylon wool-purified splenic B cells stimulated either with rIL-2, lipopolysaccharide (LPS), or Salmonella typhimurium mitogen (STM) as a thymus-independent antigen. Reserve plaque assay showed no difference between FR and AL rats in the secretion of anti-IgM and anti-IgG antibodies. In addition, no difference was found in proliferation of B cells stimulated by LPS, STM mitogens or rIL-2. Although purified splenic T cells demonstrated an equally proliferative response in FR and AL rats when cultured with concanavalin A (Con A) or phytohemagglutinin (PHA), T cells in FR rats developed higher responses when stimulated with an alloantigen and rIL-2. Time-course studies carried out to measure high-affinity (HA) IL-2 receptor (R) molecules by using purified T cells with rIL-2 and 125I-labeled IL-2 revealed a higher expression of IL-2R molecules on T cells of FR rats than on T cells of AL rats at 72 h after culturing with Con A. These results indicate that the increased production of antibody to a T cell-dependent antigen (SRBC) and the enhanced responsiveness to alloantigen are attributable to the greater expression of HA IL-2 on T cells, which may facilitate an increased response to and a higher production of IL-2 in FR rats.

Maternal antigenic stimulation actively produces suppressor activity in offspring

Pregnant B10.A(3R) and B10.A(5R) mice were immunized with heterologous erythrocyte and protein antigens and the active immune responsiveness of their offspring was investigated by the plaque-forming cell (PFC) assay. In offspring derived from mothers which were stimulated with optimal amounts of antigens and which had fully developed antibody-forming cells, there was a clear-cut suppression in development of specific PFC over a significant period after delivery. In order to characterize the suppressor cell population, spleen cells were prepared from offspring whose mothers were immunized and thereafter treated by anti I-J k or anti I-J b monoclonal antibody and complement (C′) followed by adoptive transfer to normal corresponding mouse strain. Only the group that received 3R spleen cells treated with anti I-J b monoclonal antibody and C′ had no suppressed PFC. To clarify the suppressor site in offspring, precursor B-cells of experimental offspring responded as hapten specific antibody forming cells by employing homologous hapten and heterologous carrier antigens. These results suggest mechanisms for suppression in offspring whose mothers were stimulated during pregnancy.