Production Of Antibody-forming Cells Research Articles

Effect of Stress on the Production of Antibodies and IL-2, IL-4, and IFNγ Depending on the Time of Antigen Administration and Evaluation of the Role of Opioid Receptors.

The time of stress exposure relative to the moment of immunization affects the direction of the immunoregulatory effect of stress. In case of stress exposure preceding immunization, rotation stress stimulated the production of antibodies, while immobilization depressed it. After antigen injection, these types of stress had no significant effect on the formation of antibody-producing cells. Acute cold stress did not affect the number of antibody-forming cells before immunization, but stimulated the humoral response after it. At the same time, the effect of stress on the production of antibodies was leveled by blockade of opioid receptors with naloxone for rotation and immobilization, but was not canceled for acute cold stress. A similar pattern was revealed when analyzing the effect of stress exposure on cytokine production. Cold stress before antigen administration to mice had almost no effect on the production of IL-2, IL-4, IFNγ, while rotational and immobilization stress naloxone-dependently modulated the synthesis of IL-2 and IL-4. On the contrary, in animals subjected to stress after antigen administration, only cold stress significantly modulated the production of IL-2 and IL-4.

Synthesis of Glycyrrhizic Acid Conjugates with Amino-Acid Methyl Esters and their Ability to Stimulate Antibody Genesis in Mice.

Conjugates of glycyrrhizic acid (GA) with methyl esters of L-amino acids (valine, methionine, and glutamic acid) containing the amino-acid residues in the carbohydrate moiety of the glycoside were synthesized using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide. The resulting GA conjugates at a dose of 2 mg/kg stimulated a primary immune response (production of antibody-forming cells, AFCs) in outbred mice by 1.6 – 3 times as compared with the control. The conjugate of GA with Glu(OMe)2 stimulated antibody genesis in outbred mice 1.7 times more efficiently than N-acetylmuramyl dipeptide and showed a stimulating effect on AFC production in the spleen of CBA mice.

Sensitivity of immunocompetent cells of DBA/2 and C57Bl/6 mice to cyclophosphamide.

T cells were most sensitive to cyclophosphamide in DBA/2 mice, while in C57Bl/6 mice both T and B cells were sensitive. The formation of antibody-producing cells and the production of specific antibodies were delayed in DBA/2 mice immunized after pretreatment with antitumor drug. Accumulation of antibody-producing cells in the spleen was more active in immunized C57Bl/6 mice treated with cyclophosphamide compared to animals not treated with cyclophosphamide.

Immunotoxic effects during acute ethylene glycol poisoning.

Experiments on outbred and inbred male CBA mice showed that acute poisoning with ethylene glycol (0.8 LD50) increased mortality from infections, decreased the number of spleen colony-forming units, inhibited antibody formation (mainly to thymus-dependent antigens, and suppressed natural and antibody-dependent cytotoxicity and delayed-type hypersensitivity. Ethylene glycol in concentrations of 10 and 100 mM in vitro inhibited the formation of antibody-producing cells by acting on T and B lymphocytes.

The effect of chorionic gonadotropin on the functional activity of some populations of splenocytes

Selective immunomodulating effects of high (50 IU/ml) and low (10 IU/ml) doses of chorionic gonadotropin (CG) on the functional activity of A cells and intact splenic T and В lymphocytes were revealed for the syngeneic transfer system. One- hour incubation of nonfractionated splenocytes with CG suppressed the formation of antibody-producing cells (APC) in le- thally irradiated syngeneic recipients. Removal of macrophages from the suspension of intact splenocytes arrested CG-induced immunosuppression and, in case of a low dose, activated the formation of APC. Addition of CG in a low dose to cell culture rich in В lymphocytes activated APC formation. A high dose of the hormone exerted no immunostimulating effect of this kind but selectively suppressed the function of T lymphocytes fractionated by fdtration through nylon fiber filters. Study of the functional activity of A cells by luminol-dependent chemiluminescence showed CG in the studied doses to suppress the generation of active oxygen forms, thus determining the key role of A cells as inducers of CG-dependent immunosuppression in a heterogeneous suspension of splenocytes.

Effect of adamantane amino derivatives on the immune response

In the previous paper [ 1 ] we reported that remantadine and deutiphorin exhibit moderate immunosuppressive activity and are capable of inhibiting the cytotoxicity of natural killer cells and the reaction of lymphocyte blast-transformation in the mononuclear fraction of peripheral blood l~om healthy donors. Earlier, Clark et al. [2] reported similar properties for l-aminoadamantane. In our work [1], it was established that the immunosuppressive activity of the main amino derivatives of adamantane is due to their ability to inhibit activation of Ca2+-phospholipid dependent protein kinase (protein kinase C), and it was suggested that this property may also account for the antiviral activity of these compounds. The ability ofadamantane amino derivatives, as anti-influenza preparations, to inhibit the activation of protein kinase C (thus suppressing the lymphocyte blast-transformation and the functional activity of natural killer cells) suggest that these derivatives may also be capable of affecting the humoral immune response and the post-infection response related to the formation of immune memory cells. Both these processes include the interaction of various T and B cell subclasses, whereby the activated T-lymphocytes belonging to different subclasses may activate or suppress the functional activity of the B cells [3, 4]. As is known, the activation of Tand B-lymphocytes is related to the activation of protein kinase C [5, 6]. In this connection, we have studied the effect of remantadine, 1-aminoadamantane, and deutiphorin on the formation of humoral immunity to bacterial antigen, a living antituberculous BCG vaccine, in CBA line mice. At the same time, we have studied the effect of these compounds on the delayed hypersensitivity that develops simultaneously with the formation of antibodyproducing cells and reflects the T-cell immunity reaction.

Immunotoxicological characteristics of sodium methyldithiocarbamate

This study was conducted to assess immunotoxicological effects and selected general toxicological effects of sodium methyldithiocarbamate (SMD). Initially, the compound was administered orally to female B6C3F1 mice at 300 mg/kg/day for 3, 5, 10, or 14 days. Body, liver, kidney, spleen, and thymus weights were measured. Selected hematological and bone marrow parameters were examined. Flow cytometric analysis was used to assess changes in lymphocyte subpopulations in the thymus and spleen, and production of antibody-forming cells in vitro was measured. Major effects included decreased thymus weight at all time points; increased spleen weight after 10 or 14 days of exposure, increased bone marrow cellularity after 10 or 14 days of exposure, significant decreases in mature lymphocyte subpopulations which were greater in the thymus than in the spleen, relatively selective depletion of the major subpopulation of thymocytes (CD4 +CD8 +), and decreased body weight. Overall patterns of changes were consistent with the conclusion that SMD rapidly depletes most CD4 +CD8 + thymocytes, more slowly depletes a smaller number of mature lymphocytes in the thymus and spleen, and induces compensatory and/or detoxication mechanisms after 10–14 days of exposure. Subsequent experiments were done to assess selected immune function parameters. SMD at 50–300 mg/kg/day for 7 days caused substantial, dose-dependent suppression of NK cell activity. No suppression of antibody production in vivo or splenocyte responses to mitogens or allogeneic lymphocytes in vitro was detected. NK cell activity, thymus weight, and CD4 +CD8 + thymocyte numbers were suppressed by dermal administration of SMD. These results indicate that NK cell activity is significantly decreased by SMD, but mature T and B lymphocytes which survive treatment are generally not significantly impaired with regard to the functions tested. In several experiments, immunological parameters were significantly suppressed in the absence of a significant decrease in body weight, suggesting that most of the effects of SMD on the immune system are not secondary to generalized toxicity.

Comparative immunotoxicity of 2,2′-dichlorodiethyl sulfide and cyclophosphamide: Evaluation of L1210 tumor cell resistance, cell-mediated immunity, and humoral immunity

The immunotoxicity of 2,2′-dichlorodiethyl sulfide (sulfur mustard, SM), on humoral and cell-mediated immunity was compared with that of the nitrogen mustard 2-[bis(2-chloroethyl)amino]tetrahydro-2H-1,3,2-oxazophosphorine 2-oxide (cyclophosphamide, CP). SM and CP had similar effects on thymic and splenic weights, spleen cell number, and the formation of antibody producing cells to sheep red blood cells (sRBC) when examined 5 days after exposure, but differed in their effects on body weights. Although there were no differences in the delayed hypersensitivity response to keyhole limpet hemocyanin, CP and SM had different effects in the L1210 tumor cell allograft rejection assay. CP, but not SM, decreased the 28 day survival rate of allogeneic mice exposed to a sublethal L1210 tumor challenge. The differing effects on survival to the L1210 tumor challenge could not be attributed to a direct cytotoxic effect of SM on the L1210 tumor cells as SM did not increase the survival rate or median survival time of syngeneic mice exposed to a lethal L1210 tumor cell challenge. In summary, SM and CP had immunosuppressive effects in the humoral immune assay. Although neither compound suppressed the delayed hypersensitivity response, CP was found to suppress host resistance to L1210 tumor cells.

Effects of NO2 on immune responses.

The effects of NO2 on immune responses of mice were investigated. Mice were exposed to various concentrations of NO2 in inhalation chambers. After exposure the following parameters were measured: phagocytosis of polystyrene beads by both peritoneal and alveolar macrophages, production of antibody-forming cells from mice immunized with sheep erythrocytes, lymphocyte blastogenesis of splenic cells, and susceptibility to influenza virus. The production of antibody-forming cells was reduced in mice that were exposed to 5 ppm NO2. The serum antibody titers, phagocytosis, and other immune parameters measured were not affected. Exposure to NO2 did not affect mortality to influenza virus. These data indicate that certain immune parameters were altered by exposure to NO2; however, NO2 does not appear to be a major immunosuppressive factor at the concentrations tested.

Immunomodulating activities of ethylene-2,2'-bis(dithio)bis(ethanol) and related compounds. Effects on murine lymphocyte proliferation and functions in vitro

HEDS (2-hydroxyethyl disulfide) and structurally related compounds were found to affect lymphocyte proliferation and functions in vitro. HEDS, and ADA 202–718 (ethylene-2-2'-bis(dithio)bis(ethanol)), were shown to stimulate the proliferation of murine spleen cells. The growth of populations of murine T-cells (thymocytes) was not stimulated. HEDS and ADA 202–718 enhanced the allogeneic response in the mixed lymphocyte reaction and stimulated the formation of antibody producing cells in a primary humoral immune response. A similar stimulatory effect was observed in a secondary humoral response towards a T-cell specific antigen, DNP-keyhole limpet hemocyanine. Neither ADA 202–718 nor HEDS exhibited γ-interferon inducing ability, when tested on quiescent Balb/c spleen cell cultures in absence of antigen. However, HEDS, and especially ADA 202–718 potentiated the allogen-induced γ-interferon production and release in the mixed lymphocyte reaction. Both HEDS and ADA 202–718 seemed to induce or stimulate the release of an interleukin-1-like activity as well as interleukin-2 in Balb/c spleen cells. No obvious effect, however, was seen with either compound on maturation or immune-phagocytic activity of bone-marrow derived macrophages.

Role of interleukin 1 in anti-immunoglobulin-induced B cell proliferation.

In this report we describe conditions for polyclonal activation of small numbers of highly purified mouse B lymphocytes. Three signals are required for induction of DNA synthesis by the particular subset of small B lymphocytes investigated: a signal delivered by antibodies specific for the IgM receptor expressed on the B cell membrane; a signal delivered by a T cell-derived factor (B cell growth factor [BCGF]); and a signal delivered by the macrophage-derived factor interleukin 1 (IL-1). The conclusion that IL-1 has B cell co-stimulator activity is based on the findings that highly purified preparations of mouse and human IL-1 have the capacity to cause proliferation in B cells treated with anti-IgM and BCGF. Such cultures show an absolute dependence on exogenously added IL-1 when 2-mercaptoethanol is omitted from the medium. BCGF and IL-1 each act in a non-antigen-specific, non-H-2-restricted, synergistic manner. Their requirement is not observed when B cells are cultured at high density, presumably reflecting accessory cell contamination and endogenous factor production under these conditions. The B cell activation induced by these three signals is restricted to proliferation without the production of antibody-forming cells.

60] General considerations in measurement of interferon-induced antibody suppression

This chapter discusses interferon-induced antibody suppression and the assay systems. The detailed analysis of the kinetics of immunosuppression, particularly in relation to the heterogeneity of responding cells are presented, which directly affects the level of suppression observed. Therefore, the discussion in the chapter concentrates on the means whereby an assay system can be devised to minimize the problems that result from having a complex cell system to assay possible complex effects of an immunosuppressive agent such as IF. The main approach has been to stimulate the appropriate precursor cells with antigens or mitogens in the presence of IF and to measure the number of specific antibody-forming cells (AFC) which are produced. In studying the effect of IF (or any other soluble mediator) on a complex, multicomponent system, two important points emerge which are (a) that there are a number of possible targets on which IF might act and (b) that its effect will depend not only on the time at which it is added, but also on the time at which the response is measured. The production of AFC involves a combination of two processes, which are the activation of antigen-specific precursor B cells and the clonal expansion of each activated precursor. By employing various lymphocyte fractionation techniques and clonal analyses, it is possible to gain a clearer understanding of the mechanics of IF-induced antibody suppression. Much of the investigation of IF-induced immunosuppression has been derived from work with mouse interferon and murine in vitro responses.

Adjuvant and immunostimulating activities (in the absence of freund's incomplete adjuvant) of chemically modified low molecular weight mycobacterial peptidoglycans.

Relatively low molecular weight peptidoglycan fragments extracted from two strains of Mycobacterium tuberculosis var. hominis were chemically coupled with lauric acid. The fatty acid conjugates were compared with the native substances with respect to some immunopotentiating activities. In vitro, the mitogenic effect on murine spleen lymphocytes was significantly enhanced following conjugation. One of the lauric acid conjugates stimulated, upon intravenous administration in mice, the formation of antibody-producing cells in the spleen, while the native substance was devoid of such activity. In adjuvanticity tests performed in the guinea pig in the absence of mineral oil, the fatty acid conjugates generally exerted a higher adjuvant effect on antibody production or on delayed type hypersensitivity reactions than did the native preparations.

Rate of development of immunological memory and characteristics of the secondary immune response in mice of strains with high and low reactivity

Immunological memory for sheep's red blood cells develops in mice of strains CBA and DBA/2 and (CBA×C57BL/6)F1 hybrids 24 h after injection of a small dose of the antigen, but 48 h after injection in C57BL/6 mice. The level of the secondary immune response in CBA, C57BL/6 and F1 hybrids is significantly higher than in DBA/2 mice. Maximal production of antibody-forming cells in the spleen of the CBA mice is observed after two injections of small doses of the antigen. By contrast to this, to obtain a marked immune response in the case of adoptive transfer of spleen cells of C57BL/6 mice a second injection of a large dose of antigen is required.

B-cell suppression of antibody response to sheep red blood cells in mice of high- and low-responding genotypes

CBA and C57B1 mice (high and low responders to sheep red blood cells, respectively) were injected intravenously with syngeneic lymph node, marrow, spleen, or thymus cells together with sheep red blood cells (SRBC), and the production of antibody-forming cells (AFC) was assayed in the spleen. Transfer of lymph node, marrow, spleen, or thymus cells led to a significant enhancement of immune responsiveness in low-responding C57B1 mice. In contrast, transfer of marrow, lymph node, or spleen cells to high-responding CBA mice was accompanied by a decline in AFC production. These effects were magnified if syngeneic cell donors had been primed with SRBC; suppression in CBA mice and stimulation in C57B1 mice were especially pronounced after transfer of SRBC-primed lymphoid cells. Pretreatment of CBA donors with cyclophosphamide in a dose causing selective B-cell depletion completely abrogated the suppression of immune responsiveness. A large dose (10 7) of syngeneic B cells injected together with SRBC suppressed the accumulation of AFC in both CBA and C57B1 mice. No suppression of immune responsiveness was observed after transfer of intact thymus cells, hydrocortisone-resistant thymocytes, or activated T cells. We conclude that suppression of the immune response to SRBC is induced by B cells. At the same time, there is a possibility that the addition of “excess” B cells acts as a signal, triggering suppressor T cells.

Correction of immune response to sheep's erythrocytes by polyelectrolytes in different strains of mice

Inbred mice of strain C57BL, A, C57BR, C3H, and CBA, with low, medium, and high reactivity to sheep's red cells (SRBC), were injected with the polymers poly-4-vinylpyridine (P4VP) and polyacrylic acid (PAA) and immunized with SRBC; production of antibody-forming cells (AFC) in their spleen was then determined. Injection of P4VP into C57BL mice was shown to produce a fivefold increase, and injection of PAA a fourfold increase in the immune response. Injection of P4VP and PAA into mice of strain A increased the immune response by 2.5 and 2.8 times respectively. The immune response in C57BR mice was increased fourfold by P4VP and by 4.7 times by PAA. Treatment of C3H mice with P4VP increased the immune response by twice, and treatment with PAA by 2.4 times. Injection of P4VP and PAA into CBA mice did not affect the intensity of the immune response. With an increase in the immunological reactivity toward SRBC in the mice of these strains a decrease in the potentiating effect of polymers on the immune response was thus observed.

Migration and interaction of T- and B-lymphocytes in mice of different genotypes

Interaction between bone marrow cells and thymus cells in CBA, C57BL, and F1 (CBA×C57BL) mice was investigated during the production of antibody-forming cells against sheep's erythrocytes when the thymus and part of the bone marrow was screened during lethal irradiation. Genetically determined differences between CBA mice, reacting strongly to erythrocytes, and C57BL mice, with a low level of reaction, were shown to be associated with differences in the intensity of cooperative interaction between the B- and T-cells; under the conditions used this difference in intensity evidently depends on differences in the intensity of migration of the B-lymphocytes from the bone marrow or T-lymphocytes from the thymus.

Changes in the number of antibody-forming and stem cells in the spleen of unilaterally nephrectomized mice

A suspension of spleen cells from intact and unilaterally nephrectomized mice, obtained in the latter case 19–21 h after the operation, was injected into lethally irradiated (CBA×C57BL/6) F1 mice 24 h after irradiation. On the 8th day after injection of the cells the number of colony-forming and plaque-forming cells in the spleen of the irradiated recipients was determined. To determine the number of plaque-forming cells, a mixture of spleen cells and sheep's red cells was injected into the irradiated recipients. The number of colonies in the recipients' spleen 19–21 h after the operation either was unchanged or was significantly reduced. Stimulation of the production of antibody-forming cells was observed at these same times, and it coincided in time with the period of manifestation of the ability of the splenic lymphoid cells of the unilaterally nephrectomized mice to induce proliferation in the kidney of the intact recipients.

Immunosuppression mediated by Ehrlich ascites fluid

One intraperitoneal injection of cell-free Ehrlich ascites fluid, which did not induce the tumour, inhibited very strongly, and for a long time, the formation of antibody-producing cells (PFC's) in the spleens of recipient mice. The same procedure did not alter cellular immunological reactivity of these splenocytes (assayed by graft-vs-host reaction). At the time when the strongest suppression of PFC formation was registered, cellularity of the spleens increased. Several interpretations of these data are given, the most attractive one being that immunosuppressive agent(s) present in the ascitic fluid of Ehrlich tumour selectively eliminated or inhibited function of the population of B lymphocytes.

Immune processes in vitamin deficiency states

Studies conducted in our laboratory, relating to the development of immune processes in vitamin-deficient experimental animals, have been reviewed. 1) The significant participation of these nutritional factors in the production of circulating antibodies to a variety of antigens, and in the manifestation of delayed hypersensitivity reactions, including the rejection of tissue transplants, has been described. 2) Investigations on the mode of action of vitamin B6 and pantothenic acid have demonstrated a marked reduction in the production of antibody-forming cells following antigenic stimulation in both deficiency states. The metabolism of antigen appeared to be normal. However, these two vitamins seem to function at different loci in the development of the immune process. Whereas vitamin B6 appears to be necessary for the production of “C1” units from serine, which are required for the biosynthesis of nucleic acids, it seems likely that pantothenic acid is involved in the secretion of newly-synthesized proteins into the extracellular compartment.