Characteristics of the immune response to staphylococcal anatoxin in healthy persons

The immunization of young healthy males with adsorbed staphylococcal toxoid revealed the existence of two main types of postimmunization humoral response. One of them was characterized by an early (on days 7-14 after immunization) rise in the titer of antibodies with the subsequent gradual decrease of their content, while the number of circulating T-suppressors remained unchanged. The characteristic feature of the other type was a slow rise in the level of antitoxins by day 21 after immunization with the subsequent drop of their titers, preceded by a considerable increase in the number of circulating T-suppressors. The maximum antibody titer was definitely higher in the first type of response than in the second type (14.8 +/- 1.41 and 9.0 +/- 1.53 I. U./ml respectively). A single plasmapheresis procedure on day 21 after immunization produced no essential effect on the dynamics of the characteristics of cell-mediated and humoral immunity.     

The role of T-suppressors in the formation of plague immunity in mice

Antiplague immunization of mice causes an increase in the number of T-suppressors in their thymus and spleen; this increase is especially pronounced after injection of a low-immunogenic dose of the vaccine strain. T-suppressors specific to Yersinia pestis antigens were detected in the thymus on day 3 after a single injection and on day 14 after two injections of the vaccine strain.     

Analysis of certain mechanisms of antigen-specific suppression of the immune response

CBA mice were immunized with sheep red blood cells (SRBC) to obtain immune spleen cells (ISc) which were used to suppressor cells. Administration of ISC to intact syngeneic recipients on the immunization day led to a more powerful suppression of the immune response as compared to that seen one day after antigen injection. Four days after immunization the animals' immune response was not liable to be suppressed. ISC extract possessed similar effects with respect to the immune response of normal spleen cells which were transplanted to the cyclophosphamide-treated recipients. The immune response of spleen cells from mice immunized with SRBC in a dose of 10(6) was less liable to be suppressed. Hyperimmune spleen cells from donors immunized with SRC in a dose of 10(9) were insensitive to ISC or to the extract. Experiments with the use of adoptive transfer of a mixture of immune and intact T- and B-cells have disclosed that B-cells from hyperimmune donors were resistant to suppression. Therefore, B-lymphocytes are the most probable target cells exposed to T-suppressors in the given system. The mechanism is discussed of the selective effect of T-suppressors on B-cells in the course of the immune response development during immunization with high doses of antigen.     

Immunomodulating properties of non-steroidal anti-inflammatory drugs]

There has been studied the influence of a number of nonsteroidal antiinflammatory agents (NSAID ) on the humoral immune response of mice in immunization with erythrocytic and viral antigen. It has been found out that NSAID have immunomodulating effect, stimulating humoral immune response (4-iodantipyrine, 4-bromantipyrine) or suppressing it (butadione, sodium salicylate). Apparently the mechanism of NSAID ++ immunostimulating effect is related to the inhibition of T-suppressors ++ function by the latter ones.     

Specificity of suppressor cells of the delayed hypersensitivity type for Staphylococcus studied by immunoadsorption

The immunological specificity of T-suppressors obtained from mice after intravenous immunization with corpuscular antigen was shown. The splenocytes of such a mice suppressed DH to staphylococcal antigens, but not to sheep red blood cells. The suppressor cells under study were specifically adhesive to staphylococci.     

The effects of 2-methoxyoestrogen sulphamates on the in vitro and in vivo proliferation of breast cancer cells

2-Methoxyoestrogen sulphamates are a new class of compounds, which inhibit breast cancer cell proliferation and are also potent inhibitors of steroid sulphatase (STS) activity. In the present study, we have used two cell proliferation assays (MTS and AB) to identify potent new compounds in this class. Similar IC(50) values were obtained using these assays with two of the most potent compounds identified being 2-methoxyoestradiol-bis-sulphamate (2-MeOE2bisMATE) and 2-methoxyoestradiol-17beta-cyanomethyl-3-O-sulphamate (2-MeOE2CyMATE). Both compounds inhibited the proliferation of MCF-7 (ER+) and MDA-MB-231 (ER-) breast cancer cells. Using the AB assay, which allows repeat measurements of cell proliferation without killing cells, both compounds were shown to inhibit cell proliferation in an irreversible manner. As STS may be involved in the removal of the sulphamoyl moiety of these compounds, which could reduce their potency, their ability to inhibit the proliferation of MCF-7 cells transfected with the cDNA for STS was also examined. Although the STS activity was 20-fold higher in these cells than in non-transfected MCF-7 cells, no decrease in the ability of these compounds to inhibit cell proliferation was detected. To test the efficacy of these compounds in vivo, nude mice were inoculated with MCF-7 cells in Matrigel and stimulated to grow with oestradiol. Three weeks after the oral administration of 2-MeOE2bisMATE or 2-MeOE2CyMATE (20mg/kg/day, 5 days/week) tumour volumes had regressed by 52% and 22%, respectively. Both compounds also inhibited liver and tumour STS activity by >90%. The potent anti-proliferative effects of these compounds, and their ability to inhibit tumour growth and STS activity in vivo, indicates that they are suitable for development as novel therapeutic agents, which should be active against a wide range of cancers.     

Anti-idiotype responses abrogate anti-CD4–induced tolerance to a tumor-specific antigen and promote systemic tumor immunity

Purpose: Immunologic-based cancer treatment modalities represent an active area of investigation. Included in these strategies are passive administration of monoclonal antibodies which recognize tumor-associated antigens and active vaccination with identified tumor antigens. However, several problems associated with these types of treatment strategies have been identified. Methods: In this report, we address certain issues by employing a murine model for experimental pulmonary metastasis and a tumor antigen vaccination strategy that induces complete tumor immunity in this system. Utilizing this model, we attempt to address issues related to unresponsiveness to tumor antigen immunization induced by passive administration of a rat monoclonal anti-CD4 and the induction of anti-idiotype responses to a passively administered monoclonal antibody and the effects on the induction of tumor immunity. Results: The results presented indicate that passive administration of rat monoclonal anti-CD4 exhibits immunosuppressive effects that inhibit the production of antibodies to the tumor antigen immunization and abolishes tumor immunity. Repeated administration of the rat monoclonal anti-CD4 results in an anti-idiotype response that can abrogate unresponsiveness to tumor antigen immunization and promote systemic tumor immunity. Conclusions: The data examine a number of potential problems associated with immunologic-based treatments for cancer. These problems include the potential for tolerance to the tumor antigen and establishing an immunocompromised state where immunization with a tumor antigen failed to generate tumor immunity. Approaches to eliminate tolerant T cells by targeting anti-CD4 via anti-idiotype responses that could be generated in vivo without CD4⁺ T cells allowed for recovery of nontolerant T cells, and an antibody response to the tumor antigen that results in tumor immunity.Abbreviations CTL Cytotoxic T lymphocyte - FITC Fluorescein isothiocyanate - OD Optical density - PBS Phosphate-buffered saline - SV40 Simian virus 40     

Interrelations of cellular and humoral immune response and different doses of sheep erythrocytes in mice]

In experiments on CBA and C57BL/6 mice the generation of antibody-forming cells respectively either in the popliteal lymph nodes or spleen as well as a rate of delayed type hypersensitivity response (DTHR) on the background of subcutaneous (into foot) or intraperitoneal injection of different doses of sheep erythrocytes (from 10(4) to 10(8)) have been studied. In so doing two types of immune response can be isolated on the dependence upon the sensitivity threshold to antigen of DTHR and humoral immunity. Thus in C57BL/6 mice the antigen threshold for DTHR is of one time (in intraperitoneal immunization) or of a two times (in subcutaneous) lower order than for antibody response. In CBA line mice under subcutaneous immunization there can be seen quite an opposite picture while intraperitoneal immunization causes exact correlation of antigen threshold for cellular and humoral immune response.     

Enhancement of the in vivo antibody response by an 8-derivatized guanine nucleoside

The capacity of the C8-substituted guanine ribonucleosides to enhance the in vivo humoral immune response to the protein antigen, human gamma globulin (HGG), in A/J mice was evaluated. It has been shown recently that the C8-substituted guanine ribonucleosides are a new class of potent adjuvant for humoral immune responses to the sheep erythrocyte antigen. The current studies extend these findings to HGG with 8-bromoguanosine (8BrGuo), a representative of this group of nucleosides. The adjuvant activity of 8BrGuo in this system is highly dose and time dependent. Although 8BrGuo enhanced responses when injected either early (Day 0) or late (Day 4 or 5) after immunization, its administration on Day 1 or 2 most often led to no enhancement, suggesting that 8BrGuo may act on two events separated by a resistant stage in an ongoing immune response. The plaque-forming cell (PFC) response to HGG was enhanced optimally at doses as low as 1 mg 8BrGuo/mouse administered either on the day of immunization or 4 days thereafter. In contrast, however, serum anti-HGG antibody concentration assayed by enzyme-linked immunosorbent assay (ELISA) was enhanced only at doses of 10 mg or more, injected on the day of immunization, but doses as low as 1 mg were effective on Day 4. 8BrGuo was also an effective adjuvant when injected after antigen administration in incomplete Freund's adjuvant or when administered by several different routes (intraperitoneal, subcutaneous, oral).     

Immunosuppression in experimental staphylococcal infection and its effect on antimicrobial resistance

Experiments on CBA and (CBA X C57BL)F1 mice have revealed that a prolonged period of antigen-nonspecific immunosuppression of humoral immunity develops in experimental staphylococcal infection; this period of suppression may be preceded by a short phase of antigen-nonspecific immunostimulation. Immunosuppression is linked with the accumulation of antigen-nonspecific T-suppressors in the spleen, these T-suppressors being capable of the manifestation of their activity both in vitro and in vivo in cases of their transplantation to semi-syngeneic recipients. Immunosuppression does not aggravate the course of staphylococcal infection and is accompanied by an increase in resistance to Pseudomonas aeruginosa superinfection, which is due to the stimulation of inflammatory reaction at the site of the injection of the superinfecting agent.     

Relationship between tumour resistance and the in vitro and in vivo cytotoxicity elicited by Salmonella antigens in immunized mice

Immunization of mice with the attenuated 11RX strain of Salmonella enteritidis (11RX) induces resistance to intraperitoneal (i.p.) tumour growth. Tumour resistance is much greater and lasts for a longer time following i.p. immunization than following intravenous (i.v.) immunization. This paper extends our previous observations that, after this resistance is lost, it can be recalled by a T cell-mediated reaction to an antigenic extract of the bacteria (11RX antigen) which is not protective in unimmunized mice. The duration of this sensitization to 11RX antigen was determined in mice immunized i.p. or i.v. with live 11RX by challenging them at various times after immunization with 131I (or 125I)-labelled Ehrlich ascites tumour (EAT) cells alone or mixed with 11RX antigen. In vivo killing of EAT cells was assayed by monitoring whole-body retention of radioactivity and this was correlated in the same mice with suppression of tumour growth and survival of the mice. The resistance recalled by 11RX antigen was short-lived and in vivo cytotoxic activity had subsided by 6 days after antigen injection. 11RX antigen also recalled the ability of the peritoneal cells to lyse 51Cr-labelled EAT cells in vitro and a close correlation was found between this activity and the cytotoxicity measured in vivo. The adherence properties of the cytotoxic cells and their inhibition by trypan blue indicated that they were macrophages.     

Modulation of mitogen-induced spleen cell proliferation and the antibody-forming cell response by beta-endorphin in vivo

Experiments were conducted which compared the in vivo effects of beta-endorphin (BEP), gamma-endorphin (gamma EP), methionine-enkephalin (Met-ENK), and acetylated BEP(1-27) on the in vitro proliferative response of rat spleen cells to concanavalin A (ConA). In addition, the influence of BEP administration on the primary and secondary antibody-forming cell (AFC) response to the soluble antigen keyhole-limpet hemocyanin (KLH) was examined. Intravenous administration of BEP enhanced the spleen cell proliferative response to ConA assessed 3 hr after a single bolus infusion. Conversely, infusion with AcBEP(1-27) suppressed the proliferative response, whereas no effects of intravenous gamma EP or Met-ENK treatment were observed. The enhancing effect of BEP administration was not detectable 24 hr after a single infusion, but could be maintained over a 44 hr period by multiple infusions. The primary AFC response to KLH was suppressed by a dose of 1 nmole BEP only. On the other hand, the secondary IgG AFC response to KLH was enhanced by 10 pmoles BEP, while the IgM and IgA AFC responses remained unaltered by BEP treatment. The anamnestic in vitro proliferative response of spleen cells cultured with KLH was not altered if BEP was administered at the time of secondary KLH immunization. These results extend previous observations of BEP-induced modulation of in vitro immune function by demonstrating that opioid and nonopioid forms of BEP administered in vivo alter the capacity of spleen cells to proliferate and develop antibody responses to antigen.     

Determinant specific suppression of antigen-induced T cell proliferation in the guinea pig. I. Quantitation of suppressed antigen-specific T cell responses as a consequence of prior exposure to antigen in incomplete Freund's adjuvant

We have asked whether a correlation exists between T cell proliferation and the in vivo suppression of delayed type hypersensitivity observed after administration of antigen in incomplete Freund's adjuvant before exposure to antigen in complete Freund's adjuvant. We find that in vivo suppression is indeed paralleled by diminished in vitro responsiveness to the immunogen. Suppression of T cell proliferation is antigen-specific, dependent upon prior immunization of antigen in IFA, and can be transferred adaptively into unprimed but not primed animals by lymphoid cells from actively suppressed syngeneic donors.     

The effect of sodium thiosulfate on the metabolism of cis-platin in human plasma in vitro

The anticancer drug cis-platin (CP) is widely used to treat patients, but it is also associated with significant side effects, including nephrotoxicity. Given that this metallodrug is intravenously (iv) administered, its biotransformations in the bloodstream are likely to be involved in mediating these side-effects. Previous studies have revealed that the iv administration of patients/mammalian model organisms with sodium thiosulfate (STS) can ameliorate the side effects of CP, but the underlying molecular basis remains elusive. We have studied the effect of STS on the metabolism of CP in human plasma in vitro by determining the platinum (Pt) distribution using size exclusion chromatography (SEC) coupled on-line to an inductively coupled plasma atomic emission spectrometer (ICP-AES). The addition of STS to plasma 10 min before CP was added accelerated the hydrolysis of CP and resulted in the formation of a Pt-STS complex. Conversely, when plasma was incubated with CP for up to 3 h and STS was added thereafter the analysis of the obtained mixture revealed that the formation of the same Pt-STS complex which in turn greatly diminished the plasma protein binding of CP-derived hydrolysis products. Thus, the observed amelioration of the side effects of CP by STS can be rationalized in terms of the rapid formation of a biologically inactive Pt-STS complex in the bloodstream. This is the first mechanism that can explain the amelioration of the side effects of CP by STS. Based on the fact that cis-platin remained in plasma for a considerable amount of time, the optimization of the administration sequence, the molar ratio and the time delay between the administration of both drugs emerges as a viable strategy to achieve a careful balance between ameliorating the side effects while leaving the antitumour activity intact. Our results demonstrate that in vitro studies can be useful to develop feasible strategies to mitigate the side-effects of Pt-based anticancer drugs in patients.     

Local but not systemic administration of IFN-gamma during the sensitization phase of protein antigen immunization suppress Th2 development in a murine model of atopic dermatitis

Biphasic Th1/Th2 development plays a central role in the pathogenesis of atopic dermatitis. In the sensitization phase after protein antigen exposure, an immune response polarized toward Th2 differentiation, which is due to the hosts' genetic proneness to the disease, initiates the skin lesions. Th1/Th2 antagonism is a potential mechanism that could be manipulated to suppress the initial Th2 deviation. IL-12 is the key cytokine for Th1 differentiation. Interferon gamma (IFN-gamma) can assist Th1 development through several mechanisms and suppress Th2 differentiation. We took advantage of a recently developed murine model of atopic dermatitis elicited by epicutaneous sensitization with protein antigen through patch application to examine the effects of different routes of IFN-gamma administration on Th1/Th2 differentiation during the sensitization phase of antigen exposure. Our data showed that systemic administration of IFN-gamma during the sensitization phase could not promote serum levels of specific IgG(2a). However, local administration (intradermal injection or patch application) of IFN-gamma during the sensitization phase could promote serum levels of specific IgG(2a) and suppress serum levels of specific IgE. Moreover, pretreatment of local IFN-gamma with protein antigen has a long-term modulatory effect on serum levels of specific IgG(2a) and IgE after repeated antigen immunization. Our results demonstrate that local but not systemic administration of IFN-gamma during the sensitization phase of protein antigen immunization could suppress the Th2 deviation in this murine model of atopic dermatitis. Thus, this may represent a novel strategy for the treatment and prevention of atopic dermatitis.     

Inhibition of experimental autoimmune thyroiditis in mice by anti-I-A antibodies

Experimental autoimmune thyroiditis is induced in mice by immunization with thyroglobulin emulsified in Freund's complete adjuvant. The disease is characterized both by thyroid infiltration with mononuclear cells and by circulating thyroglobulin antibodies. The magnitude of the thyroid infiltration and the titer of thyroglobulin antibodies are controlled by genes in the I-A subregion of the major histocompatibility complex (H-2). We investigated the in vivo effect of monoclonal anti-Ia antibodies on experimental autoimmune thyroiditis in susceptible mice. Antibodies were given around the time of immunization, later after immunization, and to mice with established disease. Monoclonal antibody produced by the hybridoma line 10-3.6 (anti-I-Ak, s, u, v, z, f) completely prevented both production of thyroglobulin antibodies and thyroid infiltrates, when given shortly before or at the time of antigen administration. This effect was dose-dependent and this monoclonal antibody decreased the severity of the disease when given after the antigen challenge but did not fully suppress established thyroiditis. The same antibody markedly decreased the number of B lymphocytes in the spleen and decreased the thyroglobulin-induced spleen cell proliferation when either given in vivo or added in vitro to cell cultures. Antibodies produced by the hybridoma line 11.2.12 (anti-I-Ak) did not show an inhibitory effect on the disease. These experiments suggest that in this model of murine thyroiditis anti-Ia antibodies act on antigen-presenting cells. Furthermore, only one monoclonal antibody, anti-Ia, suppressed the immune response to thyroglobulin, suggesting a possible role for the isotype and specificity of anti-Ia antibody.     

Production of monoclonal antibody to bovine leukemia virus glycoprotein gp51 by in vitro immunization

The monoclonal antibody against glycoprotein gp51 of bovine leukemia virus (BLV) envelope antigen was produced by in vitro immunization. This monoclonal antibody reacted with viral antigen was observed at the 69 kilodalton (kDa) glycoprotein. However, this monoclonal antibody was not involved in neutralizing. It was shown that in comparison with in vivo immunization, in vitro immunization has some advantages, namely a short immunization period and a small antigen quantity.     

Hormonal regulation of four urea cycle enzymes in postnatal rat liver in organ culture

The administration of hydrocortisone to 3- to 15-day-old rats increased the levels of hepatic argininosuccinate synthetase (ASS) and arginase. In 13-day-old rat liver explants maintained in organ culture, ornithine carbamoyltransferase (OTC), carbamoylphosphate synthetase (CPS) and arginase were stimulated by betamethasone. Actinomycin D prevented the responses of the latter two enzymes. Dibutyryl cyclic AMP raised OTC, CPS, ASS and arginase in vitro. The responses of the latter three enzymes were blocked by cycloheximide and puromycin and partially inhibited by actinomycin D. The simultaneous presence of betamethasone and dibutyryl cyclic AMP in the culture medium raised CPS and OTC in an additive manner. The sequential treatment of the cultures with betamethasone followed by dibutyryl cyclic AMP increased CPS and arginase synergistically and amplified the response of ASS to dibutyryl cyclic AMP.     

Mechanism of the suppressive effect of interferon on antibody synthesis in vivo.

Mouse interferon preparations significantly suppress the in vivo antibody response to sheep red blood cells (SRBC), a thymus-dependent antigen, and to Salmonella typhimurium lipopolysaccharide (LPS), a thymus-independent antigen. It is also possible to effect the late responses of antigen sensitive "memory" cells observed during secondary immunization by administration of interferon prior to primary immunization. The immunosuppressive activity of interferon was time- and dose-dependent. Maximum suppression was produced when animals were given 1.5 times 10-5 units of interferon between 4 and 48 hr before antigenic stimulation. These findings suggested that interferon affects some early event(s) in the process of antibody synthesis which might be related to the general inhibitory effect of interferon on rapidly dividing cells and viral m-RNA translation. In addition, the use of nonadherent spleen cell cultures from interferon-treated mice, immunized in vitro with a thymus-independent antigen, indicated that in this situation the inhibitory effect of interferon was due to an action on B lymphocytes. A variety of soluble "suppressive" factors are secreted by T cells as a consequence of activation by mitogens or specific antigens in vitro. Since T cells are recognized as one of the sources of interferon, it is suggested that interferon should be investigated as a suppressor T cell-produced lymphokine which can regulate B cell expression.     

Parallel formation of delayed-hypersensitivity effectors and T-suppressors in the intraperitoneal administration of a massive dose of ram erythrocytes

Injection of 6 X 10(9) sheep red blood cells (SRBC) to mice led to parallel formation, on days 4-5, of delayed hypersensitivity effector cells (the activity was tested in local transfer experiments) and delayed hypersensitivity T-suppressors preventing sensitization of syngeneic recipients. After massive injection of SRBC the activity of spleen suppressors gave 2 peaks: on days 5 and 14. Five days after massive antigen injection only T-cells capable of sorbing on a specific antigen manifested suppressor activity. On day 14 T-cells capable of sorbing on specific antibodies showed a specific activity, whereas T-cells capable of sorbing on a specific antigen retained only part of their activity. The mechanism of delayed hypersensitivity inhibition following massive antigen injection by suppressors obtained by day 5 is reviewed in terms of Germain and Benacerraf's theory postulating that delayed hypersensitivity is regulated by Ly 2+, I-J+ antiidiotypic suppressors capable of sorbing on specific antibodies and formed upon injection of Ly 1+, I-J+, Id+ inductor cells capable of sorbing on a specific antigen.