Anti-theta Serum Research Articles

Characteristics of DTH suppressor cells in mice infected with Candida albicans

Inoculation of 10(8) C. albicans intraperitoneally into Balb/c mice at given dosage was reported to induce suppression of antigen-specific delayed-type hypersensitivity. Adoptive transfer of spleen cells into normal syngeneic mice pre-treated with Cyclophosphamide confirmed the existence of suppressor cells in mice. Such cells were sensitive to treatment with anti-theta serum and complement, non-adherent to Sephadex G-10. A pretreatment of the mice with Cyclophosphamide eliminated DTH suppression. Treatment with antimacrophage agents via intraperitoneal abrogated suppression only if being effected before inoculation of alive 10(8) Candida albicans. It is concluded that the spleen suppressor cell is a T-lymphocyte whose precursor is Cyclophosphamide-sensitive, requiring the macrophage to be induced.

Babesia microti in mice. Adoptive transfer of immunity with serum and cells.

Protection against infection with Babesia microti in mice was passively transferred with either serum or cells. Immune spleen cells were more effective than were immune mesenteric lymph node (MLN) cells in reducing parasitaemias in recipient mice, and the level of protection correlated with the donor to recipient ratio rather than with the number of cells transferred. Protection against primary infection in recipient mice was adoptively transferred by nylon wool adherent, B cell enriched subpopulations of spleen cells. In contrast, higher peak parasitaemias were apparent in mice which received nylon wool non-adherent, T enriched spleen cells in comparison with the control mice. However, if the recipient mice were reinfected, control of this second infection was greatest in the recipients of the T enriched cell subpopulation. Treatment of the protective nylon wool adherent, B cell subpopulation with anti-theta serum and complement abolished both the protective effect and the anamnestic antibody response in the recipient mice. This suggested that primed T cells controlled the expression of the protective antibody response by primed B cells.

Legionella pneumophila-induced blastogenesis of murine lymphoid cells in vitro.

Legionella pneumophilia antigen preparations, either killed whole cell vaccine, a soluble sonic extract, or a purified large-molecular-weight somatic antigen, stimulated blastogenic responses by splenocytes from both normal and Legionella-sensitized mice. Graded amounts of the bacterial preparations, when added to cultures of normal spleen cells, resulted in increased uptake of thymidine into cellular DNA, indicating that the preparations were mitogenic for normal mouse splenocytes. Spleen cells from mice injected with graded numbers of living bacteria showed blastogenic responsiveness to Legionella preparations generally at a higher level than spleen cells from normal animals. The heightened blastogenic response was mainly evident with spleen cells obtained from mice injected with living bacteria 2 to 3 weeks earlier. Splenocytes from mice infected with legionella less than 1 to 2 weeks or for more than 4 to 5 weeks responded generally similar to those obtained from uninjected mice, indicating that sensitization with living organisms had a relatively short duration. Spleen cell suspensions responding to the L. pneumophila antigens appeared to be mainly B-lymphocytes since cell suspensions from athymic nude mice deficient in T-cells responded as well as cells from conventional mice. Furthermore, passage of splenocytes over nylon wool columns to obtain B-cell-enriched preparations resulted in cell populations capable of responding to Legionella antigen. The cell fractions rich in T-cells were much less capable of responding to the Legionella antigens. In addition, treatment of spleen cell populations with antitheta serum plus complement failed to inhibit the blastogenic response, whereas the same spleen cell preparations treated with anti-mouse immunoglobulin serum plus complement markedly diminished blastogenic responsiveness, again consistent with the likelihood that B-lymphocytes were the major cell class responding to the Legionella preparations.

Staphylococcus aureus inhibits contact sensitivity to oxazolone by activating suppressor B cells in mice.

Killed Staphylococcus aureus strain Cowan I cells inhibit contact sensitivity to oxazolone in mice, when given intravenously 24-72 h before sensitization. With transfer experiments it was found that the cells responsible for the suppression are antigen-specific, nylon-adherent, resistant to antitheta serum + C, and sensitive to anti-mouse Ig serum + C. These suppressor B cells bear anti-oxazolone immunoglobulins and appear to exert their suppressive activity by preventing the contact sensitizer from reaching the specific reactive T cells.

Naturally Occurring Cell-Mediated Cytotoxicity Against Simian Virus 40-Transformed 3T3 Murine Tumor Cells<xref ref-type="fn" rid="fn2">2</xref><xref ref-type="fn" rid="fn3">3</xref>

The tumorigenicity and host protective mechanisms induced by simian virus 40 (SV40)-transformed 3T3 cells (SV403T3) were evaluated in syngeneic BALB/c mice. Tumors were regularly produced by sc inoculation of SV403T3 cells; the incidence, latent period, and survival were proportional to the size of the initial inoculum. With the use of an in vitro 18-hour 51Cr cytotoxicity assay, spleen cells from normal mice showed a dose-related killing activity against the SV403T3 cells. At an effector cell-to-target cell ratio of 200:1, the average lysis was 56 +/- 6%. This reaction appeared specific for the virally transformed targets; the mean lysis of parent 3T3 cells was 23 +/- 5%. Effectors were resistant to anti-theta serum and not removed by adherence to plastic or nylon wool. Tissue distribution studies indicated that these effectors were present in high concentrations in spleen, bone marrow, lymph nodes, and peritoneal cavity. Low levels of activity were associated with cells from the thymus. In the present studies specific T-cell cytotoxicity against the SV403T3 cells could not be demonstrated. Animals challenged with nonviable SV403T3 cells prior to tumor cell inoculation did not show increased in vivo resistance. In parallel, the in vitro cytotoxicity of animals inoculated with SV403T3 tumor cells showed no heightened cell killing compared to the cytotoxicity of normal controls.

Herpes simplex virus type 1-induced interferon production and activation of natural killer cells in mice.

Cultures of peritoneal exudate cells (PEC) of non-immune C57BL/6 mice reacted to infectious herpes simplex virus type 1 (HSV), but not to non-infectious virions by producing interferon. Similarly, when the virus preparations were injected intraperitoneally (i.p.) and interferon was determined in the wash-out fluid, interferon could only be detected after injection of infectious HSV. All interferons were mixtures of interferon-alpha and -beta. The cell that produced interferon was not sensitive to anti-theta serum or anti-asialo-GM1 serum, it was retained on nylon wool columns and removed by plastic adherence. Silica treatment of PEC abolished HSV-induced interferon production whereas irradiation by 2000 R was without effect. Pure cultures of mouse T cells did not produce interferon when treated with infectious HSV whereas pure cultures of mouse bone marrow macrophages did produce interferon. Thus, we suggest that in PEC the cells which produced interferon were of the macrophage lineage. In further experiments we have analysed in vivo activation of natural killer (NK) cells in the peritoneal cavity of mice. Non-infectious virions were as effective as infectious HSV and doses of HSV too low to induce measurable titres of interferon caused a marked activation of NK cells.

Studies on experimental murine histoplasmosis: host protection and cellular immunity.

Mice immunized subcutaneously with 10(6) viable yeast cells of Histoplasma capsulatum exhibited host protection to a challenge infection and also exhibited concomitant manifestations of cellular immunity to a culture filtrate antigen (yeast extract dialysate (YED)-histoplasmin). Protective immunity was determined by quantitative culture of organisms from infected spleens on a solid medium containing a growth factor produced by the organism. As early as 4 days following immunization, host protection was observed in mice. Positive footpad swelling, indicative of delayed-type hypersensitivity, also was apparent at the early time period (4-7 days) after immunization. By days 14 and 21 following immunization, the mice exhibited maximum footpad responses and maximum splenic clearance of yeast cells. As an additional correlate of cellular immunity, YED-histoplasmin was coupled with chromic chloride to murine erythrocytes (M-RBC) and mixed with immune or nonimmune splenic lymphocytes to detect antigen-specific rosette forming cells (RFC's). Only splenic lymphocytes from mice immunized with H. capsulatum formed antigen-specific RFC's (greater than or equal to 0.2%). Pretreatment of splenic lymphocytes with antitheta (theta) serum plus complement greatly reduced the numbers of RFC's, illustrating their T cell nature. Host protection and cell-mediated immune responses decreased substantially by day 28 following immunization.

Local passive transfer of delayed-type hypersensitivity in the mouse.

A means for local passive transfer of DTH in the mouse has been discovered and partially characterized. Briefly, peritoneal exudates (PE) are induced with incomplete Freund's adjuvant 5 to 7 days after immunization with antigen (usually picGPA) emulsified with complete Freund's adjuvant. Five days later, PE cells are harvested and transferred with soluble antigen into a hind footpad of a naive mouse. Antigen-specific swelling is defined as the change in diameter of footpads over the 24 hr after injection of PE cells plus immunizing antigen minus change in diameter of footpads injected with PE cells plus control antigen. Antigen-specific footpad swelling of 20 to 80% was found in contrast to meager swelling when lymph node or spleen cells were used. The magnitude was dependent on dose of both cells and antigen over a broad range (1 to 12 X 10(6) cells, 1 to 100 micrograms of antigen per recipient). Specific swelling was reduced to nil after treatment of PE cells with anti-theta serum, indicating T cell dependence. Carrier specificity of the response was similar to DTH in the guinea pig. Time course and histology are compatible with a DTH response as well. The superiority of PE cells in local transfer of DTH and the modest requirements of this murine local transfer system provide potent tools for identification of the cell(s) that mediate DTH and their interactions. This system, in addition, should provide a sensitive indicator of immunomodulating activity in cellfree supernatants.

Studies on mouse auto-reactive cells. II. Cytotoxicity exerted by mouse lymphocytes against syngeneic erythrocytes.

In mice, an in vitro cell-mediated cytotoxicity is directed towards syngeneic erythrocytes used as targets. This phenomenon occurs in lymphoid organs of normal, non-immunized mice. Its intensity is greater in the thymus than in spleen or lymph nodes. The study of the nature of the cell responsible for this spontaneous syngeneic cytotoxicity ruled out the role of macrophages. The involvement of T cells in this phenomenon is assessed by its disappearance after the lymphoid cells had been treated with anti-theta serum plus complement, by its disappearance from lymphoid organs of mice previously treated by hydrocortisone, and by its decrease in the presence of synthetic thymic factor. Moreover, spontaneous syngeneic cytotoxicity is lost when lymphoid cells are depleted at autologous rosettes by centrifugation on a Ficoll-Hypaque gradient after rosette formation. Cytotoxic lymphocytes might belong to the population of auto-rosettes previously characterized as a population of immature T cells.

Induction of interferon by mycoplasmas in mouse spleen cell cultures.

Interferon production in cocultures between lymphoma cells and lymphocytes was previously found to be caused by mycoplasma contamination of the tumor cells. Here we have shown that M. arginini caused interferon production in spleen cells of C57BL/6 mice. The producer cell in spleen cell cultures was insensitive to treatment with anti-theta serum plus complement and it was present in spleen cell cultures of homozygous nude mice. Macrophage depletion did not adversely affect interferon production whereas interferon production was abolished in cultures of cells obtained after passage through nylon wool columns. Collectively, these data suggest, but cannot definitively prove that interferon is produced by B cells. By the use of specific antisera it was found that by all probability the interferon produced represented interferon alpha.

Effect of immune heterozygous spleen cell transfer on resistance to mouse hepatitis virus infection in nude mice.

The role of cell mediated immune response to mouse hepatitis virus (MHV) infection in mice was studied by transferring spleen cells from immune heterozygous littermates (nu/+). A suppressive effect on viral growth was seen in infected nude (nu/nu) mice, whereas immune nu/+ serum transfer had no effect. The protective effect of immune nu/+ spleen cells was significantly reduced by treatment with anti-theta serum plus complement but not with anti-Ig serum. In infected nu/nu mice which received transfers of immune nu/+ cells, neutralizing antibody appeared although the titer was not high enough to protect nu/nu mice from fatal infection. Histopathologically, lymphocyte infiltration in hepatic lesions was evident in infected nu/nu mice with nu/+ cell transfer, while it was slight without nu/+ cell transfer.

Anémie induite par Le BCG chez La souris: Nécessité de la présence de lymphocytes thymodépendants

An anaemia and a decreased number of erythroid bone marrow cells were observed on the first weeks following the intravenous injection of 1 mg of BCG into normal mice. No such modification of erythropoiesis was detected in Nude mice. The graft of thymus from new born mice to Nude adults before BCG infection allowed to obtain a drop of packed-cell volume, In previously infected Nude mice, the injection of spleen cells providing from infected normal mice also modified the erythropoiesis. This effect was lost by pretreatment of injected spleen cells with anti-theta serum and complement. Thus, the conditions necessary to the development of the anaemia during BCG infection correlated with the conditions which allowed an immune response against infection.

Studies of the Producer Cell of Herpes Simplex Virus-Induced Interferon in Mouse Spleen Cell Cultures

The producer cell of type I interferon was studied in spleen cell cultures of C57BL/6 mice stimulated by inactivated Herpes Simplex Virus (HSV). Interferon production was not abolished by pretreatment of the spleen cells by anti-theta serum plus complement. The producer cell of interfere on was not removed by plastic adherence and was not destroyed by the addition of silica. It was present in spleens of 3 day old C57BL/6 mice and in spleens of nu/nu mice. It was not inactivated by treatment of nu/nu spleen cells by anti-theta serum plus complement. HSV-induced interferon production was abolished by passage of the spleen cells through nylon wool columns and by irradiation (1000 R) of the spleen cells. Collectively these data suggest that in murine spleen cell cultures type I interferon is produced by B cells. However, our data do not allow to rule out that the interferon producing cell may be an immature macrophage or an immature T cell. References

Migration and cell recruiting activity of specifically sensitized lymphocytes in sponge matrix allografts.

The migration of indium-III-labeled specifically sensitized lymphocytes (SSLs) and unsensitized spleen lymphocytes (ULs) into sponge matrix allografts was studied in irradiated mice. Intravenously injected SSLs were preferentially recruited to the graft bearing the sensitizing alloantigen. High numbers of these SSLs, however, were attracted regardless of their immunological specificity to a third-party graft, when cells sensitized against the third-party antigen were given simultaneously. When SSLs were injected locally into a specific and third-party graft, i.v. injected ULs also homed preferentially to the specific graft. Prior treatment of SSLs with anti-theta serum and complement abrogated their cell recruiting capacity. The percentage of injected lymphocytes recruited to the specific allograft during the first 36 hr increased with the number of SSLs injected. The duration of augmented lymphocyte recruitment, however, was inversely related to the number of SSLs injected into the graft. Furthermore, there is some evidence that SSLs which have been recruited to an allograft migrate to a second site of antigen challenge and initiate there a high lymphocyte recruitment. These results suggest that increased lymphocyte recruitment to an allograft is initiated by an immunologically specific interaction between SSLs and alloantigen. The cells, however, which are preferentially recruited to an allograft, comprise small unsensitized lymphocytes and SSLs regardless of their immunological specificity. Lymphocytes with cell recruiting activity are T cells and appear to be very mobile in vivo. The mechanism described might have an important amplifying effect on allograft rejection.

Production and properties of immune interferon from spleen cell cultures of Toxoplasma-infected mice.

In response to antigenic stimulation, spleen cells from Toxoplasma-infected mice produce a factor showing inhibitory activity against vesicular stomatitis virus infection in L cell cultures. When BALB/C and ICR mice were inoculated intraperitoneally with the low-virulent S-273 strain of T. gondii, such activity was first detected in 4 and 7 days and reached maximum levels at 10 and 14 days respectively, and retained these levels for at least three weeks. However, BALB/C mice, which are considerably more sensitive to Toxoplasma infection than ICR mice, produced significantly smaller amounts of interferon (IF) after challenge with the high virulent strain. The IF produced in this system possessed certain known properties of immune (type II) IF and was not neutralized by rabbit antiserum against mouse type I IF. The immune IF preparation also inhibited multiplication of Toxoplasma within nonphagocytic L cells in an IF-like fashion, whereas Newcastle disease virus-induced (type I) IF had no effect on this parasite. The antiviral and anti-Toxoplasma activity in immune IF preparations could not be distinguished solely on the bases of their molecular weight and isoelectric point. The experiments with anti-theta serum plus complement and with nylon wool column effluent cells strongly suggest that immune IF was produced by T lymphocytes and required the assistance of macrophages.

Chemical enhancement of tumor immunogenicity.

C57BL/6 spleen cells immunized in vitro against syngeneic methylcholanthrene-induced sarcoma cells(MC-1 cells) modified with various chemical reagents show cytotoxic activity against unmodified MC-1 cells in a short-term 51Cr release assay, whereas unmodified MC-1 cells are nonimmunogenic. The effector cells cross-react widely with many other fibroblastic and epithelioid tumors, even those that are not H-2 matched, as well as with some nonneoplastic cells. Priming mice in vivo with a hapten leads to enhanced anti-tumor cytotoxicity developed by spleen cells from these mice immunized in vitro against tumor cells modified with the same hapten. Cytotoxic activity is largely, but not completely, removed by treatment with anti-theta serum and C. The existence of suppressor cells that can inhibit an anti-tumor response is demonstrasted in in vitro immunizations of mixtures of spleen cells from normal mice and mice primed with mitomyhcin-treated tumor cells, the latter suppressing the former.

Enhancement of growth of a radiation-induced lymphoma by T cells from normal mice.

The effect of lymphocytes from normal mice on the growth of a syngeneic, radiation-induced, T-cell-derived lymphoma was investigated. Thymus and spleen cells enhanced the growth of admixed lymphoma cells in a reproducible manner. Growth enhancement was manifested by the earlier appearance and higher final incidence of tumours. Lymphocytes also enhanced the growth of radiation-damaged lymphoma cells. The enhancing activity of spleen cells was predominantly a property of T cells, since it was abolished by treatment with anti-theta serum plus complement and significantly less in spleen cells of nude mice. Tumour-enhancing thymocytes seem to belong to the immature thymic subpopulation, as indicated by their binding to peanut agglutinin.

Regulation of the T 15 idiotype-positive, T-independent immune response to Streptococcus pneumoniae R 36 a (Pn). I. Specific suppressor T cells induced by Pn in vitro are antigen-specific and idiotype-positive.

Normal BALB/c spleen cells incubated for 2 days with Streptococcus pneumoniae R 36 a (Pn) vaccine in vitro acquired the specific ability to inhibit the T-independent anti-Pn response of fresh lymphocytes. The activated suppressor cells were susceptible to cytotoxic treatment with mouse anti-theta serum plus complement, and they inhibited the anti-Pn response of B lymphocytes deprived of T cells. The suppressive effect was eliminated by adsorbtion of cells on dishes coated with the C polysaccharide from Pn, and treatment of the suppressor cells with a rabbit antibody specific for TEPC-15 myeloma protein, in the presence of complement, abolished the suppressive effect. The results indicate that stimulation of lymphocytes with a T-independent antigen in vitro activates suppressor T cells bearing a specific antigen receptor that may express the idiotype(s) characteristic of anti-Pn antibody, and that these cells inhibit the Pn-reactive B cells directly.

Augmentation of erythroid burst formation by the addition of thymocytes and other myelo-lymphoid cells.

Bone marrow from barrier-sustained specific pathogen-free (SPF) CBA and C57BL/6 mice gave relatively low numbers of BFU-E colonies in methylcellulose culture, as compared to conventional mice. Addition of thymocytes to the marrow cultures increased the yield of BFU-E colonies more than fourfold in SPF mice but only 1.5-fold in conventional mice. Colony size was also increased. Increased yield of BFU-E colonies was also obtained by co-culture of bone marrow with lymph node cells or with bone marrow or spleen cells from 900R whole-body irradiated mice. The effect appeared to be cellular rather than humoral. It was not reproduced by conditioned medium from thymus or pokeweed mitogen stimulated spleen cells. The helper effect of thymus cells was eliminated or reduced by freezing and thawing, or by 48 hours of incubation after irradiation. Treatment of bone marrow cells in vitro with anti-theta serum and complement did not decrease the number of BFU-E colonies. The putative helper cells appear not to be T cells, were non-adherent to the plastic culture dish, and were cortisone resistant and radioresistant. The low BFU-E colony yield from SPF mouse marrow is presumed to be largely the result of deficiency of these non-T helper cells in SPF bone marrow, rather than of BFU-E progenitor cells.

Requirements for T cells in alloantigen-induced generation of non-T cell-mediated cytotoxicity against syngeneic mouse sarcoma cells.

In vitro stimulation of BALB/c spleen cells with allogeneic normal cells results in the generation of effector cells cytotoxic for syngeneic SV40 virus-induced MKSA fibrosarcoma cells. The conclusion that natural killer (NK) cells are activated that lyse the syngeneic tumor cells derives from findings that 1) treatment of the allostimulated BALB/c effector cells with anti-theta serum and complement does not ablate cytotoxicity against syngeneic MKSA tumor cells and 2) allostimulation of congenitally athymic (nu/nu) BALB/c mice results in significant cytotoxicity against MKSA tumor cells but not against allogeneic target cells. The requirement for the presence of theta-positive cells in alloantigen-induced activation of NK cells cytotoxic for syngeneic tumors was studied. Spleen cells from conventional BALB/c mice are cytotoxic for syngeneic MKSA tumor cells after stimulation with anti-theta serum-treated allogeneic cells or with allogeneic nude cells. In contrast, stimulation of BALB/c nude spleen cells with anti-theta serum-treated allogeneic cells or allogeneic nude cells fails to give rise to anti-tumor effector cells. Thus, theta-positive cells (presumably T cells) are required for alloantigen induced non-T cell-mediated lysis of syngeneic tumors.