X-irradiated Mice Research Articles

Detection of DNA Damage in Cells Exposed to Ionizing Radiation by Use of Anti-single-stranded DNA Monoclonal Antibody

An immunochemical method has been developed for quantitative detection of DNA damage in mammalian cells. The method is based on the binding of a monoclonal antibody to single-stranded DNA. The clone producing this antibody (D1B) was obtained as a by-product from fusion of mouse myeloma cells with spleen cells isolated from a mouse immunized with chemically modified DNA. The technique is based upon the determination of the percentage single-strandedness resulting from the time-dependent partial unwinding of cellular DNA under alkaline conditions. Single- and double-strand DNA breaks, or lesions converted into such breaks in alkaline medium, form initiation points for the unwinding. The extent of unwinding from these points under defined conditions is a measure of the number of such sites. The method is rapid, does not require radioactive labelling of DNA or physical separation of single-from double-stranded molecules, is sufficiently sensitive to detect damage induced by 1 Gy of ionizing radiation and needs only small numbers of cells. The usefulness of the technique was demonstrated in a study of the induction of DNA damage and its repair in cultured Chinese hamster cells and in human white blood cells after exposure to 60Co-gamma-rays, and in white blood cells and bone marrow cells of X-irradiated mice. A dose-related DNA unwinding was observed and repair of DNA lesions was observed up to 60 min after irradiation.

Protection of bone marrow progenitor cells by superoxide dismutase.

Intravenously administered cuprozinc-superoxide dismutase in X-irradiated mice hastens the recovery of peripheral blood cells. This effect is consistent with protection of the pluripotent stem cells by the enzyme. Amongst the bone marrow cells committed to differentiation along the myeloid pathway, there exists in mice a subpopulation of macrophage progenitor cells that is inactivated by superoxide radicals, generated photochemically or by X-rays. This cell killing effect is inhibited by superoxide dismutase, in part because it acts intracellularly. Human bone marrow also contain a superoxide-sensitive subpopulation of myeloid progenitor cells that is protected by superoxide dismutase but not by catalase. As well, human myeloid progenitor cells contain a subpopulation with enhanced sensitivity to X-rays in vitro. Treatment of these cells with exogenous superoxide dismutase reduces the sensitivity to X-rays by a factor of 2.

Effects of Trichinella spiralis Infections upon Bone Marrow Stem Cells in BALB/c Mice

Trichinella spiralis infections provoke a variety of responses in the host, some of which involve stem cell proliferation and myeloid cell maturation, increases in the mast cell precursor cell populations, and maturation and eosinopoiesis. Very little is known about the influence of T. spiralis upon bone marrow stem cells and splenic colony formation. In the present communication we report that T. spiralis infection in mice stimulates the generation of colony-forming units in the spleen (CFU-S). Passive transfer of bone marrow cells from uninfected BALB/c mice to X-irradiated (650 R) T. spiralis-infected recipients resulted in a significant increase of CFU-S at 14 and 24 days postinfection. Passive transfer of bone marrow cells from T. spiralis-infected mice to X-irradiated uninfected mice also resulted in increased numbers of CFU-S in the donor mice at 24 days postinfection. These findings strongly suggest that T. spiralis infection conditions the microenvironment in the spleen which stimulates CFU-S.

Self-reactive delayed type hypersensitivity induced in mice by syngeneic lymphoblasts. III. Immunological characterization of the small and large antigens of the blast cells.

X-irradiated or normal A mice injected with syngeneic concanavalin A-induced lymphoblasts (syn-Con A blasts) developed inflammatory responses in their footpads 24 to 72 h after the injection of syngeneic lipopolysaccharide-induced lymphoblasts (syn-LPS blasts) into these tissues. This response was designated syngeneic delayed type hypersensitivity (syn-DTH). The Con A blast extracts contain small (apparent MW of 6000-7000) and large (apparent MW of 160,000-175,000) syn-DTH-stimulating antigens, which are found in the total volume (low molecular weight fraction) and the void volume (high molecular weight fraction), respectively, of AcA 44 gel filtrations of this extract. The small and large antigens exhibit different immunological properties. The small antigen of A mouse lymphoblasts induced syn-DTH in X-irradiated (250 rad) mice but not in normal mice, and this immunological activity was elicited with syngeneic but not allogeneic lymphoblasts. The syn-DTH induced with the small antigen was inhibited by Lyt-1+2+, I-Jk+ suppressor T cells or a factor extracted from these cells. In contrast to the small antigen, the large antigen of A mouse lymphoblasts induced syn-DTH in both normal and X-irradiated mice, and this immunological activity was elicited by both syngeneic and allogeneic LPS lymphoblasts. The small and large antigens do not immunologically cross-react, but their immunogenicity is not affected by ultraviolet irradiation, indicating that the immune response against both of them is relatively class II-independent. The possibility that the cellular autoanti-lymphoblast response observed in our studies is in fact a mechanism that down-regulates the lymphoblast activity and thus suppresses the immune response is discussed.

Tumourigenesis by partial body X-irradiation in mice.

The aim of the present experiment was to investigate the late effects of partial body X-irradiation on mice. A total of 428 ddY/SLC female mice 10 weeks old were assigned to the following four groups: (1) head exposure with 950 rad, (2) trunk exposure with 950 rad, (3) lower body exposure with 950 rad, (4) unirradiated control. Mean after survival times with their standard errors were as follows: 430±13 days for head exposure, 354±8 days for trunk exposure, 435±13 days for lower body exposure, 472±14 days for control. Life shortening was 9 percent by head exposure and 25 percent by trunk exposure. Irradiation to the lower body did not induce statistically significant life shortening. Head exposure with 950 rad induced pituitary tumours. Trunk exposure with 950 rad induced ovarian tumours and reduced malignant lymphomas. The reduction may be attributable to the extensive life shortening after trunk exposure. An increase of lung tumours after trunk exposure was not statistically significant. Lower body exposure with 950 rad did not change the tumour spectrum of the control group.

The role of new vessels and macrophages in the development and resolution of edema following a cortical freeze lesion in the mouse.

The role of an influx of macrophages and neovascularity in the resolution of vasogenic edema is not well defined. The inhibition of these processes with x-irradiation or parenteral corticosteroid administration was used to evaluate their contribution to the resolution of edema around a cortical freeze lesion in mice. The resorption of Evans blue, a marker of protein extravasation, was delayed in x-irradiated mice on the second day following a freeze lesion (p = 0.0075), which correlates with a delay in macrophage infiltration around the lesion. The specific gravity of the lesion and its border regions was significantly less in x-irradiated animals on day 7 than in controls (p = 0.00062), which correlates with a delay in new vessel formation around the lesion. Administration of corticosteroids from the time of production of the freeze lesion resulted in a specific gravity significantly less than control when measured eight days after the lesion (p = 0.01). Macrophages may participate by inhibiting the development of the macromolecular portion of vasogenic edema. The development of neovascularity correlates with the resorption of the aqueous portion of vasogenic edema. As with x-irradiation, corticosteroids administered from the time of freeze lesion inhibited the resorption of the aqueous portion of vasogenic edema, but they may suppress the spread of edema in this experimental model.

Suppression of delayed type hypersensitivity to herpes simplex virus type 1 following immunization with anti-idiotypic antibody: an example of split tolerance.

Intraperitoneal (i.p.) immunization with herpes simplex virus type 1 (HSV-1) or an anti-idiotypic antibody (anti-id C) prepared against a monoclonal antibody specific for glycoprotein C of HSV-1, tolerizes mice for an HSV-1 delayed type hypersensitivity (DTH) response. This tolerization could be adoptively transferred to naive X-irradiated mice by splenic T cells, and was specific for HSV-1 DTH. Thus, DTH-tolerized mice responded to vaccinia virus or HSV-2 challenge, while remaining tolerized for HSV-1 DTH. In addition, these animals demonstrated a form of split tolerance such that HSV antibody, cytotoxic T cell and lymphoproliferative responses were detected in vitro. Thus, anti-id C induced a T cell population capable of specifically suppressing the HSV-1 DTH response, mimicking the effect of i.p. and intravenous immunization with HSV-1 found previously.

Susceptibility of normal and X-irradiated animals toAeromonas hydrophila infections

The virulence of threeAeromonas hydrophila strains isolated from humans and water were compared by intramuscular inoculation of normal and x-irradiated mice and goldfish. Selected strains, irrespective of source, were pathogenic for mice and goldfish with maximum mortality of normal hosts observed at 24–48 h after injection. Immunosuppression resulted in increased mortality of hosts by two of the threeA. hydrophila strains. The variation in virulence potential of differentA. hydrophila isolates may depend upon the in vivo interaction of endotoxins, surface antigens, and extracellular bacterial products.

Studies on the Mechanism of Systemic Suppression of Contact Hypersensitivity by UVB Radiation. II. Differences in the Suppression of Delayed and Contact Hypersensitivity in Mice

Exposing mice to UV radiation in the UVB range (280-320 nm) causes a selective immune suppression that contributes to the development of UVB-induced skin cancers. Among the immune responses suppressed by UVB irradiation are contact and delayed hypersensitivity reactions to haptens administered at unexposed sites. In these studies we provide evidence that delayed and contact hypersensitivity to the same hapten are not equivalent reactions and that they are suppressed in UVB-irradiated mice by 2 different mechanisms. This conclusion is based on the findings that: suppression of contact hypersensitivity could not be overcome by immunizing UVB-irradiated mice with hapten-coupled antigen-presenting cells derived from normal donors; and treatment of UVB-irradiated mice with methylprednisolone before immunization prevented the suppression of delayed hypersensitivity but had no effect on the suppression of contact hypersensitivity. The decreased ability to induce contact hypersensitivity in UVB-irradiated mice could be transferred to x-irradiated mice by reconstituting them with spleen cells from UVB-irradiated donors. The induction of hapten-specific suppressor cells, however, required both UVB irradiation and priming with hapten. Based on these results, we postulate that UVB irradiation induces a population of suppressor-inducer cells with specificity for a modified skin antigen and that this antigen serves as a carrier molecule for haptens that induce contact hypersensitivity and for tumor-specific transplantation antigens on UVB-induced tumors.

Immunologic Aspects of Acute Cutaneous Graft-Versus-Host Disease: Decreased Density and Antigen-Presenting Function of Ia+ Langerhans Cells and Absent Antigen-Presenting Capacity of Ia+ Keratinocytes

Cutaneous graft-versus-host disease (GVHD) provides a unique model for studying the pathogenesis of several important lymphocyte-mediated skin diseases. Morphologic studies have suggested that Ia antigen (Ia)-bearing epidermal Langerhans cells (LC) may be specific targets for destruction in these conditions. Keratinocytes synthesize and express Ia in GVHD and some other lymphocyte-mediated skin disorders; Ia+ keratinocytes, constitutively able to secrete epidermal cell-derived thymocyte activating factor (ETAF)/interleukin 1, may possess antigen-presenting capacity, thus leading to enhanced cutaneous immune responses and disease chronicity. We therefore investigated the fate of Ia+ LC, and the potential antigen-presenting capacity of Ia+ keratinocytes, in a murine model of GVHD. Lethally irradiated C3H/He (H-2k) mice developed acute cutaneous GVHD, and expressed keratinocyte Iak, 8 days after injection of BALB/c (H-2d) bone marrow and spleen cells. Immunofluorescence studies showed a progressive decrease in the density of Ia+ epidermal LC during the evolution of GVHD. This decrease was paralleled by a progressive reduction in the allostimulatory capacity of GVHD epidermal cells (EC) in the allogeneic EC-lymphocyte reaction (ELR). The fall in the density of Ia+ LC, and in EC allostimulatory capacity in both primary and secondary ELRs, was consistently greater in GVHD mice than in mice treated only with x-irradiation. The allostimulatory capacity of GVHD and x-irradiated EC could not be restored by addition of indomethacin or exogenous ETAF to ELR cultures. The decreased allostimulatory capacity was not the result of inhibition of the ELR, since EC from GVHD and x-irradiated mice did not cause suppression when added to control ELR cultures. The capacity of EC to present ovalbumin, purified protein derivative of tuberculin, 2,4,6-trinitrobenzenesulfonic acid coupled to EC, and native cytochrome c (CYTc) to antigen-specific T-cell lines, clones, or hybridomas was reduced in x-irradiated mice and markedly decreased in GVHD mice. The capacity of EC from x-irradiated and GVHD mice to present CYTc fragment 81-104, which does not require further processing or catabolism by accessory cells, was similarly decreased. Taken together, the results indicate that: the function of LC is markedly and progressively impaired in acute GVHD; LC function is also decreased, but to a lesser extent, following x-irradiation alone; and Ia+ keratinocytes from lethally irradiated mice undergoing GVHD do not exhibit antigen-presenting capacity.

Suppression of the anti-hapten IgE antibody response with hapten-modified spleen cells.

Spleen cells of BALB/c mice were chemically modified with phosphorylcholine or benzylpenicilloyl hapten. The i.v. administration of such cells into syngeneic animals suppressed the formation of specific IgE antibodies against the respective hapten. The IgE antibody response against ovalbumin, which was used as an immunogenic carrier for the haptens, was not affected and the anti-hapten IgG or IgG1 response remained at the levels of the controls. The suppression could be transferred into X-irradiated mice by T cells from tolerized animals. Moreover, it was demonstrated that not only the induction of IgE, but also an established anti-hapten IgE antibody response is accessible to suppression by treatment with hapten-modified spleen cells from syngeneic animals. The results indicate that the i.v. administration of antigen coupled to syngeneic spleen cells induces T cells which suppress the formation of specific IgE antibodies in the primary and the secondary response without significantly affecting the formation of IgG antibodies.

Effect of X-irradiation on Epidermal Immune Function: Decreased Density and Alloantigen-Presenting Capacity of Ia+ Langerhans Cells and Impaired Production of Epidermal Cell-Derived Thymocyte Activating Factor (ETAF)

The mechanisms involved in the modulation of cutaneous immune responses by UV radiation have been extensively investigated; by contrast, few studies have addressed the effects of x-irradiation on epidermal immune function. We therefore investigated the effect of x-irradiation of mice on: (a) the density of epidermal Ia+ Langerhans cells (LC) in immunofluorescence studies, (b) epidermal cell (EC) allostimulatory capacity in the allogeneic EC-lymphocyte reaction (ELR), and (c) production of epidermal cell-derived thymocyte activating factor (ETAF). C3H/He and BALB/c mice were irradiated with 900, 1,800, 2,700, or 3,600 rad from a 137Cs source, and sacrificed 10 h or 3 days later. X-irradiation of mice 10 h previously only slightly decreased the density of epidermal Ia+ LC and did not affect the capacity of their EC to stimulate allogeneic responder lymphocytes in the ELR. X-irradiation of mice 3 days previously, however, resulted in a dose-dependent decrease in the density of Ia+ LC. This decrease was accompanied by a substantial reduction in EC allostimulatory capacity in the ELR at all doses of x-irradiation. ETAF production by cultured EC from mice x-irradiated 3 days previously was also found to be diminished at all doses of x-irradiation. Trypan blue exclusion studies demonstrated that the observed decreases in EC allostimulatory capacity and ETAF production were not the result of a generalized lethal effect of x-irradiation on EC. The reduction in EC allostimulatory capacity following in vivo x-irradiation could not be reversed by addition of exogenous ETAF or interleukin-1 in the ELR. Taken together, these results indicate that x-irradiation decreases the density of Ia+ LC, impairs LC alloantigen-presenting function, and reduces ETAF production. Thus cutaneous x-irradiation may affect inflammatory and neoplastic processes not only by its antimitotic activity, but also by a direct effect on EC which subserve immunologic functions.

The effect of cyclosporin on murine autoreactive delayed type hypersensitivity induced with syngeneic lymphoblasts

The effect of cyclosporin on an immunological autoreactive experimental model was analyzed. The experimental system consisted of X-irradiated A mice injected with syngeneic concanavalin A-induced lymphoblasts and footpad-challenged 7 days later with syngeneic lipopolysaccharide-induced lymphoblasts. 24–72 h after challenge, the footpads of these mice responded with significant swelling, accumulation of 125I-UdR or massive cellular infiltration revealed by histological examination. Since the immunological activity was transferred by Lyt-1+ T cells from the sensitized donors to naive recipients, we designated it ‘syngeneic delayed-type hypersensitivity’ (syn-DTH). This DTH was induced and elicited mostly by antigens of the syngeneic lymphoblasts and not by contaminants attached to them, indicating the immunological autoreactive nature of this system.Multiple doses of 60 mg/kg cyclosporin, given daily in the time interval between immunization and challenge, or on the last four days before challenge, inhibited the syn-DTH. Multiple injections of cyclosporin before or close to the induction phase of the syn-DTH was ineffective, whereas single or multiple injections of cyclosporin close to the effector phase (the challenge time) markedly reduced the syn-DTH. Even a single injection of cyclosporin 24 h after the challenge efficiently reduced the 48-h syn-DTH.Adoptive transfer experiments revealed that T cells from X-irradiated mice immunized with concanavalin A-induced lymphoblasts and injected with cyclosporin, failed to efficiently transfer the syn-DTH response to naive recipients. Similarly, the syn-DTH response of naive X-irradiated recipient mice injected with cyclosporin, failed to be reconstituted with primed T cells derived from X-irradiated mice immunized with concanavalin A-induced lymphoblasts. Since the nonspecific footpad swelling response of X-irradiated mice challenged with lymphoblasts alone is resistant to the standard protocol of the cyclosporin treatment, we suggest that cyclosporin inhibits the ability of T cells to produce or release lymphokines at the effector phase of DTH, while phagocytic cells involved in the DTH response are not affected by it. The practical and the theoretical implications of this research are discussed.

Influence of added catalase on chromosome stability and neoplastic transformation of mouse cells in culture

The generation of hydrogen peroxide (H2O2) and the derivative free hydroxyl radical (. OH) in cultures of mouse cells grown in the presence of visible light and ambient oxygen was shown previously to be implicated in chromatid damage. Furthermore, chromosome alterations appear to be associated with the spontaneous neoplastic transformation of mouse cells in culture. An attempt was made in this study to reduce the incidence of chromosomal aberrations and delay or prevent the onset of spontaneous neoplastic transformation of freshly isolated mouse cells, both fibroblasts and epidermal keratinocytes, by adding catalase to the culture medium, shielding the cultures from wavelengths less than 500 nm and providing a gas phase of 0-1% O2. These conditions significantly decreased the incidence of chromosomal aberrations in both cell types, and in fibroblasts prevented tumourigenicity in non-irradiated syngeneic mice, and increased latent periods for tumour development in X-irradiated mice. The epidermal keratinocytes were particularly resistant to spontaneous neoplastic transformation under all conditions tested. These observations on the protective effect of extracellular catalase suggest that H2O2, a normal metabolite, and/or the derivative .OH can directly or indirectly produce genetic damage and neoplastic transformation in mouse fibroblasts.

Visceral leishmaniasis in congenic mice of susceptible and resistant phenotypes: T-lymphocyte-mediated immunosuppression.

This paper continues a comparative study (A. D. Nickol and P. F. Bonventre, Infect. Immun. 50:160-168, 1985) describing immune responses exhibited by congenic, Lshs mouse strains C57B1/10 (cure) and B10.D2 (noncure) during the course of disseminated leishmaniasis. We report that sublethal whole-body irradiation of B10.D2 mice before challenge with Leishmania donovani converted the noncuring mice to a curing phenotype. Splenic lymphocytes from L. donovani-infected B10.D2 mice failed to proliferate in response to parasite antigen stimulation in vitro. Splenic lymphocytes from irradiated, cured B10.D2 mice regained the capacity to respond to the parasite antigen stimulus. Transfer of T cells but not B cells from L. donovani-infected B10.D2 mice prevented the acquisition of immunity and recovery from infection in X-irradiated mice. In addition, a splenic T-cell population from L. donovani-infected B10.D2 mice suppressed the proliferation in vitro of parasite antigen-stimulated lymphocytes of irradiation-cured B10.D2 mice. Suppressor T cells were not demonstrable in the spleens of spontaneously cured C57B1/10 mice. Splenic lymphocytes from infected B10.D2 mice were deficient in the production of macrophage-activating factor (MAF) upon stimulation by L. donovani antigens in vitro. Deficient MAF production was specific for parasite antigen stimulation, because MAF production subsequent to concanavalin A stimulation of splenic lymphocytes from infected B10.D2 mice was not suppressed. The data suggest that a genetically based immunological defect in B10.D2 mice prevents the acquisition of effective cell-mediated immunity and subsequent elimination of L. donovani from tissue macrophages. The immunological deficit, not apparent in the curing C57B1/10, appears to be caused by the development of parasite antigen-specific suppressor T cells during the course of the disseminated leishmaniasis.

Self Reactive Delayed Type Hypersensitivity (DTH) Induced in Mice by Syngeneic Lymphoblasts

Normal and x-irradiated A mice injected with syngeneic concanavalin A (Con A)-induced lymphoblasts revealed after challenge with syngeneic lipopolysaccharide (LPS)-induced lymphoblasts delayed type hypersensitivity (DTH) measured both by footpad swelling and by 125IudR accumulation. Mice injected with allogeneic Con A-induced lymphoblasts and challenged with syngeneic LPS-induced lymphoblasts or vice versa, also generated an appreciable DTH response. In contrast, Con A-induced blast cells of human origin (xenogeneic cells) generated a considerably less effective DTH. The DTH response was more profound and consistent in x-irradiated mice, suggesting that irradiation sensitive cells control this response. The syngeneic DTH response was efficiently transferred to naive recipients with Thy-1 +, nylon wool passed cells. The establishment of the DTH activity was associated with the lymphoblasts own (differentiation?) antigens and not with contaminants attached to the cells, such as Con A or fetal calf serum. The results were compared with similar results reported by other groups and the biological significance of all findings was evaluated.

Restoration of radiation injury by ginseng. IV. Stimulation of recoveries in CFUs and megakaryocyte counts related to the prevention of occult blood appearance in X-irradiated mice.

A single post-irradiation injection of the thermostable fraction of a ginseng extract enhanced the recoveries of the numbers of both blood-forming stem cells (10 days CFUs) and megakaryocytes in bone marrow of irradiated mice. It also protected hemorrhaging tendency of X-irradiated mice determined by a quantitative measurement of occult blood appearance in daily feces. The present study and our previous findings suggest that ginseng protected the irradiated animals from bone marrow death by stimulating thrombopoietic hematogenesis (CFUs, megakaryocytes and thrombocytes) and finally preventing hemorrhage.

Whole-body X-irradiation of mice accelerates polyploidization of hepatocytes.

Male C57BL/6 mice were whole-body irradiated with 4.75 Gy of X-rays at the age of 2 months and killed at 2, 6, 12 and 19 months after irradiation. The percentage survival began to decline earlier and faster in the irradiated group than the controls up to 19 months after exposure when the study was terminated. The nuclear DNA content of individual hepatocytes was measured by a Feulgen-DNA microfluorometric method, and hepatocytes were classified into various ploidy classes. In the irradiated mice, the degree of polyploidization was significantly higher than the controls by 2 months after exposure and steadily increased up to 6 months after exposure. Thereafter, however, a slow return to the control level was found up to 19 months after irradiation. These results appear to support a hypothesis that radiation accelerates the ageing process as judged from hepatocyte polyploidization.

The Effect of a Sunscreen Containing Para-aminobenzoic Acid on the Systemic Immunologic Alterations Induced in Mice by Exposure to UVB Radiation

Application of a sunscreen containing para-aminobenzoic acid partially abrogated certain of the systemic immunologic alterations produced in mice by exposure to UVB (280-320 nm) radiation from sunlamp bulbs. The sunscreen reduced the degree of UVB-induced suppression of contact hypersensitivity to a chemical applied subsequently to nonirradiated skin. In addition, it reduced the frequency with which mice became susceptible to the growth of a highly antigenic, syngeneic, UVB-induced tumor following chronic treatment with UVB radiation, but this effect was not statistically significant. The tumor-susceptible state was transferred from animals treated with sunscreen and UVB radiation to lethally x-irradiated mice by injection of spleen cells. The gross morphology of the skin of mice treated with sunscreen and UVB radiation was normal but the histologic changes induced by UVB irradiation in skin were only partially abrogated by the sunscreen.

Auto-delayed-type hypersensitivity induced in immunodeficient mice with modified self-antigens. III. Suppressive T-cell factor controls the autoreactivity against self-antigens.

X-irradiated (250 rad), cyclophosphamide-treated or ATx A mice injected with syngeneic trinitrophenylated spleen cells (TNP-SC) and footpad challenged with syngeneic lymphoblasts generated delayed-type hypersensitivity (DTH) responses 24, 48 and 72 h after challenge. The syngeneic-DTH (syn-DTH) response was mediated by Lyt-1+ cells and suppressed with Lyt-1+2+3+, I-Jk+ cells. The suppressor cells were obtained from spleens or thymuses of normal syngeneic mice. Suppressor factor (SF) was extracted or released from Lyt-1+2+3+, I-Jk+ cells obtained from normal A mice (but not from X-irradiated A mice). The factor blocked the DTH responses of X-irradiated mice injected with syngeneic TNP-SC and challenged with syngeneic lymphoblasts when injected into the mice both at the induction phase and the elicitation phase of the DTH. The factor failed to abrogate allogeneic and xenogeneic DTH. However, allogeneic factor (derived from C57BL/6 mice) abolished the syn-DTH response of mice injected with syngeneic TNP-SC and challenged with syngeneic lymphoblasts. The SF was produced by Lyt-1+2+3+, I-Jk+ T cells or by thymocytes. The combined extracted product of Lyt-1+ and Lyt-2+ cells did not abrogate the syn-DTH response. Normal spleen cells depleted of phagocytes by a magnetic procedure also produced the SF. These findings indicate, therefore, that suppressive factor (or factors; see Discussion in the accompanying paper, Ref. 17) controls the immunological autoreactivity against syngeneic TNP-SC.