concanavalin-a and Graft-vs-Host-Disease

This review addresses the issue of histone deacetylase (HDAC) inhibitors as developed for the treatment of cancer and for the investigation of the inhibition of inflammation. The review focuses on both in vitro and in vivo models of inflammation and autoimmunity. Of particular interest is the inhibition of pro-inflammatory cytokines. Although the reduction in cytokines appears paradoxical at first, upon examination, some genes that are anti-inflammatory are upregulated by inhibition of HDAC. Whether skin diseases will be affected by inhibitors of HDAC remains to be tested.

Topics: Animals; Anti-Inflammatory Agents; Antineoplastic Agents; Concanavalin A; Cytokines; Disease Models, Animal; Enzyme Inhibitors; Genes, Viral; Graft vs Host Disease; Hepatitis; Histone Deacetylase Inhibitors; Histone Deacetylases; Humans; Inflammation; Interleukin-1; Lipopolysaccharides

Phytohemagglutinin was prominent in the evaluation of the immunosuppressive effects of mitogenic lectins that started in 1965 and continued for two decades. Basic studies elucidated the inhibitory actions of PHA on humoral and cellular immune responses in mice, rats, and guinea pigs. Some suppressive effects of this and other mitogens including lentil lectin and Con A were demonstrated on renal, skin, pancreas, and heart allograft rejections in mice, rats, and dogs. In addition to their inherent suppressive activities, these substances have been shown to potently augment the suppression generated by conventional agents. More relevant to tolerance-inducing modulations, the Rigas group showed that in vitro incubation of the parental spleen cells with PHA in the F1 hybrid model before giving them to the F1 mouse virtually abolished the GvH responses. Their explanation was that the donor cells were rendered unresponsive by their reversion to an immature state in which they lost the essential surface receptors. Similar spleen cell suppression was generated by systemic administration of PHA to the parental donor prior to their administration. A method is proposed here for establishing tolerance to either newly introduced or firmly established antigens applying the L4 isolectin of PHA to make non-reactive all T lymphocytes that remain functional in conjunction with full suppression by conventional agents. During the vulnerable period of drug-induced suppression, the host would be protected from infection and bleeding by full nonspecific proliferative activation of the lymphoid and myeloid systems. The establishment of allograft tolerance by preemptive inhibition of responses to newly introduced transplant antigens would be easier to achieve than the reversal of firmly established responses to antigens involved in GvH reactions and autoimmune diseases. The studies of von Boehmer and Kisielow in a transgenic mouse model confirmed that clonal deletion does occur in the thymus whereby corresponding specific thymocytes are destroyed when they encounter self-antigens. Their work suggested that tolerance to self-antigens might be established if immune responses against putative antigen peptides were blocked sufficiently that they would be released to reach the central or peripheral residence sites of their immunologically reactive clones for deletion to occur. This approach to suppressive therapy would be useful for managing the problems of allograft rejection, GvH rea

Topics: Animals; Autoimmune Diseases; Concanavalin A; Dogs; Graft Rejection; Graft vs Host Disease; Humans; Immunosuppressive Agents; Lectins; Mice; Phytohemagglutinins; Rats; Transplantation Immunology

Topics: CD3 Complex; Cell Proliferation; CHARGE Syndrome; Concanavalin A; Cyclosporine; Diarrhea; DiGeorge Syndrome; Epstein-Barr Virus Infections; Fatal Outcome; Graft vs Host Disease; Herpesvirus 4, Human; HLA Antigens; Humans; Infant, Newborn; Lymphocyte Transfusion; Male; Methotrexate; Mutation; Phenotype; Phytohemagglutinins; Siblings; T-Lymphocytes

NF-κB is a crucial mediator of inflammatory and immune responses and a number of phytochemicals that can suppress this immune-regulatory transcription factor are known to have promising anti-inflammatory potential. However, we report that inducer of pro-inflammatory transcription factor NF-κB functions as an anti-inflammatory agent. Our findings reveal that a plant derived flavonoid baicalein could suppress mitogen induced T cell activation, proliferation and cytokine secretion. Treatment of CD4+ T cells with baicalein prior to transfer in to lymphopenic allogenic host significantly suppressed graft versus host disease. Interestingly, addition of baicalein to murine splenic lymphocytes induced DNA binding of NF-κB but did not suppress Concanavalin A induced NF-κB. Since baicalein did not inhibit NF-κB binding to DNA, we hypothesized that baicalein may be suppressing NF-κB trans-activation. Thioredoxin system is implicated in the regulation of NF-κB trans-activation potential and therefore inhibition of thioredoxin system may be responsible for suppression of NF-κB dependent genes. Baicalein not only inhibited TrxR activity in cell free system but also suppressed mitogen induced thioredoxin activity in the nuclear compartment of lymphocytes. Similar to baicalein, pharmacological inhibitors of thioredoxin system also could suppress mitogen induced T cell proliferation without inhibiting DNA binding of NF-κB. Further, activation of cellular thioredoxin system by the use of pharmacological activator or over-expression of thioredoxin could abrogate the anti-inflammatory action of baicalein. We propose a novel strategy using baicalein to limit NF-κB dependent inflammatory responses via inhibition of thioredoxin system.

Topics: Animals; Anti-Inflammatory Agents; Cell Line, Tumor; Cell Proliferation; Concanavalin A; Cytokines; DNA; Flavanones; Graft vs Host Disease; Lymphopenia; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mitogens; NF-kappa B; Protein Binding; Spleen; T-Lymphocytes; Thioredoxins; Transcriptional Activation

Ursolic acid (UA), a pentacyclic triterpenoid carboxylic acid, is the major component of many plants including apples, basil, cranberries, peppermint, rosemary, oregano and prunes and has been reported to possess antioxidant and anti-tumor properties. These properties of UA have been attributed to its ability to suppress NF-κB (nuclear factor kappa B) activation. Since NF-κB, in co-ordination with NF-AT (nuclear factor of activated T cells) and AP-1(activator protein-1), is known to regulate inflammatory genes, we hypothesized that UA might exhibit potent anti-inflammatory effects.. The anti-inflammatory effects of UA were assessed in activated T cells, B cells and macrophages. Effects of UA on ERK, JNK, NF-κB, AP-1 and NF-AT were studied to elucidate its mechanism of action. In vivo efficacy of UA was studied using mouse model of graft-versus-host disease. UA inhibited activation, proliferation and cytokine secretion in T cells, B cells and macrophages. UA inhibited mitogen-induced up-regulation of activation markers and co-stimulatory molecules in T and B cells. It inhibited mitogen-induced phosphorylation of ERK and JNK and suppressed the activation of immunoregulatory transcription factors NF-κB, NF-AT and AP-1 in lymphocytes. Treatment of cells with UA prior to allogenic transplantation significantly delayed induction of acute graft-versus-host disease in mice and also significantly reduced the serum levels of pro-inflammatory cytokines IL-6 and IFN-γ. UA treatment inhibited T cell activation even when added post-mitogenic stimulation demonstrating its therapeutic utility as an anti-inflammatory agent.. The present study describes the detailed mechanism of anti-inflammatory activity of UA. Further, UA may find application in the treatment of inflammatory disorders.

Topics: Animals; Anti-Inflammatory Agents; Antibodies, Monoclonal; Antioxidants; B-Lymphocytes; CD28 Antigens; CD3 Complex; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Proliferation; Concanavalin A; Cytokines; Graft vs Host Disease; Lymphocyte Activation; Macrophages; Mice; Mitogen-Activated Protein Kinases; Mitogens; NF-kappa B; NFATC Transcription Factors; Oxidation-Reduction; Reactive Oxygen Species; Transcription Factor AP-1; Triterpenes; Up-Regulation; Ursolic Acid

Rhodiola quadrifida (Rq) roots and rhizomes are traditionally used in Asia as a tonic, adaptogen, antidepressant and anti-inflammatory drug. The aim of this work was to study the in vivo effect of aqueous and 50% hydro-alcoholic extracts of Rq rhizomes on some parameters of cellular immunity in mice and rats. The metabolic activity of blood phagocyting cells was determined based on the measurement of intracellular respiratory burst after stimulation by PMA in RBA test. Potential bactericidal activity of phagocyting cells was determined in isolated blood leukocytes stimulated with killed microorganisms, according to the PKA test. Proliferative response of lymphocytes stimulated by mitogen concanavaline A (ConA) or lipopolysaccharide (LPS) were determined by MTT assay. Both extracts stimulated granulocytes activity in vitro and increased lymphocyte response to mitogens. The ability of parental strain mice lymphocytes to induce local cutaneous graft-versus-host reaction (GVH) in F1 hybrids was stimulated by 50% hydro-alcoholic extract only.

Topics: Animals; Concanavalin A; Cytotoxicity, Immunologic; Dose-Response Relationship, Drug; Female; Graft vs Host Disease; Graft vs Host Reaction; Immunity, Cellular; Lipopolysaccharides; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred DBA; Phagocytosis; Plant Extracts; Plant Roots; Random Allocation; Rats; Rats, Inbred Lew; Respiratory Burst; Rhodiola

Acute, lethal graft-versus-host disease (GvHD) develops in B6D2F1 hybrid recipients of wild-type, C57BL/6, parental strain grafts; however, when interferon-gamma (IFN-gamma) gene knockout (gko) donors are used, the disease is prolonged and associated with a higher level of engraftment, particularly of T cells. Lesions containing large, mixed cellular infiltrates develop in the skin, liver, pancreas, salivary gland, lung and kidney. In our current study, we wished to determine whether GvHD features a preponderance of T helper 2 (Th2) cytokines in the absence of donor-derived IFN-gamma, and whether autoantibody production, commonly associated with chronic GvHD, also occurs. Because mitogen responsiveness is consistently suppressed in mice with acute GvHD, we wished to measure this response in recipients of IFN-gamma gko grafts. Our findings indicate that spleen cells from the latter produce interleukin (IL)-4, IL-5 and IL-13 in culture, but respond poorly to concanavalin A (Con A) and lipopolysaccharide (LPS). Their sera contain anti-nuclear antibodies (ANA), some of which are specific for double-stranded (ds)DNA and are predominantly immunoglobulin (Ig)M and IgG1. We also noted the presence of numerous eosinophils in the infiltrates developing within the target organs. In some respects, this syndrome bears resemblance to both systemic lupus erythematosus (SLE) and chronic GvHD. However, histological evidence of glomerulonephritis is lacking and proteinuria fails to develop in recipients of IFN-gamma gko grafts, suggesting that IFN-gamma may be necessary for the development of lupus nephritis. On a broader scope, our findings underscore the importance of IFN-gamma in the pathogenetic mechanism of GvHD, and demonstrate that the absence of this cytokine promotes the development of chronic GvHD and autoimmunity.

Topics: Animals; Antibodies, Antinuclear; Concanavalin A; Cytokines; Eosinophilia; Female; Graft vs Host Disease; Immunoglobulin G; Immunoglobulin M; Interferon-gamma; Interleukin-13; Interleukin-4; Interleukin-5; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Mice, Knockout; T-Lymphocytes, Helper-Inducer

Clinical application of cytotoxic T lymphocytes (CTL) induced in vitro is extensively used for the treatment of viral infection and malignant diseases. We produced anti H-2d CTL in vitro from C57BL/6 (B6) splenocytes presensitized with (B6 x DBA/2) F1 (BDF1) splenocytes to establish a model system of CTL therapy. The specificity and cytotoxic activity were high enough (E/T ratio 1:1 = 38.8%) to induce graft versus host reaction. Though the total number of B6 splenocytes decreased by 0.27 during the 4 days of culture, the number of CD8+ lymphocytes increased 1.3-fold. When more than 5 x 10(6) cells of H-2d-reactive CTL were transplanted into BDF1 mice, mice died within 2 days postinduction. This lethal effect was not seen in the mice induced with ConA-stimulated T cells. Histological examination of the lungs and liver revealed massive infiltration of neutrophils in alveoli and the necrosis of hepatocytes. Therefore, this protocol was shown to be effective to produce alloantigen-specific CTLs and applicable to in vitro manipulation such as retrovirus-mediated gene transfer.

Topics: Animals; Cell Transplantation; Cells, Cultured; Chromium; Concanavalin A; Female; Flow Cytometry; Graft vs Host Disease; Liver; Lung; Mice; Mice, Inbred Strains; Pulmonary Alveoli; Spleen; T-Lymphocytes, Cytotoxic; Transplantation, Isogeneic

We have previously reported that cluster of differentiation (CD)4+ T cells induced autoimmune liver diseases in mice with graft-versus-host reaction (GVHR) because of major histocompatibility complex (MHC) class II disparity. To analyze the progression of the autoimmune-related mechanism in the liver, concanavalin A (Con A) was injected in mice undergoing GVHR. The aim of this study is to clarify whether Con A deteriorates murine hepatic lesions induced by GVHR, and to elucidate the participation of the cytokines of liver-infiltrating CD4+ T cells.. Mice (F1; B6.C-H-2(bm12) x B6) were intravenously injected with B6 T spleen cells. Concanavalin A (15 mg/kg) was administrated 5 days after cell transfer. We examined serum transaminase, antimitochondrial antibodies (AMA), antinuclear antibodies (ANA) and histological changes. Liver-infiltrating CD4+ T cells were sorted and their cytokine mRNA expression was examined by the use of reverse transcription-polymerase chain reaction (RT-PCR).. Graft-versus-host reaction + Con A mice revealed an elevated serum transaminase, elevated AMA and ANA titers, increased periportal cellular infiltration, piecemeal necrosis and bridging necrosis in the liver. In this group, interferon (IFN)-gamma mRNA expression was more elevated than it was in the GVHR mice. However, there was no difference in the expression of interleukin (IL)-10 mRNA between the two groups.. The results suggest that Con A deteriorates the GVHR-induced hepatic lesions, and IFN-gamma and IL-10 of CD4+ T cells might be implicated in the progression of autoimmune-related hepatic lesions. This model might offer an aspect for the investigation of progressive mechanisms in T-cell- mediated hepatobiliary injury.

Topics: Analysis of Variance; Animals; Autoantibodies; Autoimmune Diseases; CD4-Positive T-Lymphocytes; Cell Transplantation; Concanavalin A; Cytokines; Disease Models, Animal; Graft vs Host Disease; Hepatitis; Liver Function Tests; Mice; Mice, Inbred C57BL; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Spleen

Despite a 10-fold increase of T cell dose, the incidence and severity of acute GVHD following allogeneic transplantation of G-CSF-mobilized PBSC is not increased compared to BMT. Experimental murine studies demonstrate that G-CSF polarizes donor T cells toward a type 2 cytokine response. To determine whether G-CSF alters T cell cytokine responses, we investigated the effects of G-CSF administration on T cell proliferative and cytokine responses to alloantigen and Con A in nonadherent PBMC (NAC) and CD3+ T cells obtained from normal individuals before and after G-CSF administration (10 microg/kg x 4 days). Although T cell proliferative and cytokine (IFN-gamma and IL-4) responses to alloantigen stimulation and Con A were significantly reduced in post-G-CSF NAC, they were restored by the removal of non-T cells from post-G-CSF NAC. Furthermore, there was less T cell alloreactivity in MLR in the presence of autologous post-G-CSF monocytes than in the presence of pre-G-CSF monocytes. This alteration was not replicated in vitro by culturing PBMC with G-CSF. These results suggest that G-CSF administration suppresses T cell proliferative and cytokine (IFN-gamma and IL-4) responses to allogeneic stimulation by indirectly modulating monocyte function. Bone Marrow Transplantation (2000).

Topics: Adult; Cells, Cultured; Coculture Techniques; Concanavalin A; Depression, Chemical; Female; Gene Expression Regulation; Graft vs Host Disease; Granulocyte Colony-Stimulating Factor; Hematopoietic Stem Cell Transplantation; Humans; Interferon-gamma; Interleukin-4; Isoantigens; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Lymphocyte Depletion; Male; Monocytes; Monokines; Recombinant Proteins; T-Lymphocyte Subsets; Transplantation, Homologous

To investigate the clinical applicability of prophylaxis of post-transplant graft-versus-host disease by UV-B irradiation of stem cell preparations, the UV-B sensitivities of human lymphocytes and primitive hematopoietic progenitors were compared. The mononuclear cell fractions (MNC) derived from human cord blood and granulocyte-colony-stimulating factor-mobilized peripheral blood were used. After UV-B irradiation, lymphocyte proliferation ability, hematopoietic colony-forming cells, and apoptotic cells were analyzed. At a dose of 33 J/m(2), significant differences were observed in the residual percentages of hematopoietic progenitors and lymphocyte functions [ANOVA, F (5,46) = 19.4; P <.0001], and the difference between CFU-C (85.2% + 24.0%; n = 8) and MLR (12.7% + 12. 6%; n = 10) was significant (P <.0001). There were no significant differences in the residual percentages of CFU-C, HPP-CFC, and LTC-IC. Percentages of annexin V-positive cells in the total MNC and the CD34(+) cell population in MNC after UV-B irradiation were 69.8% and 18.7%, respectively. In conclusion, there was a range of UV-B doses over which T lymphocytes were inactivated but hematopoietic progenitors, including HPP-CFC and LTC-IC, were preserved.

Topics: Annexin A5; Antigens, CD34; Apoptosis; Cell Membrane; Cells, Cultured; Colony-Forming Units Assay; Concanavalin A; Fetal Blood; Flow Cytometry; Graft vs Host Disease; Granulocyte Colony-Stimulating Factor; Hematopoietic Stem Cells; Humans; Lymphocyte Culture Test, Mixed; Lymphocytes; Ultraviolet Rays

Graft-versus-host disease (GVHD), in which immunocompetent donor cells attack the host, remains a major cause of morbidity after allogeneic bone marrow transplantation (BMT). To understand the role of cytokines in the pathobiology of GVHD, we used cytokine knockout (KO) mice as a source of donor T cells. Two different MHC-disparate strain combinations were examined: BALB/c (H2(d)) donors into lethally irradiated C57BL/6 (H2(b)) recipients or C57BL/6 (H2(b)) donors into B10.BR (H2(k)) recipients. Donor cells were from mice in which either the interferon-gamma (IFN-gamma) or the IL-4 gene was selectively disrupted to understand the role of these cytokines in acute GVHD. In both strain combinations the same pattern was noted with regard to GVHD onset and morbidity. All mice exhibited the classic signs of acute GVHD: weight loss with skin, gut, and liver pathology resulting in morbidity and mortality. Surprisingly, donor cells obtained from mice lacking IFN-gamma gave rise to accelerated morbidity from GVHD when compared with cells from wild-type control donors. Similar results were obtained using normal donors when neutralizing antibodies to IFN-gamma were administered immediately after the BMT. These results suggest that IFN-gamma plays a role in protection from acute GVHD. In marked contrast, cells obtained from IL-4 KO mice resulted in protection from GVHD compared with control donors. Splenocytes from IFN KO mice stimulated with a mitogen proliferated to a significantly greater extent and produced more IL-2 compared with splenocytes obtained from IL-4 KO or control mice. Additionally, there was increased IL-2 production in the spleens of mice undergoing GVHD using IFN-gamma KO donors. These results therefore indicate, with regard to the TH1/ TH2 cytokine paradigm, the absence of a TH1-type cytokine can be deleterious in acute GVHD, whereas absence of a TH2 cytokine can be protective.

Topics: Animals; Bone Marrow Transplantation; Cell Division; Concanavalin A; Graft vs Host Disease; Interferon-gamma; Interleukin-2; Interleukin-4; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mitogens; Spleen; Transplantation, Homologous

We previously used peripheral newborn blood (NBB) as a possible in vivo experimental model for cord blood (CB) transplantation and showed that B10.D2 NBB cells successfully reconstituted adult (DBA/2 x B10.D2)F1 mice without causing graft-versus-host disease (GVHD), probably because of their phenotypic and functional immaturity. Here we investigated the influence of T-cell maturation occurring in NBB cells during the early postbirth period on the degree of engraftment, the incidence of GVHD, and the graft-versus-leukemia (GVL) potential. These parameters were compared in recipients grafted with bone marrow (BM) cells. We observed an increased percentage of CD4(+) mature T cells accompanied by the acquisition of proliferative responses to phytohemagglutinin (PHA) and to allogeneic cells of day-5 NBB cells. The capacity of day-2 NBB to engraft was moderately reduced and recipients developing GVHD were occasionally observed after the graft of day-5 NBB cells. No GVL effect was evidenced regardless of the time of postbirth blood collection. However, the GVL effect can be obtained by the delayed infusion of donor mature T cells to recipients grafted with day-0 NBB, without causing GVHD. In contrast, the same protocol applied to mice grafted with BM cells induced GVHD mortality of all recipients. Interleukin (IL)-10 but not IL-2 messenger RNA was expressed in NBB cells as opposed to BM cells. These findings suggest that, in terms of GVHD incidence, delayed infusion of mature T cells as post-transplant tumor immunotherapy would be more effective when applied after CB than after BM transplantation.

Topics: Animals; Animals, Newborn; Bone Marrow Cells; Bone Marrow Transplantation; CD4-Positive T-Lymphocytes; Cell Differentiation; Concanavalin A; Fetal Blood; Graft Survival; Graft vs Host Disease; Graft vs Tumor Effect; Hematopoietic Stem Cell Transplantation; Hematopoietic Stem Cells; Humans; Interleukin-10; Interleukin-2; Lymphocyte Activation; Mice; Mice, Inbred DBA; Radiation Chimera; RNA, Messenger; Spleen; T-Lymphocytes

Using murine models, we have shown that the lysosomotropic amine, chloroquine, is effective in the prevention of graft-versus-host disease (GVHD) mediated by donor T cells reactive with recipient minor histocompatibility antigens (MiHCs). Because lysosomotropic amines can suppress major histocompatibility complex (MHC) class II antigen presentation, their mechanism of action is potentially different from current immune suppressant drugs used to control GVHD such as cyclosporine.. We investigated the use of cyclosporine and the lysosomotropic amines chloroquine and hydroxychloroquine in combination for additive or synergistic immunosuppression on T-cell responses in vitro to MiHC and MHC in mice.. We found that similar concentrations of chloroquine and hydroxychloroquine suppress the T-cell response to MiHC in mice (C57BL/6 anti-BALB.B) and that lysosomotropic amines in combination with cyclosporine result in synergistic suppression of a proliferative response to MiHC. Similar suppression and synergy appear to be present in an alloreactive response (C57BL/6 anti-BALB/c). Direct inhibition by chloroquine of T-cell proliferative responses induced by anti-CD3epsilon in the absence of antigen-presenting cells is present at higher concentrations than that required to suppress responses to MiHC or MHC. Chloroquine appears to induce decreased T-cell viability at high concentrations. This effect does not appear to be due to decreased T-cell production of interleukin-2 or interferon-gamma. At lower concentrations (<25 microg/ml), chloroquine can also decrease the ability of antigen-presenting cells to stimulate an a C57BL/6 anti-BALB/c T-cell response and can inhibit MHC class II expression after activation with lipopolysaccharide.. Lysosomotropic amines in combination with cyclosporine appear to be synergistic in the suppression of T-cell proliferation to MiHC and MHC. Use of chloroquine in combination with cyclosporine may result in improved control of GVHD.

Topics: Animals; Cell Survival; Cells, Cultured; Chloroquine; Concanavalin A; Cyclosporine; Drug Synergism; Female; Graft vs Host Disease; Histocompatibility Antigens Class II; Hydroxychloroquine; Interferon-gamma; Interleukin-2; Kinetics; Lymphocyte Activation; Lysosomes; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Minor Histocompatibility Antigens; Spleen; T-Lymphocytes

The development of graft-versus-host disease (GVHD) is associated with long-lasting and profound deficits in immune function that lead to increased morbidity and mortality after bone marrow transplantation (BMT). We investigated a mechanism of T-cell immunodeficiency in response to mitogen or alloantigen in an experimental model of acute GVHD by analyzing the roles of two immunosuppressive moieties: interferon gamma (IFN-gamma) and nitric oxide (NO). Splenocytes from mice with GVHD did not proliferate either to the T-cell mitogen, concanavalin A (Con A), or to host alloantigens, but only mitogen-activated cultures produced increased levels of NO. The abrogation of NO synthesis with LG-mono-methyl-arginine (NMMA) restored mitogen-induced proliferation but not the response to host antigens. The mechanism of impared proliferation to mitogen was dependent on IFN-gamma because blockade of this cytokine in culture inhibited NO production and restored proliferation to Con A to levels similar to those in transplanted control mice without GVHD. NMMA did not substantially reduce IFN-gamma levels, demonstrating that NO acted distally to IFN-gamma in the pathway of immunosuppression in response to mitogen. Furthermore, the prevention of IFN-gamma production in vivo after allogeneic BMT, by transplantation of polarized type 2 donor T cells (secreting interleukin-4 but not IFN-gamma), also prevented NO production and restored splenocyte responses to mitogen. Our data demonstrate the existence of NO-dependent and NO-independent pathways involved in suppression of T-cell proliferation during acute GVHD. Excess NO synthesis appears to be one mechanism by which IFN-gamma induces immunodeficiency after allogeneic BMT.

Topics: Acute Disease; Animals; Arginine; Bone Marrow Transplantation; Concanavalin A; Enzyme Activation; Enzyme Inhibitors; Female; Graft vs Host Disease; Interferon-gamma; Interleukin-4; Isoantigens; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mitogens; Nitric Oxide; Nitric Oxide Synthase; omega-N-Methylarginine; Radiation Chimera; Spleen; T-Lymphocytes

Transplantation of allogeneic bone marrow (BM) is often complicated by the development of acute graft-vs-host disease (GVHD) caused by contaminating T cells in BM inocula because of activation of the host reactive T effector cells. Using a murine model for acute GVHD caused by injection of parental C57Bl/6 splenocytes into unirradiated (C57Bl/6 x DBA/2) F1 hybrids, we demonstrated that pretreatment of the inocula with a novel immunosuppressant, B subunit of cholera toxin (CT-B) impaired the ability of C57Bl/6 T cells to induce acute GVHD in F1 recipients. F1 mice injected with CT-B-treated C57Bl/6 splenocytes did not develop significant splenomegaly, and no antihost CTLs were found in their spleens. Moreover, these mice did not suffer from aplastic anemia, nor from general immunosuppression. Immunofluorescence studies suggest that treatment of the inducing inocula with CT-B selectively prevents accumulation of the host-reactive CD8+ T cells in F1 mice. Furthermore, our experiments demonstrated that CT-B treatment does not impair the ability of BM progenitors to form colonies in semisolid culture or in lethally irradiated hosts. Thus, taken together, our data suggest that ex vivo CT-B treatment can be used in allogeneic BM transplantation to prevent acute GVHD.

Topics: Animals; Bone Marrow Transplantation; Cholera Toxin; Colony-Forming Units Assay; Concanavalin A; Cytotoxicity Tests, Immunologic; Enzyme-Linked Immunosorbent Assay; Female; Flow Cytometry; Graft vs Host Disease; Immunosuppressive Agents; Lipopolysaccharides; Lymphocyte Culture Test, Mixed; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Inbred DBA

The response of IFN-gamma, IL-2 and IL-5 mRNA expression to the stimulation of concanavalin A (Con A) in peripheral blood mononuclear cells (PBMC) after bone marrow transplantation (BMT) was analyzed using reverse-transcriptase polymerase chain reaction (RT-PCR) to assess the recovery of T cell function. The subjects were 23 patients undergoing allogeneic BMT, 1 syngeneic BMT, 1 autologous BMT and 2 normal individuals. IFN-gamma mRNA expression increased after Con A stimulation in 6 patients who had limited chronic graft versus host disease (GVHD), 14 patients who did not have chronic GVHD, each one patient receiving syngeneic and autologous BMT and 2 normal individuals. On the other hand, IFN-gamma mRNA expression was not increased by Con A stimulation in 4 patients who had extensive chronic GVHD. Also, the concentration of IFN-gamma in cultured medium in a patient with extensive chronic GVHD was not detectable. A similar low response of IL-2 and IL-5 mRNA expression to Con A was observed in these patients with extensive chronic GVHD. These findings indicate that the cytokine productive capacity of T cell (IFN-gamma and IL-2 could be produced by type 1 T helper (Th1) cells and IL-5 could be produced by type 2 T helper (Th2) cells) was suppressed in patients who had extensive chronic GVHD, while that capacity was almost normal in patients without chronic GVHD and with limited chronic GVHD. Therefore, the analysis of cytokine gene response to Con A stimulation may provide useful information regarding immune reconstitution after BMT.

Topics: Adolescent; Adult; Base Sequence; Bone Marrow Transplantation; Chronic Disease; Concanavalin A; Cytokines; DNA Primers; Female; Gene Expression; Graft vs Host Disease; Humans; In Vitro Techniques; Interferon-gamma; Interleukin-2; Interleukin-5; Leukocytes, Mononuclear; Male; Molecular Sequence Data; Polymerase Chain Reaction; RNA, Messenger

Topics: Animals; Cells, Cultured; Concanavalin A; Glycine; Graft vs Host Disease; Immunosuppressive Agents; Lymph Nodes; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Lymphocyte Transfusion; Lymphocytes; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred CBA; Skin Transplantation; Spleen; Sulfides; Thiophenes; Transplantation, Homologous

The generation of nitric oxide (.N = O) during in vitro assays involving lymphocyte-macrophage interaction can result in profound inhibition of lymphocyte proliferation. The present study examined whether .N = O synthesis plays a role in the suppression observed in immune function assays during graft vs host disease (GvHD). By using a parent to F1 model to induce GvHD (C57BL/6J to C57BL/6J x DBA 2J F1), a mild but transient increase in serum NO2- plus NO3- levels was observed on day 12 after inoculation. Resident peritoneal macrophages obtained from mice with GvHD demonstrated enhanced .N = O synthesis in response to LPS, compared with control F1 peritoneal macrophages. Similarly, when splenocytes from GvHD mice were cultured with Con A or LPS enhanced supernatant NO2- levels were observed, compared with control F1 mice. Addition of NG-monomethyl-L-arginine (NMA), a competitive inhibitor of .N = O synthesis, resulted in decreased NO2- levels and greatly enhanced proliferation in response to Con A. Addition of NMA to LPS-stimulated cultures did not enhance proliferation, perhaps as the result of the paucity of B cells in the GvHD population. LPS-induced .N = O synthesis by GvHD splenocytes was blocked by anti-IFN-gamma mAb, whereas Con A-induced .N = O synthesis was relatively unaffected by similar concentrations of anti-IFN-gamma mAb, suggesting different mechanisms of induction of .N = O synthesis. A proliferative response of splenocytes from mice with GvHD to third-party alloantigen was not detectable, even in the presence of NMA. The suppression observed when splenocytes from GvHD animals were added to control TNP-modified self cultures was partially reversed in the presence of NMA. These results demonstrate that .N = O synthesis in both splenocyte and peritoneal macrophage populations from GvHD mice is enhanced, revealing that in vivo priming of macrophages for .N = O synthesis occurs during GvHD. Some, but not all, in vitro tests of immune function by using GvHD splenocytes are suppressed by the generation of .N = O.

Topics: Animals; Arginine; Concanavalin A; Female; Graft vs Host Disease; Immune Tolerance; Lipopolysaccharides; Lymphocyte Activation; Macrophages; Mice; Mice, Inbred Strains; Nitrates; Nitric Oxide; Nitrites; omega-N-Methylarginine; Peritoneal Cavity; Trinitrobenzenes

Irradiated C57BL/6(B6) mice, when they were injected with spleen cells of C57BL/6J-lpr/lpr(B6-lpr) mice, developed splenomegaly at 2 weeks post-transfer, but afterward displaced by GVH-like disease. At 2 weeks the enlarged spleen in the chimeric mice, designated as [B6-lpr----B6] chimera, contained about 70% of the total cell population as CD8-positive T cells. Spleen cells from [B6-lpr----B6] chimeras were unresponsive to Con A and LPS stimulation and suppressed the mitogenic response of B6, B6-lpr, and C3H spleen cells to Con A. However, they had no cytotoxic activity towards Con A blasts of B6 and B6-lpr spleen cells. The suppressor activity found in the [B6-lpr----B6] spleen cells was removed by pretreatment of them with anti-Thy-1.2 or anti-CD8(Lyt2.2) plus complement. The present experiment showed that enormous proliferation of CD8-positive suppressor T cells was induced in the [B6-lpr----B6] chimeras. These cells were probably responsible for the GVH-like lymphoid atrophy observed in these [B6-lpr----B6] chimeras.

Topics: Animals; Antigens, Surface; CD4 Antigens; CD8 Antigens; Cell Division; Complement System Proteins; Concanavalin A; Cytotoxicity, Immunologic; Disease Models, Animal; Female; Flow Cytometry; Graft vs Host Disease; Immune Tolerance; Interleukin-2; Leukocyte Common Antigens; Lipopolysaccharides; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Radiation Chimera; Spleen; T-Lymphocyte Subsets; Thy-1 Antigens

Ultraviolet irradiation inhibits alloreactive and mitogen-induced responses and might reduce both graft-versus-host and host-versus-graft reactions after bone marrow transplantation (BMT). We have studied proliferative responses to mitogens and reactivity in mixed lymphocyte culture after irradiation with ultraviolet (UV)-B light using splenocytes from Balb/c (H-2d) and CBA (H-2k) mice. Response to mitogens and in MLC was strongly inhibited by 20 J/m2 and abolished at 50 J/m2. Clonogenic cell recovery (CFU-GM; CFU-S) after UV-B irradiation was also reduced. When bone marrow and spleen cells were transplanted from parent (Balb/c) animals into F1 hybrid (Balb/c X CBA) recipients, all animals died with features indicative of graft-versus-host disease (GVHD) in 34 days. If the grafts were first irradiated with 100 J/m2 of UV-B at a mean wavelength of 310 nm, then 76% survived to day 80 when they were killed and shown to have normal marrow cellularity. The remainder died in marrow aplasia or of GVHD. H-2 typing in a group of surviving recipients showed either donor hematopoiesis only (8 of 15), mixed allogeneic chimerism (5 of 15), or recipient type hematopoiesis (2 of 15). Higher doses (200 to 300 J/m2) were detrimental to survival with 88% of recipients dying in marrow aplasia. Syngeneic BMT in Balb/c mice showed slower hematopoietic reconstitution when the grafts were first irradiated with 100 J/m2. After BMT from Balb/c to CBA mice all recipients of unirradiated grafts died within 54 days. By contrast, after graft irradiation with 100 J/m2 survival of recipient animals to day 80 was 59%. If these grafts were treated with 50 J/m2 survival was only 26% with an increase in deaths due to GVHD. Hematopoiesis at day 80 in a group of survivors studied by Ig heavy chain allotyping indicated donor type hematopoiesis in 6 of 10 (50 J/m2) and 2 of 9 (100 J/m2). These data indicate that UV-B irradiation inhibits lymphocyte reactivity and can prevent GVHD. However, there is clear in vitro and in vivo evidence of stem cell damage, such that autologous marrow recovery was demonstrated in a proportion of recipients. In parent----F1 UV-irradiated transplants, sustained hematopoietic recovery was effected in the majority by donor stem cells.

Topics: Animals; Bone Marrow; Bone Marrow Transplantation; Cell Division; Colony-Forming Units Assay; Concanavalin A; Graft vs Host Disease; Hematopoiesis; Hybridization, Genetic; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Lymphocytes; Mice; Mice, Inbred BALB C; Mice, Inbred CBA; Phytohemagglutinins; Spleen; Ultraviolet Rays

There is growing interest in studying pathways of mast cell activation. In a mouse model of chronic graft-vs-host disease (cGVHD) extensive mast cell activation and degranulation occurs in vivo coincident with the development of dermal fibrosis. An interesting feature of this model is that the mast cell reaction is slow to develop, occurring over a period of weeks and waning by 300 days. The aim of our work was to investigate the effects of supernatants from splenocytes of such cGVHD mice (cGVHD sups) on mouse and rat peritoneal mast cells cocultured with 3T3 skin fibroblasts. We found that cGVHD sups are able to release histamine from both mouse and rat cultured mast cells in a slow fashion. Histamine release became evident only after 5-8 days of coculture of the mast cells with the cGVHD supernatants and thereafter decreased to basal levels. Mast cell activation due to cGVHD supernatants was a noncytotoxic event as demonstrated by mast cell counts in the cocultures and by the ability of mast cells to exclude trypan blue. Mast cells that had been activated by incubation with the cGVHD sups were as responsive to stimulation with either anti-IgE antibodies or compound 48/80 as were mast cells incubated with control sups. Supernatants from mice early in GVHD (Days 11-28) were most active in promoting histamine release. Supernatants from spleens of mice which had GVHD for 290 days and where the mast cells had returned to full granulation in vivo were inactive. This is the first in vitro study demonstrating slow mast cell histamine release instituted by other cells, namely the splenocytes of cGVHD mice.

Topics: Animals; Cell Degranulation; Cells, Cultured; Concanavalin A; Culture Media; Fibroblasts; Graft vs Host Disease; Graft vs Host Reaction; Histamine Release; Male; Mast Cells; Mice; Mice, Inbred BALB C; Peritoneal Cavity; Rats; Spleen

Immunosuppression is a well-characterized consequence of chronic graft-versus-host disease (GVHD). We have previously shown that interferon (IFN) is produced in high levels during acute GVHD. Our objective in this study was to determine if IFN, as a cytokine with known immunosuppressive qualities, could be detected in mice experiencing chronic GVHD-induced immunosuppression. Two different experimental models were used to induce chronic GVHD. The first model involved the injection of parental strain spleen cells into adult F1 hybrids (AJ----B6AF1), while the second model utilized GVHD induced across minor histocompatibility barriers (B10.D2----BALB/c). Results indicated that significant levels of serum IFN-alpha/beta are present in mice undergoing chronic GVHD. Spleen cells from chronic GVHD mice were also shown to produce significant levels of IFN-alpha/beta upon in vitro culture in medium only. This IFN-alpha/beta production was greatly increased when GVHD spleen cells were cultured with either concanavalin A (Con A) or IL-2. In contrast, IFN-gamma production was undetectable in these Con A- or IL-2-containing cultures. Additionally, these same spleen cells which produced high levels of IFN-alpha/beta were immunosuppressed as measured by mitogen-induced cell proliferation. These results suggest that IFN-gamma production is defective in GVHD spleen cells, and that the presence of high IFN-alpha/beta production by GVHD mice may contribute to the immunosuppression associated with chronic GVHD.

Topics: Acute Disease; Animals; Bone Marrow Transplantation; Cells, Cultured; Chronic Disease; Concanavalin A; Female; Graft vs Host Disease; Immune Tolerance; Interferon Type I; Interferon-gamma; Interleukin-2; Lymphocyte Activation; Mice; Mice, Inbred Strains; Spleen

Topics: Animals; Antigens, Surface; Bone Marrow Transplantation; Cell Differentiation; Cell Division; Cells, Cultured; Concanavalin A; Cyclosporins; Female; Graft vs Host Disease; Phenotype; Rats; Rats, Inbred Lew; T-Lymphocytes

Topics: Adolescent; Adult; Bone Marrow Transplantation; Concanavalin A; Graft vs Host Disease; Humans; Interleukin-2; Lymphocyte Activation; Middle Aged; Phytohemagglutinins; T-Lymphocytes, Regulatory; Transplantation, Homologous

Although isolated intraepithelial lymphocytes (IEL) have been shown to have specific and non-specific cytolytic functions, their ability to proliferate in response to T-cell mitogens or alloantigens is controversial. Here we show that IEL from mouse small intestine do not respond to mitogens such as concanavalin A and phytohaemagglutinin A or in mixed lymphocyte reactions unless an accessory spleen cell is also present. Adherent spleen cells possess the most potent helper function, but a dividing accessory cell may also be required. Supernatants from stimulated lymphocytes also assist IEL to proliferate in vitro, particularly in the presence of adherent accessory cells. IEL could not mediate lethal graft-versus-host disease in irradiated hosts, but could produce popliteal lymph node hypertrophy or splenomegaly in unirradiated hosts. Thus, IEL have the potential for proliferative activities characteristic of T cells, but they require accessory cells and/or factors such as interleukin-2 for their function in vitro and in vivo.

Topics: Animals; Antigen-Presenting Cells; Concanavalin A; Female; Graft vs Host Disease; Graft vs Host Reaction; In Vitro Techniques; Intestine, Small; Lipopolysaccharides; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Lymphocytes; Male; Mice; Mice, Inbred Strains; Mitogens; Mitosis; Phytohemagglutinins; Spleen

The following findings were noted among 45 bone marrow transplant recipients. The patients without cytomegalovirus (CMV) infection or chronic graft-versus-host disease (GVHD) showed normal lymphocyte stimulation in vitro by concanavalin A (Con A) more than 3 months after transplantation, and normal stimulation by phytohemagglutinin (PHA), anti-beta 2-microglobulin (A-beta 2m) and protein A (SpA) after 6 months. In contrast, the patients who had CMV infection without chronic GVHD had Con A and SpA responses within the normal range after 12 months and reduced lymphocyte responses to PHA and A-beta 2m more than 12 months after transplantation. The patients with chronic GVHD had reduced responses to all of these four mitogens after more than 12 months. In comparison with other patients those who later developed chronic GVHD showed an increased mixed lymphocyte culture stimulation during the first 3 months that decreased between 6-12 months. Patients with chronic GVHD still had reduced IgA levels at 12 months after transplantation. Patients with CMV infection, but without chronic GVHD, had higher percentages of lymphocytes with surface membrane Ig than healthy controls during the first 3 months after transplantation. The data suggest that CMV infection, regardless of chronic GVHD, delays immunologic recovery after marrow transplantation.

Topics: Antigens, Surface; beta 2-Microglobulin; Bone Marrow Transplantation; Concanavalin A; Cytomegalovirus Infections; Graft vs Host Disease; Humans; Immunoglobulins; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Phytohemagglutinins; Staphylococcal Protein A

We have been studying the mitogen hyporesponsiveness and immunosuppression induced in chronic murine graft-vs.-host disease (GVHD) induced across minor histocompatibility (MiHA) barriers. In this system, donor and recipient mice are major histocompatibility complex- and mls-identical, and are nonreactive in primary mixed leukocyte reactions. Spleen cells from B10.D2 (H-2d, mls b) mice were injected into irradiated (600 rad) BALB/c (H-2d, mls b) recipients. Recipient spleen cells are hyporesponsive to mitogens, and contain natural suppressor (NS) cells. We investigated the cellular requirements for both the in vivo induction and the in vitro expression of this GVH suppression. T cells are required in the graft, but they are not sufficient to induce suppression, and a non-T cell population is also required for maximum induction in vivo. T cells are also required for the maximum expression of NS cell suppressive ability in vitro. Early in the course of GVH, the suppressor cells are able to suppress the Con A and LPS response of all mouse strains tested (except for the relative difficulty in suppressing the B10.D2 LPS response). Later, they become almost completely unable to suppress the B10.D2 LPS response; while still being able to suppress the Con A and LPS response of all other strains tested (including the B10.D2 Con A response). This inability to suppress a B10.D2 LPS response can be brought back to almost complete suppression by the addition of concanavalin A supernatant (CAS). We present a hypothesis to explain what may be a common mechanism for GVH-induced suppression, total lymphoid irradiation-induced suppression, and neonatal tolerance. These situations all include rapidly proliferating lymphohematopoietic stem cell populations, and also have large numbers of NS cells. NS cells can suppress proliferating lymphoid populations, and their development and activity are greatly enhanced by T cell signals such as are supplied by donor T cells in chronic GVHD. Thus, NS cells may feed back on and downregulate self-reactive T cells or T cells responding to introduced foreign antigens.

Topics: Animals; Bone Marrow Transplantation; Concanavalin A; Graft vs Host Disease; Lymph Nodes; Lymphocyte Activation; Lymphocyte Depletion; Lymphocyte Transfusion; Lymphocytes; Lymphokines; Mice; Mice, Inbred Strains; Radiation Chimera; Spleen; T-Lymphocytes; T-Lymphocytes, Regulatory; Thymus Gland

In this study we investigated the mechanism, or mechanisms, involved in graft-versus-host (GVH)-induced T cell immunodeficiency. Chronic GVH reactions were induced in normal CBA X A F1 (BAF1) hybrid mice by the injection of parental A strain lymphoid cells. At various times (43-91 days) after GVH induction, the functional status of GVH T cells was assessed using interleukin-1 (IL-1) and interleukin-2 (IL-2) as probes. The response of GVH thymocytes to IL-1 was depressed when compared with normal thymocytes. Although GVH peanut-agglutinin-negative (PNA-) thymocytes did respond to IL-2 alone or IL-2 plus phytohemagglutinin (PHA), this response was significantly lower than the response of PNA- thymocytes from normal mice. In addition, GVH spleen cells failed to produce significant amounts of IL-2 when stimulated with concanavalin A. These results suggest that the long term immunosuppression associated with murine chronic GVH disease is due, at least in part, to a decrease in the responsiveness to IL-1 and IL-2, and to a marked deficiency in IL-2 production.

Topics: Animals; Antibody Formation; Concanavalin A; Female; Graft vs Host Disease; Immune Tolerance; Interleukin-1; Interleukin-2; Lymphocyte Activation; Male; Mice; Mice, Inbred Strains; Rosette Formation; T-Lymphocytes

In the recipients of an allogeneic HLA-identical sibling transplant the blood leucocytes were reconstituted within 3 to 4 weeks but the level of lymphocytes remained low throughout the observation period of 20 weeks. Of the different lymphoid cell subsets, the large granular lymphocytes (LGL) reconstituted fastest, followed by DR-expressing lymphocytes. The reconstitution was accompanied by a significant lymphoid blastogenesis in the blood. The frequency of OKT4 and OKT8 lymphocytes was initially low; the number of OKT8-positive lymphocytes reached normal levels by the 6-8th week whereas the number of OKT4-positive lymphocytes and, consequently, the OKT4/8 ratio remained low. The responses to the T mitogens, phytohaemmagglutinin and concanavalin A, were strongly suppressed. Only a few significant changes were observed before and during acute graft-versus-host disease (aGVHD): the frequency of LGL, lymphoid blasts and OKT8-expressing lymphocytes was depressed before aGVHD and the frequency of lymphoid blasts remained depressed throughout the episode. We conclude that reproducible changes occur in the leucocyte subset frequencies during reconstitution, but the characteristic changes prior to and during aGVHD are not particularly prominent and hardly of diagnostic use.

Topics: Adult; Bone Marrow Transplantation; Concanavalin A; Cyclophosphamide; Female; Graft vs Host Disease; Humans; Lymphocyte Activation; Lymphocytes; Male; Methotrexate; Methylprednisolone; Phytohemagglutinins

Human bone marrow was harvested from surgically resected bones of 25 patients and was tested for the presence of mature T cells. An average of 6.5% (+/- 1.2% SE) of nucleated bone marrow cells formed spontaneous rosettes with sheep red blood cells. Functional T cells in bone marrow were also identified by characteristic responses to alloantigens and the T cell mitogens concanavalin A (Con A) and phytohemagglutinin (PHA). The ability of three monoclonal antibodies (OKT.3, Lyt-3, and (Leu-1) to lyse peripheral T cells in the presence of rabbit complement was examined. All three reagents were found to be specifically lytic for mature T cells in peripheral blood. One reagent (Leu-1) was selected for use in depletion of T cells in human bone marrow. Seven of 10 experiments performed showed sufficient T cell responses to be evaluable. In all of these experiments, a marked reduction of T cells and T cell functions was observed. On the average, E rosettes were reduced 89.2% (+/- 3.0% SE) below medium controls while the mean PHA, Con A, and mixed lymphocyte culture (MLC) activity were completely eliminated to levels below background. In four experiments, colony-forming units (CFU-GM) in bone marrow were assayed following treatment with Leu-1 and showed a mean increase of 194% (+/- 32% SE) over medium controls. Since mature T cells are thought to be responsible for graft-versus-host disease in allogeneic bone marrow transplantation, this method of T cell depletion may be useful for preparing marrow for human bone marrow transplants.

Topics: Antibodies, Monoclonal; Bone Marrow Transplantation; Colony-Forming Units Assay; Complement System Proteins; Concanavalin A; Graft vs Host Disease; Hematopoietic Stem Cells; Humans; Lymphocyte Culture Test, Mixed; Lymphocyte Depletion; Phytohemagglutinins; Rosette Formation; T-Lymphocytes

Topics: Animals; Antibody-Producing Cells; Bone Marrow Cells; Bone Marrow Transplantation; Cell Survival; Cells, Cultured; Concanavalin A; Graft vs Host Disease; Graft vs Host Reaction; Hematopoietic Stem Cells; Male; Mice; Radiation Injuries, Experimental; Spleen; Transplantation, Homologous

Topics: Animals; Antilymphocyte Serum; Cell Division; Clone Cells; Concanavalin A; Female; Graft vs Host Disease; Hematopoietic Stem Cell Transplantation; Hematopoietic Stem Cells; Immunity, Cellular; Lectins; Lymphocytes; Male; Mice; Radiation Chimera; Spleen; Transplantation, Homologous