ABSTRACT
Three monoclonal antibodies identified as D8, B11 and C5 of different specificities have been produced against human antithrombin III (AT). The apparent dissociation constants (Kd app) of the AT-antibody interaction were determined by ELISA method: Kd app (D8) = 2.4 nmole, Kd app (B11) = 13 nmole, Kd app (C5) = 24 nmole. All three antibodies reacted with isolated AT on immunoblots obtained with "native" PAGE. The D8 antibody also reacted with plasma and serum AT while B11 antibody reacted with serum thrombin-antithrombin (TAT) complexes as well.
Subject(s)
Antibodies, Monoclonal/immunology , Antithrombin III/immunology , Animals , Antibodies, Monoclonal/isolation & purification , Antibody Specificity , Enzyme-Linked Immunosorbent Assay , Humans , Immunoglobulin G/immunology , Kinetics , Mice , Mice, Inbred BALB C/immunology , Peptide Hydrolases/immunologyABSTRACT
Treatment of mouse donors or recipients of allogeneic spleen cells with repeated injections of lentil lectin markedly suppressed regional and systemic graft-versus-host reactions. The lectin acted selectively on lymphoid cells; hematopoietic stem cells were largely left unaffected. Repeated administration of the lectin decreased the number of lymphocytes and red blood cells and the amount of hemoglobin; other white blood cells increased in number.
Subject(s)
Graft vs Host Reaction , Hematopoietic Stem Cells , Immune Tolerance , Lectins/pharmacology , Plant Lectins , Adjuvants, Immunologic , Animals , Erythrocyte Count , Female , Injections, Intraperitoneal , Injections, Intravenous , Lectins/administration & dosage , Lectins/immunology , Leukocyte Count , Mice , Mice, Inbred Strains , Spleen/cytology , Tissue Donors , Transplantation, HomologousABSTRACT
Mouse spleen and bone marrow cells were incubated for 2 h with ZnCl2 and Li2SO4 at different concentrations and tested for the ability to evoke the graft-versus-host reactions (GVHR) and to form pluripotent haemopoietic colonies. ZnCl2 at concentrations 5 X 10(-6) to 5 X 10(-4) M inhibited the regional GVHR. At a concentration of 5 X 10(-6) ZnCl2 also inhibited the ability to elicit the systemic GVHR in irradiated mice. Haemopoiesis was stimulated in cells incubated with ZnCl2 at concentration 5 X 10(-6) M, it was inhibited after incubation of A/01a cells with 5 X 10(-4) M ZnCl2, whereas cells from non-inbred ICR mice were stimulated by the latter concentration of ZnCl2 for haemopoiesis. Li2SO4 inhibited the ability to induce the regional GVHR at concentrations of 5 X 10(-3) and 10(-2) M, had no effect on survival of mice during the systemic GVHR and exerted rather an inhibitory effect on haemopoiesis.
Subject(s)
Graft vs Host Reaction/drug effects , Hematopoiesis/drug effects , Lithium/pharmacology , Zinc/pharmacology , Animals , Bone Marrow/drug effects , Bone Marrow Transplantation , Colony-Forming Units Assay , Female , In Vitro Techniques , Male , Mice , Mice, Inbred Strains , Spleen/drug effects , Spleen/transplantation , Transplantation, HomologousABSTRACT
A synthetic polymer, Duxon, was developed and tested as a substitute of blood plasma for transfusion purposes. Tests of this preparation included a test for its influence on the haemopoietic stem cells and the graft-versus-host reaction (GVHR). A single dose or repeated doses of Duxon did not reduce the number of pluripotent haemopoietic colonies (CFU-S) in mice and short-term incubation of cells from haemopoietic organs of mice with Duxon resulted in a slight, yet significant, increase in the number of CFU-S. Injection of Duxon non-significantly diminished the manifestations of the regional GVHR. Incubation of spleen cells with Duxon significantly reduced their GVH reactivity in the regional test. The systemic GVHR was moderately inhibited by repeated doses of Duxon in sublethally irradiated mice. A significant delay in deaths of animals from GVHR was seen in lethally irradiated mice receiving a single dose of Duxon on day 4 after GVHR induction. Survival of lethally and sublethally irradiated mice was prolonged following injection of cells incubated with Duxon. Possible uses of Duxon, a preparation with mild immunosuppressive properties, not impairing haemopoiesis, are discussed.
Subject(s)
Graft vs Host Reaction/drug effects , Hematopoiesis/drug effects , Polymethacrylic Acids/pharmacology , Animals , Colony-Forming Units Assay , Female , Hematopoietic Stem Cells/drug effects , Immunosuppressive Agents , Mice , Plasma Substitutes/pharmacology , Spleen/transplantation , Transplantation, HomologousSubject(s)
Antigens/analysis , Factor VIII/analysis , Factor VIII/immunology , Hemophilia A/genetics , Adolescent , Adult , Aged , Female , Hemophilia A/blood , Hemophilia A/diagnosis , Humans , Middle Aged , Probability , von Willebrand FactorABSTRACT
The chromatographic separation of crude papain preparations on Sephadex G-50 (fine) enables pure papain to be obtained in a single step. Immunochemical techniques have been found to be very convenient for testing the purity of the individual chromatographic fractions. A general approach is presented that makes it possible to follow the course of the chromatographic purification of any immunogenic compound by simple qualitative immunochemical techniques that can be applied in any laboratory.
Subject(s)
Papain/isolation & purification , Chromatography, Gel/methods , Electrophoresis, Disc/methods , Immunodiffusion/methods , Immunoelectrophoresis/methods , Immunoelectrophoresis, Two-Dimensional/methodsABSTRACT
The effect of a cytostatic drug Benfluron-- 5(2-N,N-dimethyl-amino-ethoxy)-7-oxo-7H-benzo(c)fluorene hydrochloride was tested in mice on skin graft survival, graft-versus-host reaction (GVHR) and for mitogenic stimulation of human lymphocytes. Application of Benfluron resulted in a prolonged skin graft survival. The regional and systemic GVHR was potentially inhibited by p. o. administration of Benfluron at a dose of 100 mg/kg of body weight. Allogeneic spleen cells incubated with Benfluron at concentration of 5-20 micrograms/ml showed reduced capability to induce GVHR. PHA or PWM stimulation of lymphocytes was completely inhibited in the presence of 2 micrograms/ml Benfluron. Benfluron as a potent immunosuppressive agent can be considered useful for the prevention of GVHR.
Subject(s)
Antineoplastic Agents/pharmacology , Fluorenes/pharmacology , Immunity, Cellular/drug effects , Lymphocyte Activation/drug effects , Animals , Cell Survival/drug effects , Female , Graft vs Host Reaction/drug effects , Graft vs Host Reaction/radiation effects , Humans , Male , Mice , Mice, Inbred Strains , Skin/cytology , Skin/drug effects , Skin TransplantationABSTRACT
The effect of Benfluron-- 5-(2-N,N-dimethylamino-ethoxy)-7-oxo-7H-benzo(c)fluorene hydrochloride--on hemopoietic stem cells was tested by the production of spleen cell colonies (CFU-S) in irradiated mice following application of bone marrow cells and by the production of hemopoietic colonies (CFU-C) in semisolid agar. The reduced numbers of CFU-S were found in mice applied Benfluron or Benfluron-treated bone marrow cells. The increased numbers of CFU-S, however, were found in mice receiving bone marrow cells from Benfluron-treated donors. The production of CFU-C was decreased after 2 h incubation of bone marrow cells with 0.5-5 micrograms/ml of Benfluron, and fully inhibited at higher concentrations.
Subject(s)
Antineoplastic Agents/pharmacology , Fluorenes/pharmacology , Hematopoietic Stem Cells/cytology , Animals , Bone Marrow/drug effects , Bone Marrow/radiation effects , Bone Marrow Cells , Cell Division/drug effects , Cells, Cultured , Female , Hematopoietic Stem Cells/drug effects , Hematopoietic Stem Cells/radiation effects , Mice , Mice, Inbred StrainsABSTRACT
Using different isolation procedures (after acidification and saturation with 3 M ammonium sulphate) three fractions were isolated from bull seminal vesicle fluid and assayed for their effects on cell immunity in vitro and in vivo. Two of these preparations (ZS RNase and AS RNase) possessing a high level of ribonuclease activity at concentrations of 50 micrograms/ml showed inhibitory effects (up to 80%) on 3H-thymidine incorporation into the DNA of mitogen-or antigen-stimulated human lymphocytes. The third preparation (3M-P) possessing low ribonuclease activity showed lesser inhibitory effects. The potency of mouse spleen cells to cause regional GVH reaction was significantly decreased after preincubation of spleen cells to cause of 1 mg per AS RNase or ZS RNase whereas 3M-P was ineffective in this test. A single dose of 1 mg per mouse of ZS RNase or 3M-P administered i.p. on day 4 after skin transplantation significantly prolonged graft rejection. Both preparations at this dose potentiated the effect of cyclophosphamide on skin graft survival. All tested preparations preincubated with mouse bone marrow cells had no adverse effects on their colony-forming activity (in the spleens of irradiated mice). The possibility of utilizing the preparations with ribonuclease activity isolated from vesicle fluid in clinical bone marrow transplantation is discussed.
Subject(s)
Body Fluids , Endoribonucleases/pharmacology , Immunosuppression Therapy , Seminal Vesicles/enzymology , Animals , Body Fluids/immunology , Cattle , Colony-Forming Units Assay , Endoribonucleases/immunology , Female , Graft Survival/drug effects , Graft vs Host Reaction/drug effects , Lymphocyte Activation/drug effects , Male , Mice , Mice, Inbred ICR , Skin TransplantationABSTRACT
There was studied the influence on the cell-mediated and humoral response in vivo manifested by selected methylpropionic acid and pyridazinone-3 derivatives which had been found to possess strong immunotropic effects in the in vitro screening previously. It was shown that the compounds were generally poorly tolerated by animals, and they exerted only weak suppressive effects on antibody production, the contact hypersensitivity and survival of skin grafts. This immunosuppressive activity was accompanied by a slight decrease in the number of spleen colony forming cells (CFU-s). Only limited correlation between the biological activity of the preparations and their chemical structure was found.
Subject(s)
Hematopoiesis/drug effects , Immunity/drug effects , Pyridazines/pharmacology , Animals , B-Lymphocytes/drug effects , Dermatitis, Contact , Dinitrochlorobenzene/immunology , Graft Survival/drug effects , Guinea Pigs , Hemagglutination/drug effects , Immunity, Cellular/drug effects , Mice , Propionates/pharmacology , Rats , Skin Transplantation , T-Lymphocytes/drug effectsABSTRACT
The effect of 5-azacytidine (5-AzCR) and 5-aza-deoxycytidine (5-AzCdR) on the survival of skin grafts in mice and rats, the action of these drugs on regional GVH reaction, as well as the formation of haemopoietic colonies (CFU-5) in the spleen were studied. Both drugs prolonged the life span of skin grafts when administered 24 hr before transplantation, or on the 4th post-transplantation day. However, they were little effective when injected 24 hr after skin grafting, or after induction of the regional GVHR. Following intraperitoneal administration, they inhibited CFU-5 formation. Two-hour incubation in vitro of cells with 5-AzCR significantly reduced their GVH reactivity and capacity to form CFU-5; 5-AzCdR under the same conditions was ineffective.
Subject(s)
Azacitidine/analogs & derivatives , Azacitidine/pharmacology , Graft Survival/drug effects , Immunity, Cellular/drug effects , Animals , Antineoplastic Agents/pharmacology , Decitabine , Female , Male , Mice , Mice, Inbred A/immunology , Rats , Rats, Inbred Lew/immunology , Skin TransplantationABSTRACT
The use of semisolid medium for the culture and cloning of haematopoietic cells has helped our understanding of their proliferation and differentiation. It has been shown that mixed colonies of granulocytes and macrophages developed, in the presence of colony-stimulating factor (CSF), from their common precursor, granulocyte-macrophage colony-forming cells (GM-CFC). Attempts at recloning these colonies in semisolid medium suggested that granulocytes and macrophages were differentiated cells incapable of further proliferation. However, our studies on cultures of larger numbers of cells demonstrate that while this may be the case for granulocytes, macrophages seem to be capable of long-term proliferation.
Subject(s)
Bone Marrow Cells , Cells, Cultured , Macrophages/cytology , Animals , Cell Division , Cell Line , Culture Media , Granulocytes/cytology , Mice , Mice, Inbred ICR , Pinocytosis , Receptors, Complement/metabolism , Receptors, Immunologic/metabolismABSTRACT
The present paper deals with the effect of Damvar on hemopoiesis. The test of formation of hemopoietic splenic colonies (CFU-S) was used in mice, the peripheral blood picture and bone marrow differential after long-term administration of Damvar was tested in rats and dogs. The bone marrow cells from mice after a single dose of Damvar or cells incubated with Damvar in vitro showed no reduction of colony-forming activity. A single dose or repeated variable doses of Damvar, or combined administration of Damvar and Cyclophosphamide in some cases influenced the number of CFU-S in irradiated mice following transplantation of normal murine bone marrow cells. A number of CFU-S lower than 75% of the control count was never found. The application of Damvar to rats and dogs caused no significant changes in their blood pictures and bone marrows.
Subject(s)
Antineoplastic Agents/pharmacology , Colony-Forming Units Assay , Hematopoiesis/drug effects , Pyrimidinones/pharmacology , Animals , Bone Marrow/radiation effects , Bone Marrow Transplantation , Dogs , Female , Hematopoiesis/radiation effects , Male , Mice , Rats , Transplantation, HomologousABSTRACT
By fractionation on Sephadex columns of the supernatants from mixed cultures of H-2 incompatible lymphocytes we succeeded in partially purifying a lymph node activating factor and in determining its molecular weight. The properties of the lymph node activating factor correspond to those of substances with 16 000 +/- 5 000 D, 65 000 +/- 10 000 D, and also substances with molecular weight higher than 200 000 D which we consider to be aggregates of the minor components. The lymph node activating factor resembles in its molecular weight and effects (in vitro lymph node activation) the products of activated lymphocytes described by other investigators.
Subject(s)
Histocompatibility Antigens/analysis , Lymph Nodes/physiology , Lymphocytes/metabolism , Animals , Cells, Cultured , Chromatography, Gel , Lymphocyte Activation , Lymphocytes/immunology , Mice , Molecular WeightABSTRACT
Mouse sera obtained 14-18 h after transplantation of skin or spleen cells from H-2-disparate donors contain a "lymph node activating factor". They activate the draining lymph nodes of syngeneic recipients or F1 hybrids of the mouse strains, which have served as graft donors and recipients, and give specific precipitation lines with the antibody against LNAF. LNAF can be determined in the sera of mice, which have been sensitized with spleen cells prior to transplantation, from 7 to 48 h after transplantation.
Subject(s)
Lymphokines/blood , Skin Transplantation , Spleen/transplantation , Animals , Female , H-2 Antigens/analysis , Mice , Mice, Inbred Strains , Transplantation ImmunologyABSTRACT
The release of a lymph node activating factor is controlled by the genetic difference in the I region of the H-2 complex. Incompatibility at IA and IE loci is of decisive importance.
