Red Blood Cell Antibody Response Research Articles

HLA‐DRB1 associations in individuals with single and multiple clinically relevant red blood cell antibodies

A minority of red blood cell (RBC) alloantigen-exposed persons form antibodies. Responders are at high risk of developing additional antibodies upon subsequent transfusions. Several studies showed an association between particular HLA-DRB1 phenotypes and the development of specific RBC antibodies. This study evaluates the presence of HLA-DRB1 antigens in individuals with single or multiple RBC antibody specificities to explore whether the response against RBC antigens is associated with a summation of particular HLA-DRB1 susceptibility antigens. Frequencies of HLA-DRB1 alleles in individuals with antibodies against clinically relevant antigens were compared to a large population cohort to calculate odds ratios (ORs) for alloimmunization to different RBC antigens. The study cohort consisted of 941 individuals (female-to-male ratio, 3.8) possessing 1462 antibody specificities elicited by transfusion, pregnancy, transplantation, or a combination of these. Besides confirmation of known associations, new associations were identified for anti-E with DRB1*09 and for anti-S with DRB1*07 (ORs, 3.7 and 8.7, respectively). Multiple antibody formation was in a minority of cases associated with the presence of multiple DRB1 susceptibility genes. In multiple responders DRB1*15 was present in almost 40% of cases compared to approximately 25% in single-antibody responders and in the control population. This study suggests that HLA-DRB1 restriction plays an important role for a first RBC antibody response but multiple antibody formation seems less dependent on the presence of particular HLA restriction genes, while HLA-DRB1*15 may represent a susceptibility phenotype enhancing formation of multiple RBC antibody specificities.

Testosterone inhibits the immunostimulant effect of thymosin fraction 5 on secondary immune response in mice

The purpose of the present investigation was to examine the in vivo influence of testosterone on the immune properties of a thymic factor (thymosin fraction 5, TF5) a partially purified thymic preparation in male Swiss IOPS/OF1 mice (5–10 weeks old). Testosterone administration (100 μg/ml) significantly inhibited the enhanced anti-sheep red blood cell antibody response induced by TF5 (100 μg/ml); this inhibition was only observed on the secondary antibody response and not on the primary. These results suggest that gonadal steroids can affect the immune response by modulating the activity of thymic factors.

Antibody-directed affinity therapy applied to the immune system: In vivo effectiveness and limited toxicity of daunomycin conjugated to HPMA copolymers and targeting antibody

The applicability of targeting therapy intervention in lymphatic tissue was studied. The effect was measured as the inhibition of anti-sheep red blood cell antibody response expressed in plaque-forming cells. Daunomycin was used as the effective drug and polyclonal and monoclonal anti-Thy 1.2 or anti-Ia k antibody served for targeting. Both components were coupled to a soluble N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer with oligopeptidic side sequences which permitted a controlled release of the drug in the target tissue. HPMA copolymer conjugates with side sequences Gly-Phe-Leu-Gly cleavable by lysosomal enzymes decreased in vivo the antibody reaction by 60–85%. A comparable amount of free targeting antibody was without a significant effect. Injection of targeted daunomycin decreased the toxicity of the drug against hematopoietic precursors in bone marrow colony-forming unit-spleen 80 times compared to the same amount of free drug. The in vivo effectiveness of targeted daunomycin was confirmed morphologically. Application of free daunomycin lead to a significant irritation of Kupffer cells in liver while none of the daunomycin-antibody-copolymer conjugate had such an effect.

The Effect of Bacillus natto in Feed on the Sheep Red Blood Cell Antibody Response in Chickens

Chickens fed 10(6) or 10(7) of Bacillus natto strain BN/g of diet from hatching through 15 or 30 days of age showed an increase in ability to produce hemagglutinating antibody titer as determined by intravenous injection of sheep red blood cells, whereas no increase in antibody production ability was observed in the chickens fed 10(7) of BN/g of diet from hatching through 10 days of age.

The immunosuppressive effect of monoclonal anti-Lyt-1.1 antibodies in vivo.

Monoclonal anti-Lyt-1.1 alloantibody was produced as tissue culture supernatant and administered to mice. The antibody, given intraperitoneally, resulted in the suppression of all T cell functions studied, but was without direct effect on B cells. Thus, skin and tumour allograft survival was prolonged and there was suppression of the delayed-type hypersensitivity response; T cell help inthe anti-sheep red blood cell antibody response, responder cells in the mixed lymphocyte reaction (MLR), leucoagglutinin-responsive cells, cytotoxic T cell (Tc) function and the induction of Tc were either totally or partially suppressed, all these responses being mediated by Lyt-1+2- or Lyt-1+2+ cells in CBA/H mice. By contrast, there was no inhibitory effect on the MLR-stimulating or lipopolysaccharide-responsive cells. The administration of the anti-Lyt-1.1 antibody was accompanied by a depletion of Lyt-1.1+ T cells from both spleen and lymph node. These studies indicate that the monoclonal anti-Lyt-1.1 antibody is active in vivo with a selective effect on T cells. The results also have important implications for studies of T cell interactions in the mouse in vivo, and for similar studies in man.

Primary in vitro antibody response of rabbit lymphoid cells and T-B cell collaboration in the absence of detectable mitogens

To investigate whether the antibody response and T-B-cell collaboration in vitro can be obtained in the absence of mitogens, a method of obtaining an in vitro primary anti-sheep red blood cell antibody response by rabbit spleen and lymph node cells was developed. We used Marbrook culture vessels and a specially prepared medium containing 10% autologous serum and maintained at pH 7.4–7.6. The system was shown to be devoid of any polyclonal mitogens as assessed by [ 3H]thymidine incorporation and by direct examination for blast cells in stained smears. The primary response increased continuously over the 5-day cultivation period and only IgM but not IgG plaque-forming cells (PFC) were detected. In over 20 experiments, the response ranged from 357 ± 17 to 4425 ± 110 PFC/10 7 cultured cells with a median stimulation index of 52. The spleen cells required less antigen than the lymph node cells and 2-mercaptoethanol inhibited the response of the spleen cells but not that of the lymph node cells. Lymphocytes were separated into highly pure T- and B-cell populations by negative selection using antibody-coated human erythrocytes to rosette either T or B cells and Ficoll-Hypaque centrifugation to remove rosetted cells. Upon cultivation, B cells alone gave a low IgM response, whereas B cells reconstituted with T cells gave a response similar to that obtained with unseparated lymphoid cells. We concluded that: (a) optimal conditions for obtaining primary in vitro antibody responses using rabbit spleen and lymph node cells were established, (b) T-B-cell collaboration was demonstrated in the rabbit primary antibody response to sheep erythrocytes, and (c) the primary antibody response in vitro and T-B-cell collaboration may occur in the absence of detectable polyclonal mitogens.

CHIMAERISM OF IMMUNOCOMPETENT CELLS IN ALLOGENEIC BONE MARROW-RECONSTITUTED LETHALLY IRRADIATED CHICKENS

Injection of parental bone marrow cells into 12-day-old lethally irradiated F1 hybrid chickens resulted in chimaerism of donor-type graft-versus-host (GVH)-reactive cells and suppression of antisheep red blood cell antibody response. These manifestations of a chronic graft-versus-host reaction were prevented by pretreatment of the donor marrow with specific anti-T cell globulin. In some chimaeras donor-type GVH-reactive cells developed gradually from T cells precursors of donor origin. Transplantation of spleen and marrow cells from sheep red blood cell-primed F1 hybrid donors into lethally irradiated parental recipients resulted in the loss of memory potential within 1-2 weeks of transfer, whereas donor-type IgG allotype synthesis was preserved. Injection of goat antichicken thymocyte serum to recipients 1 day before reconstitution enabled the antibody response of memory cells at 1-2 weeks, although it failed to prevent their rejection by 8-9 weeeks after transplantation. Split chimaerism of donor-type GVH-reactive cells was demonstrated in chickens which had previously rejected the B cells derived from the same graft.

Cell-cooperation in anti-sheep red blood cell antibody response in mouse spleen cell cultures. Use of antilymphocyte globulin for selective suppression of the antigen reactive cells.

Restoration of impaired antibody response to sheep red blood cells (SRBC) in spleen cell cultures from mice treated with heterologous antilymphocyte globulin (ALG) was studied by adding normal cells from various sources, to explore the problems of cell-cooperation in anti-SRBC antibody response and the target of ALG. When spleen cells from ALG-treated mice were separated into macrophage-rich and lymphoid cell-rich subpopulations, only the latter was found to be impaired in the ability for anti-SRBC antibody response. Addition of even a small number of normal allogeneic spleen cells sufficiently restored the impaired anti-SRBC antibody response of the spleen cells from ALG-treated mice. By use of allo-antisera, most hemolysin plaque-forming cells (PFC) generated in such cultures were proved to be derived from the cells of ALG-treated mice. Restoration was also achieved by adding thymus-derived cells, which were obtained from spleens of mice heavily irradiated and repopulated with syngeneic thymus cells, or lymphoid cells directly collected from thymuses. All results indicate that ALG selectively depletes the thymus-derived antigen reactive cells (ARC) in the spleen cell population, and that ARC supplied from normal spleen or thymus can interact with plaque-forming cell precursors (PFCP) that remain intact in the spleen cell population of ALG-treated mice. The results also suggest that a single ARC interacts with more than one PFCP and makes them develop into PFC.

Cellular mechanisms of adjuvant action of bacterial lipopolysaccharide in anti-sheep red blood cell antibody response.

Adjuvant action of lipopolysaccharide (LPS) extracted from Salmonella typhimurium LT2 on anti-sheep red blood cell (sRBC) antibody response in mouse spleen was studied at the cellular level by the technique of localized hemolysis in agar. Injection of LPS caused significant increase in direct 19S and indirect 7S plaque-forming cells (PFC) in mouse spleen after the primary antigenic stimulation, while the adjuvant effect of LPS could hardly be assessed in the secondary PFC response. The adjuvant effect on the primary PFC response was most conspicuous when LPS was injected at the same time as the sRBC, and no effect could be observed when LPS was injected 1 or 5 hr before PFC determination. The mice who had received LPS with the primary sRBC stimulation showed much higher PFC level after the secondary sRBC stimulation, indicating the increase in the memory cells. Cell population consisting of lymphoid cells and macrophages was separated into ‘adherent’ (macrophage-rich) and ‘non-adherent’ (lymphoid cell-rich) fractions by incubating in petri dishes, and effect of LPS on each fraction was examined. Transfer of non-adherent lymphoid cells treated with LPS in vitro and simultaneous sRBC injection resulted in a significant increase of PFC in the spleen of the syngeneic recipient mice, but transfer of LPS-treated macrophages did not show any effect, suggesting that LPS acts on lymphocytes but not on macrophages. This idea appeared also to be supported by the results that LPS exerted antagonistic effect on the immune suppression caused by heterologous anti-lymphocyte serum, anti-sRBC serum and cortisone which are currently supposed to suppress the antigen reactive cells.