Model Of Autoimmune Hemolytic Anemia Research Articles

7 A nanobody-based complement inhibitor targeting complement component 2 reduces hemolysis in a complement humanized mouse model of autoimmune hemolytic anemia

7 A nanobody-based complement inhibitor targeting complement component 2 reduces hemolysis in a complement humanized mouse model of autoimmune hemolytic anemia

A nanobody-based complement inhibitor targeting complement component 2 reduces hemolysis in a complement humanized mouse model of autoimmune hemolytic anemia

C2 is an attractive therapeutic target for many complement-mediated diseases. We developed Nab1B10, a new anti-C2 nanobody that potently and selectively inhibits both the classical and lectin pathways of complement activation. Mechanistically, Nab1B10 binds to the C2a portion of C2 and inhibits the assembly of C3 convertase C4b2a. Nab1B10 cross-reacts with monkey but not rodent C2 and inhibits classical pathway-mediated hemolysis. Using a new complement humanized mouse model of autoimmune hemolytic anemia (AIHA), we demonstrated that Nab1B10 abolished classical pathway complement activation-mediated hemolysis in vivo. We also developed C2-neutralizing bi- and tetra-valent antibodies based on Nab1B10 and found these antibodies significantly more potent than the other anti-C2 monoclonal antibody that is already in clinical trials. These data suggest that these novel C2-neutralizing nanobodies could be further developed as new therapeutics for many complement-mediated diseases, in which pathogenesis is dependent on the classical and/or lectin pathway of complement activation.

C5aR activation in the absence of C5a: A new disease mechanism of autoimmune hemolytic anemia in mice.

IgG Fc receptors (FcγRs) and the C5a anaphylatoxin receptor (C5aR) were identified as key regulators of type II autoimmune injury in mice. However, and with respect to C5aR, the relative importance of C5a for IgG autoantibody-induced cellular destruction remained unclear. Using an experimental model of autoimmune hemolytic anemia (AIHA), we here report marked differences in the development of AIHA between mice lacking C5aR and C5-deficient (Hc0 ) strain, indicating a limited role of C5 in this type of C5aR-regulated disease. Ex-vivo-analyses of liver homogenates from anemic Hc0 mice demonstrate C5a-independent C5aR activation, upregulation of FcγR expression and amplification of erythrophagocytosis by macrophages. As assessed by pharmacological inhibition studies, targeting of C5aR, but not of C5, is effective in treating experimental AIHA. Collectively, these results define a previously unrecognized disease mechanism of C5aR activation in AIHA that does not necessarily involve C5 and C5a.

Binding of Autoantibodies and Apoptotic Response in Erythroid Cells in the Mouse Model of Autoimmune Hemolytic Anemia

Binding of Autoantibodies and Apoptotic Response in Erythroid Cells in the Mouse Model of Autoimmune Hemolytic Anemia

Inhibitors of Bruton's Tyrosine Kinase Reduce Anti-Red Blood Cell Response in a Murine Model of Autoimmune Hemolytic Anemia

Autoimmune hemolytic anemia (AIHA) is an antibody-mediated condition where the mainstays of treatment are corticosteroids and immunosuppressive therapies. This leads to substantial toxicity in patients with refractory or chronic disease and thus there is a need for improved therapies. BTK is a critical kinase in the B-cell receptor signaling cascade and we hypothesized that inhibition would decrease autoantibody responses. Supporting this, CLL patients taking ibrutinib, a first generation BTK inhibitor, experience very low rates of AIHA (Leukemia 2016). Based on this rationale we evaluated two BTK inhibitors, ibrutinib (ibr) and acalabrutinib (acal), in mouse models of AIHA to validate BTK as a target in this disease.Young C3H/B6 F1 mice were treated with acal, ibr, or vehicle (veh) via drinking water for 2 weeks then injected intraperitoneally with washed rat red blood cells (RBCs) weekly for 4 weeks using the method of Playfair and Marshall-Clarke (Nature New Biology 1973) to induce a response against autologous RBCs starting 14 days after initial injection and peaking days 28-35. Drug treatment was continued throughout the study and using this delivery method BTK occupancy was 99.1% for acal, 96.1% for ibr, and 0% for veh. Quantitation of immunoglobulin on mouse RBCs was performed by a direct enzyme-linked anti-globulin test (DELAT) and by flow cytometry for surface immunoglobulin as a confirmatory method. Results from DELAT are shown in figure 1. A repeated measures model was applied to log-transformed data. On day 7, prior to autoimmune response, there were no statistically significant differences between treatment groups. However, on day 21 significantly less autoantibody was detected on RBCs in the acal group vs. both veh (p=0.001) and ibr (p=0.008); the ibr group showed a trend toward less autoantibody compared to veh but this did not reach statistical significance (p=0.542). This relationship between treatments held true at days 42 and 56 (acal vs. vehicle, p=<0.001 and p=0.001; acal vs. ibr, p=0.008 and p=0.004; ibr vs. veh, p=0.542 and p=0.542). These results were confirmed by flow cytometry. Hematocrit tests were performed by capillary tube centrifugation with no mice developing anemia and no differences between treatment groups excluding rapid RBC turnover as a cause of reduced immunoglobulin on RBCs.As ibrutinib has been shown to alter macrophage function, we used a passive model to determine if BTK inhibitors changed clearance of antibody coated RBCs from circulation. After two weeks of treatment with acal (n=13), ibr (n=12), or veh (n=12) mice were injected with a single dose of anti-RBC antibody (anti-ter-119) and hematocrit was measured daily for 7 days. Median hematocrit values at baseline for the three groups were 53.7 (SD=1.3), 52.8 (SD=1.4), and 52.8 (SD=0.9) for acal, ibr, and veh respectively and fell to a low of 18.4 (SD=2.7), 18.0 (SD=3.2), and 15.8 (SD=1.9) on day 3 before recovering. A repeated measures model was applied to the data and no significant differences between drug treatment groups were found (p=0.177), indicating no effect of BTK inhibition on RBC clearance.To examine anti-rat RBC antibody response washed rat RBCs were incubated with mouse plasma and analyzed by flow cytometry. Treatment groups were compared at day 42 after initial rat RBC injection. An analysis of variance (ANOVA) model was applied to log-transformed data. There was significantly less anti-rat RBC antibody in the acal group vs veh (p=0.008). In addition, there was decreased, albeit not statistically significant, antibody production with ibr compared to veh and acal compared to ibr (p=0.193 and p=0.102, respectively). This is the same trend as for autoantibody production and argues that BTK inhibition may reduce antibody responses.In summary, treatment with acalabrutinib significantly reduced anti-RBC autoantibody response in our model compared to vehicle and ibrutinib, and autoantibody was consistently decreased in ibrutinib treated mice compared to vehicle although this did not reach statistical significance. The differences between inhibitors may be due to potency of BTK inhibition or effects on other targets of ibrutinib which is a less specific drug. This mechanism is under active investigation. These results support BTK as a potential therapeutic target in AIHA and the further development of acalabrutinib for this indication. [Display omitted] DisclosuresWoyach:Karyopharm: Research Funding; Morphosys: Research Funding; Acerta: Research Funding.

Identification of Stages of Erythroid Differentiation in Bone Marrow and Erythrocyte Subpopulations in Blood Circulation that Are Preferentially Lost in Autoimmune Hemolytic Anemia in Mouse.

Repeated weekly injections of rat erythrocytes produced autoimmune hemolytic anemia (AIHA) in C57BL/6 mice after 5–6 weeks. Using the double in vivo biotinylation (DIB) technique, recently developed in our laboratory, turnover of erythrocyte cohorts of different age groups during AIHA was monitored. Results indicate a significant decline in the proportion of reticulocytes, young and intermediate age groups of erythrocytes, but a significant increase in the proportion of old erythrocytes in blood circulation. Binding of the autoantibody was relatively higher to the young erythrocytes and higher levels of intracellular reactive oxygen species (ROS) were also seen in these cells. Erythropoietic activity in the bone marrows and the spleen of AIHA induced mice was examined by monitoring the relative proportion of erythroid cells at various stages of differentiation in these organs. Cells at different stages of differentiation were enumerated flow cytometrically by double staining with anti-Ter119 and anti-transferrin receptor (CD71) monoclonal antibodies. Erythroid cells in bone marrow declined significantly in AIHA induced mice, erythroblast C being most affected (50% decline). Erythroblast C also recorded high intracellular ROS level along with increased levels of membrane-bound autoantibody. No such decline was observed in spleen. A model of AIHA has been proposed indicating that binding of autoantibodies may not be a sufficient condition for destruction of erythroid cells in bone marrow and in blood circulation. Last stage of erythropoietic differentiation in bone marrow and early stages of erythrocytes in blood circulation are specifically susceptible to removal in AIHA.

Deficient NLRP3 and AIM2 Inflammasome Function in Autoimmune NZB Mice.

Inflammasomes are protein complexes that promote caspase activation, resulting in processing of IL-1β and cell death, in response to infection and cellular stresses. Inflammasomes have been anticipated to contribute to autoimmunity. The New Zealand Black (NZB) mouse develops anti-erythrocyte Abs and is a model of autoimmune hemolytic anemia. These mice also develop anti-nuclear Abs typical of lupus. In this article, we show that NZB macrophages have deficient inflammasome responses to a DNA virus and fungal infection. Absent in melanoma 2 (AIM2) inflammasome responses are compromised in NZB by high expression of the AIM 2 antagonist protein p202, and consequently NZB cells had low IL-1β output in response to both transfected DNA and mouse CMV infection. Surprisingly, we also found that a second inflammasome system, mediated by the NLR family, pyrin domain containing 3 (NLRP3) initiating protein, was completely lacking in NZB cells. This was due to a point mutation in an intron of the Nlrp3 gene in NZB mice, which generates a novel splice acceptor site. This leads to incorporation of a pseudoexon with a premature stop codon. The lack of full-length NLRP3 protein results in NZB being effectively null for Nlrp3, with no production of bioactive IL-1β in response to NLRP3 stimuli, including infection with Candida albicans. Thus, this autoimmune strain harbors two inflammasome deficiencies, mediated through quite distinct mechanisms. We hypothesize that the inflammasome deficiencies in NZB alter the interaction of the host with both microflora and pathogens, promoting prolonged production of cytokines that contribute to development of autoantibodies.

The idiotypic network in the regulation of autoimmunity: Theoretical and experimental studies

The regulation of autoimmunity is a key issue in fundamental immunology. Despite outstanding achievements on this front, we currently have more questions than answers. The idea of an immune network as a regulatory mechanism is quite attractive, since it enables us to explain the selectivity (specificity), and moreover the clonality, of the regulation. Nevertheless it remains unclear how this mysterious network of immune cells is organized, how it operates, and how it exerts control over autoimmunity. This article presents an attempt to understand how the immune network functions and how it controls autoreactivity. We present a mathematical model of the immune network that is based on principles of immune network organization and function that we arrived at from a survey of the available literature. To test the principles on which the mathematical model is based, we studied the model and compared the different responses to antigen that it generated with the results obtained from experimental studies of immune response. The modeled kinetics of idiotype and anti-idiotype in response to the administration of antigen are in good agreement with the experimental kinetics of idiotypic and anti-idiotypic antibodies. To obtain evidence of the existence of idiotypic mechanisms for regulating autoimmunity, we studied a mathematical model containing autoclones and compared the model results with data from experimental studies in a model of autoimmune hemolytic anemia in mice. Because the results from the theoretical and the experimental studies coincide, there is justification to conclude that autoreactive lymphocytes are normal components of the immune network within which they are regulated. We discuss a possible molecular/cellular mechanism for negative control of autoreactive cells as affected by anti-idiotypic antibodies.

Lack of Galactosylation Enhances the Pathogenic Activity of IgG1 but Not IgG2a Anti-Erythrocyte Autoantibodies

IgG bears asparagine-linked oligosaccharide side chains in the Fc region. Variations in their extent of galactosylation and sialylation could modulate IgG Fc-dependent effector functions, and hence Ab activity. However, it has not yet been clarified whether the pathogenic potential of IgG autoantibodies is consistently enhanced by the absence of galactose residues per se or the lack of terminal sialylation, which is dependent on galactosylation. Moreover, it remains to be defined whether the increased pathogenicity of agalactosylated IgG is related to activation of the complement pathway by mannose-binding lectin, as suggested by in vitro studies. Using a murine model of autoimmune hemolytic anemia, we defined the contribution of galactosylation or sialylation to the pathogenic activity of IgG1 and IgG2a anti-erythrocyte class-switch variants of 34-3C monoclonal autoantibody. We generated their degalactosylated or highly sialylated glycovariants and compared their pathogenic effects with those of highly galactosylated or desialylated counterparts. Our results demonstrated that lack of galactosylation, but not sialylation, enhanced the pathogenic activity of 34-3C IgG1, but not IgG2a autoantibodies. Moreover, analysis of in vivo complement activation and of the pathogenic activity in mice deficient in C3 or IgG FcRs excluded the implication of mannose-binding lectin-mediated complement activation in the enhanced pathogenic effect of agalactosylated IgG1 anti-erythrocyte autoantibodies.

STIM2 is the essential STIM protein in non-SOCE calcium responses of macrophages in vitro and in vivo

Event Abstract Back to Event STIM2 is the essential STIM protein in non-SOCE calcium responses of macrophages in vitro and in vivo Georgios Sogkas1*, Shahzad Nawaz Syed2, Eduard Rau1, David Stegner3, Timo Vögtle3, Bernhard Nieswandt3, Reinhold Ernst Schmidt1 and Johannes Engelbert Gessner1* 1 Hannover Medical School, Clinical Immunology and Rheumatology, Germany 2 McMaster University, Faculty of health Sciences, Farncombe Family Digestive Health Research Institute, Canada 3 University Hospital and Rudolf Virchow Center, DFG Research Center for Experimental Biomedicine, University of Würzburg, Chair of Experimental Biomedicine, Germany Macrophage physiology critically depends on Ca2+ signalling. STIM isoforms are mediators of the most common mode of calcium influx in non-excitable cells, Store-Operated Ca2+ Entry (SOCE). Here, the relative dependence of key aspects of macrophage physiology on the mobilization of Ca2+ from the internal stores and/or the extracellular compartment was evaluated. Among the functions studied, cytokine responses to TLR ligands and chemokine elicited motility were mainly dependent on the mobilization of intracellular Ca2+. In contrast, FcγR-mediated phagocytosis, apart from internal stores, depends on Ca2+ influx and SOCE. STIM2, in addition to contributing in SOCE, is involved in the maintenance of the internal ER-store of Ca2+. The consequences of STIM2 deficiency in these Ca2+ dependent functions were characterized and contrasted with the effect of STIM1 deficiency. The SOCE dependent FcγR-mediated phagocytosis was reduced in case of STIM2 deficiency. However, the resulting defect was smaller as compared to the consequences of STIM1 deficiency. Moreover, genetic deletion of STIM2 had no effect on the phagocytosis-promoting function of C5a in vitro and in a model of autoimmune hemolytic anemia in vivo. Effective cytokine production downstream of TLRs and elicited motility, however, strictly depends on the presence of STIM2. In vivo, Stim2-/- mice exhibited milder LPS-induced sepsis and thioglycollate-elicited peritonitis. Interestingly, none of these SOCE-independent functions were reduced in case of STIM1 deficiency, suggesting their being specifically mediated by STIM2. Acknowledgements Funded by the DFG and the International HRBS PhD program in Molecular Medicine. Keywords: STIM1, STIM2, SOCE, Calcium, Macrophages, Fcgamma receptors, LPS, C5a, CCL2, MCP1, Chemotaxis, Migration Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Innate immunity Citation: Sogkas G, Syed S, Rau E, Stegner D, Vögtle T, Nieswandt B, Schmidt R and Gessner J (2013). STIM2 is the essential STIM protein in non-SOCE calcium responses of macrophages in vitro and in vivo. Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.00444 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 02 Apr 2013; Published Online: 22 Aug 2013. * Correspondence: Dr. Georgios Sogkas, Hannover Medical School, Clinical Immunology and Rheumatology, Hannover, Germany, sogkas.georgios@mh-hannover.de Prof. Johannes Engelbert Gessner, Hannover Medical School, Clinical Immunology and Rheumatology, Hannover, Germany, Gessner.Johannes@mh-hannover.de Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Georgios Sogkas Shahzad Nawaz Syed Eduard Rau David Stegner Timo Vögtle Bernhard Nieswandt Reinhold Ernst Schmidt Johannes Engelbert Gessner Google Georgios Sogkas Shahzad Nawaz Syed Eduard Rau David Stegner Timo Vögtle Bernhard Nieswandt Reinhold Ernst Schmidt Johannes Engelbert Gessner Google Scholar Georgios Sogkas Shahzad Nawaz Syed Eduard Rau David Stegner Timo Vögtle Bernhard Nieswandt Reinhold Ernst Schmidt Johannes Engelbert Gessner PubMed Georgios Sogkas Shahzad Nawaz Syed Eduard Rau David Stegner Timo Vögtle Bernhard Nieswandt Reinhold Ernst Schmidt Johannes Engelbert Gessner Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Evidence in favor of a role of idiotypic network in autoimmune hemolytic anemia induction: theoretical and experimental studies

Formation dynamics of antibodies to rat erythrocytes (REs) and auto-antibodies to mouse erythrocytes were studied in an experimental model of autoimmune hemolytic anemia (AHA) in mice. The experimental conditions of AHA were simulated in a mathematical model of an immune network. It was found that maximal production of auto-antibodies and antibodies to REs do not synchronize. Antiserum, obtained at the peak of auto-antibodies formation, competed with REs for bounding with antibodies. This represents proof that auto-antibodies to erythrocytes and antibodies to REs are an idiotype-anti-idiotypic pair. In the autoimmune reaction, the autoreactive clone, being anti-idiotypic, responded earlier than the clone reacting to the injected antigen. Comparison of autoimmune reaction kinetics in the mathematical model of an immune network with experimental dynamics of AHA shows them to be similar. So activation of the autoreactive clone to erythrocytes during experimental AHA in mice is mediated by idiotype-anti-idiotypic interactions with the clone reacting to REs.

Targeting T Cell-Specific Costimulators and Growth Factors in a Model of Autoimmune Hemolytic Anemia

Although it is established that failure of regulatory mechanisms underlies many autoimmune diseases, the stimuli that activate autoreactive lymphocytes remain poorly understood. Defining these stimuli will lead to therapeutic strategies for autoimmune diseases. IL-2-deficient mice develop spontaneous autoimmunity, because of a deficiency of regulatory T cells, and on the BALB/c background, they rapidly die from autoimmune hemolytic anemia. To define the importance of costimulatory pathways in various components of this autoimmune disorder, we first intercrossed IL-2-deficient mice with mice lacking CD28 or CD40L. Elimination of CD28 reduced the activation of autoreactive T cells and lymphoproliferation as well as production of autoantibodies, whereas elimination of CD40L reduced autoantibody production without affecting T cell expansion and accumulation. To examine the role of IL-7, we blocked IL-7R signaling with neutralizing Abs. This treatment inhibited the production of autoantibodies and the development of autoimmune hemolytic anemia. Together, these data indicate that specific costimulatory and cytokine signals are critical for the spontaneous autoantibody-mediated disease that develops in IL-2-deficient mice.

Critical role of galectin-3 in phagocytosis by macrophages.

Galectin-3 is a member of a large family of animal lectins. This protein is expressed abundantly by macrophages, but its function in this cell type is not well understood. We have studied the effect of galectin-3 gene targeting on phagocytosis, a major function of macrophages. Compared with wild-type macrophages, galectin-3-deficient (gal3-/-) cells exhibited reduced phagocytosis of IgG-opsonized erythrocytes and apoptotic thymocytes in vitro. In addition, gal3-/- mice showed attenuated phagocytic clearance of apoptotic thymocytes by peritoneal macrophages in vivo. These mice also exhibited reduced IgG-mediated phagocytosis of erythrocytes by Kupffer cells in a murine model of autoimmune hemolytic anemia. Additional experiments indicate that extracellular galectin-3 does not contribute appreciably to the phagocytosis-promoting function of this protein. Confocal microscopic analysis of macrophages containing phagocytosed erythrocytes revealed localization of galectin-3 in phagocytic cups and phagosomes. Furthermore, gal3-/- macrophages exhibited a lower degree of actin rearrangement upon Fcgamma receptor crosslinkage. These results indicate that galectin-3 contributes to macrophage phagocytosis through an intracellular mechanism. Thus, galectin-3 may play an important role in both innate and adaptive immunity by contributing to phagocytic clearance of microorganisms and apoptotic cells.

Critical role of galectin-3 in phagocytosis by macrophages

Galectin-3 is a member of a large family of animal lectins. This protein is expressed abundantly by macrophages, but its function in this cell type is not well understood. We have studied the effect of galectin-3 gene targeting on phagocytosis, a major function of macrophages. Compared with wild-type macrophages, galectin-3–deficient (gal3–/–) cells exhibited reduced phagocytosis of IgG-opsonized erythrocytes and apoptotic thymocytes in vitro. In addition, gal3–/– mice showed attenuated phagocytic clearance of apoptotic thymocytes by peritoneal macrophages in vivo. These mice also exhibited reduced IgG-mediated phagocytosis of erythrocytes by Kupffer cells in a murine model of autoimmune hemolytic anemia. Additional experiments indicate that extracellular galectin-3 does not contribute appreciably to the phagocytosis-promoting function of this protein. Confocal microscopic analysis of macrophages containing phagocytosed erythrocytes revealed localization of galectin-3 in phagocytic cups and phagosomes. Furthermore, gal3–/– macrophages exhibited a lower degree of actin rearrangement upon Fcγ receptor crosslinkage. These results indicate that galectin-3 contributes to macrophage phagocytosis through an intracellular mechanism. Thus, galectin-3 may play an important role in both innate and adaptive immunity by contributing to phagocytic clearance of microorganisms and apoptotic cells.

Experimental model of autoimmune hemolytic anemia induced in mice with levodopa by intraperitoneal injection or oral feeding

In this paper we describe a murine experimental model of autoimmune hemolytic anemia induced with multiple injections or oral feeding of levodopa. Strain A mice were intraperitoneally injected or fed with levodopa, at a dose equivalent to the one used in human therapy, and subsequently they developed cycles of IgM, IgG and IgA anti-mouse red blood cells (MRBC) autoantibody responses. Levodopa injection induced serum IgM and IgG anti-MRBC responses and levodopa feeding enhanced the serum anti-MRBC IgA response. The appearance of autoantibodies in the serum was followed by binding of the autoantibodies to mice erythrocytes and three phases of anemia. Red cell bound IgM and IgG autoantibodies were predominant in levodopa-injected mice whereas red cell bound IgA autoantibodies were predominant in levodopa-fed mice. The specificity of the serum IgA autoantibody was not restricted since it interacted with erythrocytes of various species.

Experimental model of autoimmune hemolytic anemia induced in mice with Levodopa

This paper describes an experimental model of autoimmune hemolytic anemia induced with multiple injections of levodopa. The serum antibodies were detected by indirect enzyme-linked immunosorbent assay (ELISa) and autoantibodies bound to syngeneic erythrocytes were detected by direct ELISA. A strain of mice injected with levodopa, at doses which are approximately equivalent to those used in human therapy, developed early and late cycles of IgM and IgG anti-mouse red blood cell (MRBC) autoantibody responses. These responses were followed by binding of IgM and IgG autoantibodies to the syngeneic erythrocytes and by two phases of anemia. Different strains of mice exhibited different sensitivities to Levodopa treatment, suggesting that genetic factors affect the anti-MRBC autoantibody response to levodopa. Indirect ELISA revealed that mice subjected to both X-irradiation (250 rad) and levodopa treatment had higher levels of IgG anti-MRBC autoantibodies than mice subjected to levodopa treatment alone, suggesting that a radiosensitive suppressor mechanism controls the autoantibody response. The autoantibodies induced with levodopa exhibited an unrestricted specificity and retained significant activity after absorption of MRBC. Serum autoantibodies were also detected by indirect ELISA in the sera of normal mice. This “natural” autoimmunity was mouse strain dependent and was possibly further influenced by environmental factors. The various implications of this model of autoimmune hemolytic anemia are discussed.

Clonal analysis of autoreactive B cells in a murine model of autoimmune hemolytic anemia

Hybridoma technology was used to examine the repertoire of autoantibody producing B cells in mice with autoimmune hemolytic anemia induced by injection of rat red blood cells (RBC). The results point to the importance of suppressor T cells (Ts) in both the induction as well as the maintenance of the self-specific B-cell repertoire at the clonal level. Thus when normal BALB/c mice were immunized to provoke a high autoantibody response, the hybrids generated were mainly (97%) cross-reactive with mouse RBC, whereas after immunization to elicit Ts and a low autoantibody response, hybrids were mainly (87%) rat RBC specific. In addition, when hybrids were generated from rat RBC immunized (CBA × B10A)F1 mice, in which Ts levels had been depleted during ontogeny, hybrids with “forbidden” purely anti-self (mouse RBC) specificity were detected.

Friend lymphatic leukemia virus-induced autoimmune hemolytic anemia with glomerulonephritis in the rat.

AbstractWKA/Mk rats injected with Friend lymphatic leukemia virus at birth developed a runt ing syndrome which was associated with a high incidence of hemolytic anemia. The anemia was characterized by the presence of incomplete anti‐erythrocyte autoantibodies detected by the direct antiglobulin test and the indirect enzyme‐augmented hemagglutination assay, and, in some cases, was followed by glomerulonephritis. This system in the rat may represent a new model of autoimmune hemolytic anemia in man.