The immunologic self.

Immunology has been referred to as the science of self/non-self discrimination. Who or what is the "I" of which we often speak? It has been identified with body, brain, soul, and memories. There has been a diversity of ways to characterize the self, including denying its importance and existence, and to date no satisfactory conceptualization has been achieved. This article addresses the concept of self from an immunological perspective, a perspective that offers insights into the philosophic issues associated with the concept of self and personal identity. The article proposes that the immunological and the psychological are two aspects of one and the same self, offers criteria for an adequate conceptualization of the self, and discusses the philosophical and psychological implications of the immunological views of self. It concludes that we must move away from a simplistic, exclusively inward or mentalistic conception of the self.

Circulating brain-reactive autoantibodies and behavioral deficits in the MRL model of CNS lupus.

Brain-reactive autoantibodies (BRAA) are hypothesized to play a role in the neuropsychiatric manifestations that accompany systemic lupus erythematosus (SLE). The present study tests the proposed relation between circulating BRAA and behavioral deficits in lupus-prone MRL/lpr mice. Two age-matched cohorts born at different times were used to test the relationship in the context of altered disease severity. Significant correlations between autoimmunity and behavior were detected in both cohorts. These results are the first to report correlations between behavior and autoantibodies to integral membrane proteins of brain, supporting the hypothesis that BRAA contribute to the behavioral dysfunction seen in lupus.

Plasma cytokines and chemokines in primary graft dysfunction post-lung transplantation.

Primary graft dysfunction (PGD) after lung transplantation causes significant morbidity and mortality. We aimed to determine the role of cytokines and chemokines in PGD. This is a multicenter case-control study of PGD in humans. A Luminex analysis was performed to determine plasma levels of 25 chemokines and cytokines before and at 6, 24, 48 and 72 h following allograft reperfusion in 25 cases (grade 3 PGD) and 25 controls (grade 0 PGD). Biomarker profiles were evaluated using a multivariable logistic regression and generalized estimating equations. PGD cases had higher levels of monocyte chemotactic protein-1 (MCP-1)/chemokine CC motif ligand 2 (CCL2) and interferon (IFN)-inducible protein (IP-10)/chemokine CXC motif ligand 10 (CXCL10) (both p < 0.05), suggesting recruitment of monocytes and effector T cells in PGD. In addition, PGD cases had lower levels of interleukin (IL-13) (p = 0.05) and higher levels of IL-2R (p = 0.05). Proinflammatory cytokines, including tumor necrosis factor (TNF)-alpha, and IFN-gamma decreased to very low levels after transplant in both PGD cases and controls, exhibiting no differences between the two groups. These findings were independent of clinical variables including diagnosis in multivariable analyses, but may be affected by cardiopulmonary bypass. Profound injury in clinical PGD is distinguished by the upregulation of selected chemokine pathways, which may useful for the prediction or early detection of PGD if confirmed in future studies.

B and T cells in the brains of autoimmune mice.

Systemic lupus erythematosus (SLE) is an autoimmune disorder that can involve the central nervous system (CNS). Recently, we reported the presence of autoantibodies bound to the brain tissue of murine models of lupus; MRL/lpr and BXSB. We postulated that the source of these autoantibodies was in part due to in situ production, caused by the entry of B and T cells. Frozen brain sections of MRL/lpr and BXSB at 1 and 4 months of age were stained for CD3 (T cells) and CD19 (B cells) markers using an immunofluorescent antibody binding assay. Confocal fluorescence microscopy showed both CD3(+) and CD19(+) cells at 4 months of age only in MRL/lpr mice. There were no lymphocytes seen in the other autoimmune model, BXSB. Results suggest a difference in the mechanisms by which autoantibodies access the brain in these two autoimmune models of lupus.

Immunoglobulin binding to brain in autoimmune mice.

Brain-reactive autoantibodies (BRAA) are thought to play an important role in central nervous system (CNS) manifestations of systemic lupus erythematosus (SLE). Previous studies have shown the existence of BRAA in human and murine SLE. This study was undertaken to establish and characterize the presence of autoantibody binding to brain of autoimmune mice. Laser confocal microscopy was performed on frozen brain sections to detect the presence of immunoglobulin (Ig) in the brain of MRL/lpr and BXSB mice and compare that to control strains of MRL/mp and C57BL/6 mice. There was a dramatic increase in fluorescence in the brains of MRL/lpr and BXSB at 4 months of age. There was little or no Ig detected in the brains of control mice. This increase in presence of Ig in the autoimmune mouse brain was paralleled by an increase in the serum titers of BRAA and anti-DNA autoantibodies as determined by ELISA. These studies provide another link between the existence of brain-reactive autoantibodies and altered CNS functioning.

Silver ion microplates for immunoassays.

Microplate wells can be coated with silver ions using glutaraldehyde as a spacer molecule and thiourea as a complexing ligand. Microwells containing surface silver ions are shown to immobilize biotin-labeled horseradish peroxidase (HRP) in active form, while showing very little affinity for the unlabeled enzyme. These plates can also immobilize biotin-labeled antibodies that exhibit bioactivity after immobilization. Silver ions are needed for the complexation of the biotinylated enzyme or antibody because microwells modified to contain surface amine or thiourea molecules do not immobilize appreciable amounts of the labeled proteins. A maximum surface coverage for biotin-labeled HRP of 40 ng/cm2 and an immobilization binding constant of Km = 8 x 10(9)/M are determined from serial dilutions in a microplate. Detection of as little as 6.7 fmol HRP is achieved using antibodies immobilized on the silver ion-modified microplates. Active antibody surface densities were estimated to be between 130 and 260 nm2/antibody molecule. Background binding of HRP to the modified silver ion microplates was very low, allowing for reasonably accurate detection between 10(-14) and 10(-11) mol HRP.

A comparison of silver ion to streptavidin coated microplates.

Direct comparisons are made between covalently linked streptavidin and silver ion coated microplates. Both coatings can immobilize biotinylated molecules. Silver ion coated microplate wells can immobilize 1.8 times higher amounts of biotin labeled horseradish peroxidase. The quantitation range and capacity for the capture of horseradish peroxidase using biotin labeled horseradish peroxidase are also greater for silver ion coated microplates. Approximately twice as many anti-horseradish peroxidase antibodies can be immobilized per well using silver ion coated microplates. Higher capacities are presumed to be due to the smaller footprint of silver ions as compared to streptavidin. A direct comparison between the two coatings for a beta-galactosidase ELISA showed that while the silver ion coated microplates gave higher readings, the streptavidin coated microplates exhibited smaller well-to-well variation. However, higher well to well variation for the silver microplates is attributed to the high density of anti-beta-galactosidase antibodies on the microplates and the weak binding of clone GAL-13 to beta-galactosidase, rather than the silver coating. These studies suggest silver ion coated microplates are a desirable alternative to streptavidin plates for quantitative immunoassays.

The immune system can affect learning: chronic immune complex disease in a rat model.

Evidence is presented that the immune system can affect central nervous system functioning, leading to changes in learning. Immune complex disease is induced in rats and their behavior tested using a Lashley maze. Significant differences in behavior were found between the animals with high disease activity and those with low disease activity and the non-disease controls. These changes were not due to uremia and are most likely due to the immune response. There is some evidence immune complex deposits in the choroid plexus may play some role, but not the sole or major role in the behavioral changes. This provides a model by which immunologic processes can cause neuropsychiatric manifestations in autoimmune diseases like lupus, as well as showing that immune processes can affect behavioral functioning.

Characterization of murine brain-reactive monoclonal IgG autoantibodies.

A diversity of brain-reactive autoantibodies (BRAA) is found in the sera and cerebrospinal fluid in elevated amounts in systemic lupus erythematosus (SLE) and correlates with some CNS manifestations of both human and murine SLE. In order to test the hypothesis that BRAA mediate the CNA manifestations, we are developing a library of brain-reactive monoclonal autoantibodies (BRMA) from autoimmune mice for characterization. We have recently reported on the production of BRMA of the IgM class from unimmunized autoimmune mice. In the current study we extend our investigations and report on the production and characterization of two BRMA of the IgG class. These antibodies react against integral brain membrane antigens of approximately 58, 32, and 30 kDa by immunoblot. Reactivity to such antigens is also found in a majority of the autoimmune murine serum samples tested, but not in nonautoimmune mice. These IgG BRMA show reactivity to cell bodies of the cerebral cortex, hippocampus, and hypothalamus of murine brain but not to fiber tracts. They also react with an integral thymus membrane antigen, but not to antigens of other tissues tested. Because of their properties, BRMA such as those characterized here are likely to be of pathogenic significance in CNS involvement in SLE.

Nanometer spatial resolution achieved in hard x-ray imaging and Laue diffraction experiments.

Tapered glass capillaries have successfully condensed hard x-ray beams to ultrasmall dimensions providing unprecedented spatial resolution for the characterization of materials. A spatial resolution of 50 nanometers was obtained while imaging a lithographically prepared gold pattern with x-rays in the energy range of 5 to 8 kiloelectron volts. This is the highest resolution scanning x-ray image made to date with hard x-rays. With a beam 360 nanometers in diameter, Laue diffraction was observed from the smallest sample volume ever probed by x-ray diffraction, 5 x 10(-3) cubic micrometers.

Effects of peripheral cytokine injections on multiple unit activity in the anterior hypothalamic area of the mouse.

The brain and the immune system interact in a bidirectional manner. This study on neuroimmune circuitry investigated the hypothesis that circulating cytokines act as messengers in the communication from the immune system to the brain and that the anterior hypothalamus is an integral part of this pathway. It was predicted that, following tail vein injections in mice, the cytokines interleukin-1 beta (IL-1 beta), interleukin-2 (IL-2), and interleukin-6 (IL-6) would alter electrical activity in the anterior hypothalamus. Differential electrodes were used to record multiple unit activity, at 5-min intervals, immediately before and for 60 min following injections. IL-1 beta and IL-2 decreased activity in the lateral margin of the anterior hypothalamus. IL-2-responsive neurons seemed to be localized to this area, while IL-1 beta-sensitive neurons showed a larger area of distribution. A significant response following IL-6 administration was not demonstrated. The observed changes in electrical activity support the concept of the hypothalamus as a sensory integration site for the immune system. Alternative models for the mechanism of cytokine signaling are discussed.

Anti-brain antibodies in the sera of rheumatoid arthritis patients: relation to disease activity and psychological status.

Auto-antibodies reactive with brain are known to occur in various immunological disorders. Their significance with respect to disease activity, neurological manifestations, or psychological status is, however, not known. In previous studies it was found that there were antibodies reactive with brain in the sera of patients with rheumatoid arthritis (RA). The present study sought to confirm and extend our previous findings and determine the significance of these antibodies with respect to disease activity. It also investigated a possible connection between psychological function in RA and these auto-antibodies. Sera drawn from 14 RA patients, at bi-weekly intervals over 3 months, showed the presence of antibodies against transmembrane proteins from murine brain. These antibodies correlated positively with rheumatoid factor and joint swelling. There was a trend toward a correlation between depression and daily mood scores and the number of antibodies reactive with brain. In addition, a correlation between cognitive coping styles in RA patients and auto-antibodies was found. There was also evidence for a sub-population of pathogenic antibodies.

Brain reactive monoclonal auto-antibodies: production and characterization.

To determine the role of auto-antibodies in the pathogenesis of neuropsychiatric manifestations of systemic lupus erythematosus (NP-SLE), it will be necessary to characterize the diversity of auto-antibodies that exist. This can be done by producing a library of monoclonal, brain-reactive auto-antibodies. From such a library the antigens to which the antibodies bind, and whether there are any interesting relations between these antigens, can be determined. Behavioral effects can also be investigated. Toward these ends, brain-reactive monoclonal auto-antibodies (BRMA) were produced. The production and characterization of two monoclonal antibodies is presented in this study.

Detection of brain-reactive autoantibodies in the sera of autoimmune mice using ELISA.

There are considerable problems with developing an assay to detect the often small quantities of autoantibodies which react against antigens in a heterogeneous and complex mixture from a source such as brain. An indirect enzyme-linked immunosorbent assay (ELISA) has been developed which can detect naturally occurring autoantibodies in serum that are reactive with integral brain membrane antigens. Sera were collected from autoimmune BXSB and NZB mice and non-autoimmune C57BL/6 mice at various ages and were assayed for the presence of brain-reactive autoantibodies (BRAAs). It is shown that this technique provides a highly sensitive, specific, and rapid assay for detecting BRAAs in serum. It shows that integral membrane antigens from whole brain can be isolated and used to detect and quantitate antibodies in the sera of autoimmune and non-autoimmune mice. The data also confirm studies, using different techniques, showing higher levels of autoantibodies to brain in autoimmune as compared to non-autoimmune mice. There are numerous potential applications for this ELISA, such as in rapidly screening large numbers of samples of biological fluids, tracking autoimmune disease progression over time, detecting small quantities of antibody against brain antigens, and as an assay system for investigating the role of BRAAs in the pathogenesis of immune mediated CNS disease.

Brain specific autoantibodies in murine models of systemic lupus erythematosus.

Autoantibodies which bind to integral membrane proteins of brain were tested for their ability to bind to cross-reactive antigens on non-neural tissue. Both brain specific autoantibodies and antibodies which bind to cross-reactive antigens were found. There were two types of brain reactive autoantibodies which could not be adsorbed by non-neural tissue. One type was adsorbable by crude cell membrane preparations of brain. The second type was reactive against integral membrane proteins of brain, but not adsorbable by any of the crude membrane preparations tested. Autoantibodies of the first type reacted against integral membrane proteins with apparent molecular weights of 75, 70, 62, 50, 27, 24 and 20 kDa, as determined by gel electrophoresis and immunoblotting. As in previous studies, a diversity of brain reactive autoantibodies were found. The greatest numbers and strongest banding patterns were seen in the autoimmune strains of mice. The non-autoimmune strain displayed these autoantibodies at much lower levels. These results are the first to find brain specific autoantibodies, from autoimmune mice, against integral brain membrane antigens. The data support the idea that there is a sub-population of brain reactive autoantibodies which are involved in the pathogenesis of neuropsychiatric manifestations in immunologic disorders, particularly systemic lupus erythematosus.

Characterization of brain-reactive autoantibodies in murine models of systemic lupus erythematosus.

Using the Western blot technique we analyzed the sera of five strains of mice that develop a disease like systemic lupus erythematosus (SLE), along with two normal strains, for their binding specificities against isolated mouse integral brain membrane proteins. This report describes the distribution and frequency of the more than 200 brain-reactive autoantibodies in the 126 animals tested and verifies the hypothesis of diversity in anti-brain antibodies produced during autoimmune conditions such as SLE. These results emphasize the importance of characterizing the brain-reactive autoantibodies in the sera or cerebrospinal fluid of SLE patients with central nervous system involvement.

Identification of autoantibody reactive integral brain membrane antigens. A two-dimensional analysis.

A procedure to identify murine autoantibody reactive, integral membrane antigens of brain is described. This involved the isolation of integral membrane antigens from whole brain followed by two-dimensional (2D) gel electrophoretic separation and transfer to nitrocellulose (NC) membrane. A 2D 'map' was then constructed by an effective total protein staining procedure. This 'map' was subsequently compared with similar blots that were reacted with autoimmune sera, stained for immunoglobulin and the reactive antigens identified. Advantages of this procedure include economy in the reagents and sera used, not having to use radioactive substances and much shorter working time. This technique will permit the identification of brain reactive autoantibodies.

Characterization of cell surface antigens on the adrenergic neuroblastoma clone A2(1).

Neuroblastoma cell lines have been extensively used to identify the presence of brain reactive autoantibodies in the sera of patients with systemic lupus erythematosus (SLE) who have neuropsychiatric involvement and in the animal models (murine) of this disorder. In this study, a characterization of murine neuroblastoma cell surface antigens, from the adrenergic A2(1) cell line, have indicated both similarities and differences with the cell surface antigens of normal mouse brain. It has also shown that some of these antigens are nervous system specific, whereas others are not. These data indicate that a more precise definition of the antigens on the surface of neuroblastoma cells, with which anti-brain autoantibodies react, is necessary for an understanding of the neuropsychiatric manifestations associated with autoimmune diseases such as SLE.