Boris Sakic

Neurobehavioral alterations in autoimmune mice

Inbred MRL, NZB and BXSB strains of mice spontaneously develop a systemic, lupus-like autoimmune disease. The progress of autoimmunity is accompanied with a cascade of behavioral changes, most consistently observed in tasks reflective of emotional reactivity and the two-way avoidance learning task. Given the possibility that behavioral alterations may reflect a detrimental consequence of autoimmune-inflammatory processes and/or an adaptive response to chronic malaise, they are tentatively labeled as autoimmunity-associated behavioral syndrome (AABS). It is hypothesized that neuroactive immune factors (pro-inflammatory cytokines, brain-reactive antibodies) together with endocrine mediators (corticotropin-releasing factor, glucocorticoids) participate in the etiology of AABS. Since AABS develops natively, and has a considerable face and predictive validity, and since the principal pathway to autoimmunity is known, AABS may be a useful model for the study of CNS involvement in human autoimmune diseases and by extension, for testing autoimmune hypotheses of several mental disorders (major depression, schizophrenia, Alzheimers disease, autism and AIDS-related dementia).

Anti-NMDA receptor encephalitis. The disorder, the diagnosis and the immunobiology.

Anti-NMDAR encephalitis is a newly characterized syndrome with a progressive, predictable clinical course and the possibility of effective treatment. Accurate and timely diagnosis is critical to selection and implementation of treatments, and optimal patient outcomes. Outcomes are improved with early diagnosis via indirect immunofluorescence or cell-based assays, and the rapid and appropriate administration of immunosuppressant and anti-psychotic therapies. Three possible scenarios accounting for the immunopathogenesis of anti-NMDAR encephalitis are presented, with the most probable one being that of paraneoplastic autoimmunity. Future efforts in this disorder should focus on elucidating the mechanisms that contribute to initiation of this antibody response, as well as exploring the role of tumors, infectious triggers and immune-reactivation. Finally, accessible tools need to be developed that allow for reliable identification of specific antibody markers against synaptic proteins.

Reduced preference for sucrose in autoimmune mice: a possible role of interleukin-6.

In a continuous one-bottle sucrose intake test, 4-month-old autoimmune MRL-Ipr mice show a shift to the right along the X-axis of the concentration-intake function, compared to congenic MRL +/+ controls. Using a brief (60-min) and a continuous (48-h) two-bottle test, the present report examines potential factors that could account for the reduced responsiveness to a palatable stimulus. Study 1 examines whether preference for sucrose is associated with age, changes in food/water intake, or impaired renal function. Reduced preference for sucrose was observed in 5-6-week-old MRL-Ipr males, although food/water intake or blood creatinine levels did not differ from control values. Immunosuppressive treatment abolished this deficit, suggesting a role of immune factor(s). Study 2 tests the hypothesis that chronic upregulation of the neuroactive cytokine interleukin-6 (IL-6), reported to occur from 3 weeks of age in young MRL-Ipr mice, reduces preference for sucrose. Sustained administration of IL-6 was produced by infecting healthy MRL +/+, C3H.SW and Balb/C mice with adenovirus vector carrying cDNA for murine IL-6. This resulted in high serum IL-6 levels over 5 days, a rapid decline in preference for sucrose and low blood glucose levels. The results from Study 1 indicate that impaired sensitivity to sucrose in MRL-Ipr mice can be detected before autoimmune disease is florid in MRL-Ipr mice. The results from Study 2 are consistent with altered motivation/emotional states after infection, and point to sustained IL-6 production as an early mechanism in behavioral alterations during chronic autoimmune/inflammatory conditions.

Progressive atrophy of pyramidal neuron dendrites in autoimmune MRL-lpr mice

The autoimmune-prone MRL-lpr substrain of mice develop an autoimmunity-associated behavioral syndrome (AABS) which resembles in many respects the behavior of animals exposed to chronic stress. The present study examined whether these mice show changes in the morphology of neuronal dendrites, as found in animals exposed to chronic stress. A modified Golgi-Cox procedure was used to visualize the dendrites of pyramidal neurons in the parietal cortex and in the CA1 hippocampal field of 5-week and 14-week old MRL-lpr mice and MRL + / + controls. Reduced dendritic branching and length, and an up to 20% loss of dendritic spines were observed in parietal and hippocampal pyramidal neurons of MRL-lpr mice at both ages. In the parietal cortex, there was an age-dependent potentiation in the reduction of basilar, but not apical, dendrite branching and length, as well as in the loss of spines on basilar segments. Loss of spines in the hippocampus followed an age-related course for apical but not basilar dendrites. Moreover, compared to age-matched controls, brain weight was smaller in MRL-lpr mice at 14 but not 5 weeks of age. Considering that dendritic atrophy becomes more extensive when autoimmune disease is florid in MRL-lpr mice, it is proposed that immune/inflammatory factor(s) produce dendritic loss. Reduced dendritic complexity may represent, at least in part, a structural basis for the altered behavioral profile of MRL-lpr mice.

Neurodegeneration in autoimmune MRL-lpr mice as revealed by Fluoro Jade B staining

As in many humans suffering from lupus erythematosus, the development of systemic autoimmunity and inflammation in Fas-deficient MRL-lpr mice is accompanied by CNS dysfunction of unknown etiology. Experimental studies revealed infiltration of lymphoid cells into the choroid plexus, reduced neuronal complexity, retarded brain growth, and enlargement of cerebral ventricles. Moreover, an increased presence of cells with nicked-DNA (TUNEL+ cells) in the periventricular areas suggested accelerated apoptosis in brain cells of MRL-lpr mice. However, direct evidence that the dying cells were neurons was lacking. For this purpose, we presently use Fluoro-Jade B (FJB), a novel fluorescent dye which has high affinity for dying neurons (both apoptotic and necrotic). As expected, in comparison to the control groups, the brains of diseased, 5-month-old MRL-lpr mice showed increased numbers of FJB-positive (+) cells in cortical and periventricular regions. The FJB+ cells were significantly more numerous than TUNEL+ cells, and only approximately 7% co-localized with TUNEL. Immunostaining for CD4 and CD8 markers did not correlate with the number of FJB+ cells, suggesting that T-lymphocyte infiltration into the brain tissue is not a reliable predictor of neuronal demise. Conversely, indices of systemic autoimmunity (splenomegaly and high serum anti-nuclear antibody levels) were associated with increased FJB+ cell numbers in brains of autoimmune MRL-lpr mice, supporting the causal link between autoimmunity and neurodegeneration. Taken together, the above results suggest that factors other than T-cell infiltration and cell death mechanisms other than Fas-mediated apoptosis dominate neuronal degeneration in lupus-prone MRL-lpr mice.

Spatial learning during the course of autoimmune disease in MRL mice

The present study examines whether autoimmune MRL-lpr mice develop impairments in learning and memory that correlate with changing severity of lupus-like disease. MRL-lpr mice (n = 20) were tested in the Morris water-maze at 12, 14, 16 and 18 weeks of age. Age-matched controls were congenic MRL +/+ mice (n = 20) that develop the disease much later. Immune status was assessed by the presence of anti-nuclear antibodies (ANA), brain-reactive antibodies, proteinuria, and haematocrit. Learning rates and memory retention did not differ between the substrains, and did not correlate or deteriorate with advancing age and autoimmunity. However, the baseline performance level in autoimmune MRL-lpr mice was shifted, as evidenced by a consistently longer task-solving latencies. Thigmotaxic swimming (along the pool wall) was pronounced in the MRL-lpr group, and was associated with the observed difference in performance. The present study does not support the notion that learning/memory abilities of autoimmune MRL-lpr mice are impaired per se, but may support the hypothesis that the rapid progress of humoral autoimmunity affects the emotionality of lupus-prone mice.

Effect of cyclophosphamide on leukocytic infiltration in the brain of MRL/lpr mice

Neuropsychiatric manifestations are a poorly understood and potentially life-threatening complication of systemic lupus erythematosus (SLE). MRL/1pr mice spontaneously develop a lupus-like syndrome which is similar to the human disease in many respects, including behavioural abnormalities. Our previous findings indicated that the age at which infiltration of immune cells into the choroid plexus is first observed coincides with the appearance of behavioural dysfunction in MRL/1pr mice. This present study quantified leukocyte infiltration in relation to prolonged administration of cyclophosphamide (CY), a treatment effective in preventing some behavioural deficits. Compared to MRL + / + controls, saline-treated MRL/1pr mice had significantly more CD45-positive cells (leukocytes) and CD45R-positive (B) cells in the choroid plexus and in the brain parenchyma. A six week course of CY (100 mg/kg i.p.) significantly reduced the infiltration of CD45, but not of CD45R-positive cells into the choroid plexus of the MRL/1pr substrain. In addition, the presence of leukocytes correlated positively with measures on one behavioural test (floating in the forced swim test) but not on another test (novel object test). These findings suggest that CY treatment has a differential effect on the infiltration of leukocyte subtypes and strengthen the hypothesis that some abnormal behaviour in MRL/1pr mice may be related to the presence of immunocompetent cells in the brain.

Immunosuppressive treatment prevents behavioral deficit in autoimmune MRL-lpr mice.

MRL-lpr mice, which develop severe autoimmune disease, explore novel objects less well than do their congenic MRL(-)+/+ controls, in which symptoms of the disease are relatively mild. Moreover, diminished exploration in MRL-lpr mice correlates with the elevated titers of antinuclear antibodies (ANA) in their sera, suggesting that this behavioral deficit is caused by the autoimmune process. To test the hypothesis that autoimmunity affects behavior, in this study we examine whether treatment of the autoimmune process will reduce the difference in performance between the two MRL substrains in the novel-object test. Forty mice in each substrain were treated from 4 to 10 wk of age with the immunosuppressive drug, cyclophosphamide (100 mg/kg/wk, IP) or a saline vehicle. The immunosuppressive treatment reduced ANA titers to low levels and eliminated ANA production completely in 55% of MRL-lpr mice, suggesting an attenuation of the autoimmune process. In addition, treatment with cyclophosphamide, but not saline, abolished significant differences in exploration between the MRL-lpr and MRL +/+ groups, as measured by the latency to touch a novel object and the time spent exploring it. Thus, the present results suggest that a treatment which ameliorates autoimmune symptoms can concurrently remove the substrain difference in behavior. The effect of cyclophosphamide in the MRL-lpr group is believed to reflect the suppression of pathogenic immune factor(s) that alter behavior during the onset of autoimmune disease.

Neurotoxic properties of cerebrospinal fluid from behaviorally impaired autoimmune mice.

The chronic, lupus-like autoimmune disease in MRL-lpr mice is associated with leucocyte infiltration into the choroid plexus, brain cell death, and deficits in motivated behavior. The presence of lymphoid cells in the ventricular lumen and the increased number of TUNEL-positive cells in periventricular areas led to the hypothesis that immune cells enter into the cerebrospinal fluid (CSF) and induce primary neuronal damage in regions bordering the cerebral ventricles. Using an in vitro approach, we presently examine the possibility that CSF from autoimmune mice is neurotoxic and/or gliotoxic. The CSF and serum from diseased MRL-lpr mice, less symptomatic MRL +/+ controls, and healthy Swiss/Webster mice (non-autoimmune controls) were frozen until their effects on the viability of pyramidal neurons and astrocytes were assessed in a two-color fluorescence assay. Significant reduction in neuronal viability (in some cases as low as 67%) was observed in the co-cultures of hippocampal neurons and astrocytes incubated for 24 h with CSF from autoimmune MRL-lpr mice. The viability of astrocytes did not differ among the groups, and the CSF from autoimmune mice appeared more toxic than the serum. The behavior of MRL-lpr mice differed significantly from the control groups, as indicated by impaired exploration, reduced intake of palatable food, and excessive immobility in the forced swim test. The present results suggest that CSF from the behaviorally impaired lupus-prone mice is neurotoxic and are consistent with the hypothesis that neuroactive metabolites are produced intrathecally in neuropsychiatric lupus erythematosus.

Autoimmune and inflammatory mechanisms of CNS damage

Brain morphology and function are susceptible to various psysiological influences, including changes in the immune system. Inflammation and autoimmunity are two principal immunological responses that can compromise the function of multiple organs and tissues, including the central nervous system. The present article reviews clinical and experimental evidence pointing to structural brain damage induced by chronic autoimmune and/or inflammatory processes. Largely due to the vast complexity of neuroendocrine and immune systems, most of the principal pathogenic circuits are far from elucidated. In addition to summarizing the current knowledge, this article aims to highlight the importance of interdisciplinary research and combined efforts of physicians and scientists in revealing the intricate links between immunity and mental health.