Cellular and molecular aspects of the pathomechanism and therapy of murine experimental allergic encephalomyelitis

Experimental allergic encephalomyelitis (EAE) is an autoimmune disease of the central nervous system (CNS). Its immune mechanism is well understood at the cellular and molecular levels, which is herein reviewed. Susceptibility to EAE is under the control of the genes partially inside and partially outside the H-2 complex. There are two myelin constituents known to be encephalitogenic, myelin basic protein and proteolipid apoprotein. EAE is mediated by effector T cells sensitized to the encephalitogen. Effector T cells bear surface phenotypes of Lyt1+2-, L3T4+, and they are activated by the encephalitogen/self Ia complex or certain alloantigens and acquire encephalitogenic activity. By unknown homing mechanisms, the effector T cells invade the CNS and induce the target phase phenomena, which include Ia-antigen expression in the local tissue, activation of procoagulant activity, breakdown of the blood-brain barrier, and excretion of lymphokines which induce inflammation and demyelination, resulting in functional alteration. Possibility of specific immune therapy is postulated as a model for human autoimmune disease.     

Neurotrophic effect of hematopoietic cytokines on cholinergic and other neurons in vitro

We examined the effects of interleukin-3 (IL-3) and other hematopoietic cytokines on the neurotransmitters, neurite formation, and differentiation in cholinergic and other types of neurons. IL-3, granulocyte-macrophage colony-stimulating factor (GM-CSF), macrophage colony-stimulating factor, granulocyte colony-stimulating factor and erythropoietin (Epo) elevated choline acetyltransferase (ChAT) activity in septal cholinergic cell line SN6 as well as in primary cultured septal neurons without increasing protein contents of the cells. These effects were dose-dependent and the optimal doses were not different from those for blood cells. IL-3 had neurite-promoting activity but GM-CSF had no such effect. Both IL-3 and GM-CSF decreased intracellular acetylcholine concentration, and elevated glutamic acid decarboxylase and intracellular GABA in septal neuronal cultures. Epo elevated monoamines in PC12 cells. These effects are thought to result from direct action through their specific receptors in neurons, because (i) anti-IL-3-receptor antibody abolished the ChAT activity in septal neurons increased by IL-3; (ii) mRNA and immunoreactivity for β subunits of IL-3 receptors were expressed in septal cholinergic neurons and (iii) presence of receptos for GM-CSF and Epo in neurons has been reported. Our observation and others strongly support that neural-immune interactions are important not only in the defense mechanism in the nervous system but also in the development, differentiation and function of neurons.     

Neurotransmitter synthesis by SN6 cell lines,a family of hybrid cell lines of embryonic septal origin

Previously, we reported the presence of multiple neurotransmitters in subclones of SN6, a septal cholinergic hybrid cell line. To obtain information concerning the functionality of these transmitters, we measured transmitter contents, activities of transmitter-producing enzymes, and the effect of serum-free culture medium in two different batches (SN6.1.6 and SN6.10.2.2) and two subclones of the SN6 cell line (SN6.2a and SN6.1b). Except for SN6.1b, SN6 cell lines and subclones had basically the same neurotransmitter characteristics. Among the transmitters, only acetylcholine seemed to be functional. Monoamine oxidase was missing and activity of aromatic amino acid decarboxylase was diminished in SN6 cell lines. Even in serum-containing medium, SN6.1b had a more mature morphology than the other cell lines, and it contained choline acetyltransferase and acetylcholine but not tyrosine hydroxylase or catecholamines. Similar characteristics were acquired by the mother cell line in response to serum-free conditions. Thus, SN6.1b is the most mature of these central cholinergic neuronal cell lines, at least with regard to neurotransmitter profiles. ©1995 Wiley-Liss, Inc.     

Heterogeneous induction of 72-kDa heat shock protein (HSP72) in cultured mouse oligodendrocytes and astrocytes

The expression of 72-kDa heat shock protein (HSP72) in cultured mouse oligodendrocytes and astrocytes exposed to heat shock was investigated by double immunolabelling with anti-HSP72 monoclonal antibody (C92F3B-1) and antibodies against galactocerebroside (GalC) or glial fibrillary acidic protein (GFAP). After 3 h recovery from heat shock, an intermediate level of HSP72 immunolabelling was localized in the nucleolus and cytoplasm of astrocytes (less than 25%) and to a lesser extent in oligodendrocytes (less than 2%). After 8-48 h, HSP72 was expressed intensely in the cytoplasm and nuclear matrix of oligodendrocytes (20-40%), while weak/intermediate immunostaining was detectable in astrocytes (5-15%). The levels of HSP72 expressed in oligodendrocytes and astrocytes decreased around 72-120 h, but a few oligodendrocytes (4%) remained intensely immunolabelled. These results indicate that heat shock induces HSP72 in both oligodendrocytes and astrocytes. However, this response is heterogeneous.     

Analysis of T cell antigen receptors of myelin basic protein specific T cells in SJL/J mice demonstrates an {alpha} chain CDR3 motif associated with encephalitogenic T cells

Experimental autoimmune encephalomyelitis (EAE) is an animalautoimmune disease mediated by CD4⁺ T cells. Analysis of TCRexpression revealed that limited TCR elements (V^ß8.2,V2 or 4) were utilized by myelin basic protein (MBP) specificT cells In mice with H-2^U haplotype and Lewis rats. The usageof a particular ß chain complementarity determiningregion 3 (CDR3) motif has also been shown. However, It remainsunclear to what extent these observations can be extrapolated.Here we studied the TCR sequences of MBP 89–101/1-A^s specificT cell clones derived from SJL/J mice, using the polymerasechain reaction on reverse transcribed mRNA. Although the V_ßusage was less restricted than In H-2^U mice, they predominantlyutilized v_ß17a and expressed LGG or related motifsin the V_ß - D_ß - J_ß junctions.Furthermore, a single chain rearrangement between V1.1 andJBBM142 with no N region diversity was preferentially used.Concordantty, Immunization with a peptlde corresponding to the chain CDR3 was found to significantly alter the clinical courseof EAE. Comparison of the published TCR junctional regions demonstratesthat the CDR3 motifs (LGG in ß chain, CA*R*NY motifIn chains) are expressed by other encephalitogenic clones.Notably, the CA*R*NY was conserved in PL/J mice clones thatrecognize a distinct MBP–MHC determinant. It suggeststhat an antigen-independent mechanism may contribute to conservingthe a chain motif. The Implications of these observations arediscussed.     

Urinary homocysteic acid levels correlate with mini-mental state examination scores in Alzheimer's disease patients

Homocysteic acid (HA) has been suggested as a pathogen in a mouse model of Alzheimer's disease (AD), 3xTg-AD. However, it is not established whether HA is involved in humans. We investigated the relationship between urinary HA levels and Mini-Mental State Examination (MMSE) scores in AD patients (n = 70) and non-AD controls (n = 34). We found a positive, statistically significant relationship between the two variables (the urinary HA level and MMSE score) (r = 0.31, p = 0.0008, n = 70). This relationship was stronger in females than males (r = 0.43, p = 0.005, n = 44 in females; r = 0.48, p = 0.02, n = 22 in males). The urinary HA levels were significantly different in AD patients than controls (AD: 8.7 ± 7.5, n = 70; non-dementia control: 13.3 ± 9.4, n = 34, p < 0.01). In addition, aging and smoking were found as lowering factors for urinary HA levels. Our preliminary study showed a negative, statistically significant relationship between blood HA (micromole) and urine HA levels (r = -0.6, p = 0.007, n = 19), and between blood HA levels and MMSE scores (r = -0.79, p = 0.0000518, n = 19). On the basis of these results, we speculate that reduced urinary excretion induces elevated HA levels in blood, resulting in cognitive dysfunctions. This study also suggests that HA may be a candidate of neurotoxins for uremic encephalopathy. Since amyloid-β increases HA toxicity and HA is an agonist of N-methyl-D-aspartic acid (NMDA) receptor, we speculate that elevated blood HA affects the brain cognitive function through NMDA receptor-mediated toxicity in AD.     

Chronic stress differentially regulates glucocorticoid negative feedback response in rats

Exposure to chronic stress is thought to play an important role in the etiology of depression. In this disorder, a disrupted negative feedback response to exogenous glucocorticoids on cortisol secretion has been indicated. However, the regulation of glucocorticoid negative feedback by chronic stress is not fully understood. In the present study, we investigated the effects of chronic stress administered by water immersion and restraint (2 h/day) for four weeks on the glucocorticoid feedback in rats. In the acutely (one-time) stressed rats, the basal plasma corticosterone (CORT) level was markedly elevated, remained at high levels for 5 h after the termination of stress, and then decreased. In the chronically stressed rats, the CORT level was initially elevated similarly, but rapidly decreased at 2 h. In the dexamethasone (DEX) suppression test, the peak CORT level in response to stress was not suppressed by DEX in the acutely stressed rats, but was significantly suppressed in the chronically stressed rats. In contrast, the suppressive effects of DEX on the basal CORT secretion in naive rats were attenuated in the chronically stressed rats. In the chronically stressed hippocampus, which plays an important role in the regulation of the glucocorticoid feedback response, the binding of [3H]DEX was decreased and the increased response of activator protein-1 induced by acute stress was abolished. These results suggest that chronic stress induces a hypersuppressive state for induced CORT secretion in response to acute stress, which is caused by partial habituation, coping, and adaptation to the stressor, whereas it induces a hyposuppressive state for the basal CORT secretion, which is caused by glucocorticoid receptor downregulation. These mechanisms may be involved in the stress-induced neural abnormalities observed in depression.     

Prognostic value of mitotic metaphase rate in oesophageal cancer.

OBJECTIVE: To assess the prognostic value of the mitotic metaphase rate in patients with oesophageal cancer. DESIGN: Retrospective study. SETTING: University hospital, Japan. SUBJECTS: 41 patients with oesophageal cancer. INTERVENTIONS: We calculated the ratio of mitotic metaphase to anaphase cells among tumour cells in sections stained with haematoxylin and eosin, a ratio that shows the status of mitotic metaphase-anaphase transition. The DNA ploidy pattern was examined by flow cytometry. MAIN OUTCOME MEASURE: Correlation between survival and mitotic metaphase rate. RESULTS: A high mitotic metaphase rate was correlated with vascular invasion and is expected to be a useful prognostic factor. DNA diploidy combined with a low rate was an independent favorable prognostic factor. CONCLUSION: Mitotic metaphase rate is a useful index of malignant potential, independent of DNA ploidy. It can distinguish high malignant potential from low in a diploid tumour, which has a poor prognosis that is equal to that of the aneuploidy tumour.