Lower levels of glutathione in the brains of secondary progressive multiple sclerosis patients measured by 1H magnetic resonance chemical shift imaging at 3 T.

BACKGROUND: Disability levels for patients with secondary progressive multiple sclerosis (SPMS) often worsen despite a stable MRI T(2) lesion burden. The presence of oxidative stress in the absence of measurable inflammation could help explain this phenomenon. In this study, the assessment of an in vivo marker of oxidative stress, cerebral glutathione (GSH), using magnetic resonance chemical shift imaging (CSI) is described, and GSH levels were compared in patients with SPMS and healthy controls. OBJECTIVE: To assess whether GSH, a key antioxidant in the brain, is lower in the SPMS patients compared to matched controls. METHODS: Seventeen patients with SPMS (Expanded Disability Status Scale=4.0-7.0; length of MS diagnosis=19.4 ± 7 years) and 17 age- and gender-matched healthy controls were studied. GSH levels were measured in the fronto-parietal regions of the brain using a specially designed magnetic resonance spectroscopy technique, CSI of GSH, at 3T. RESULTS: The levels of GSH were lower for SPMS patients than for controls, the largest reduction (18.5%) being in the frontal region (p=0.001). CONCLUSION: The lower GSH levels in these patients indicate the presence of oxidative stress in SPMS. This process could be at least partially responsible for ongoing functional decline in SPMS.

Cognitive impairment in patients with multiple sclerosis: association with the APOE gene and promoter polymorphisms.

BACKGROUND: Studies examining the epsilon4 allele of the APOE gene as a factor affecting the severity of multiple sclerosis (MS) have yielded conflicting results. The focus of these studies on physical disability to the neglect of cognitive impairment is surprising in light of the associations between the epsilon4 allele and other dementia conditions. Only two studies examine the relationship between the epsilon4 allele and cognitive impairment. METHODS: A neuropsychological test battery was administered to 263 MS patients, and their current disability status was evaluated. Genotypes were determined for APOE epsilon and for two promoter region polymorphisms (-219 G/T and -491 A/T). RESULTS: Although effects were generally weak, female patients with the -491 AA genotype had a later age of disease onset, lower disability scores, and somewhat higher scores on the cognitive battery. Male patients with the epsilon2 allele had lower disability and higher scores on the cognitive battery. The epsilon4 allele was not related to physical disability, and there was no difference between epsilon4+ and epsilon4--patients in overall cognitive performance. However, when patients with severe cognitive impairment were identified, a greater proportion (52%) of these patients had the epsilon4 allele than those in the unimpaired group (27%). CONCLUSION: An association with the epsilon4 allele was evident in this study, but only in cases of severe cognitive impairment.

Depression during exacerbations in multiple sclerosis: the importance of uncertainty.

The following correlates of depression were examined in a sample of 166 patients with clinically definite relapsing-remitting (n=140) or secondary progressive (n=26) multiple sclerosis: (a) the present state of the patients' illness (i.e., whether or not they were currently experiencing an exacerbation of their symptoms); (b) their level of uncertainty concerning their illness; and (c) their strategies for coping with their illness. A current exacerbation in symptoms, greater uncertainty of illness, and greater use of emotion-centered forms of coping were all related to depression. Multivariate analyses revealed that uncertainty of illness played a pivotal role as a mediating variable. Exacerbations in illness appeared to heighten patients' levels of uncertainty, and it was largely through this heightened uncertainty that the increases in depression came about.

The relationship between disability and depression in multiple sclerosis: the role of uncertainty, coping, and hope.

The relationship between disability and depression was studied in 188 patients with clinically definite multiple sclerosis (MS). Patients were administered the Zung Self-Rating Depression Scale, Ways of Coping, Uncertainty of Illness Scale, and Hope Scale during their regular clinic appointments. Their current level of disability was rated by the attending physician using the Expanded Disability Status Scale. Even when the depression measure was corrected for items overlapping with other symptoms or consequences of MS, depression was correlated with disability. Depression was also correlated with an array of psychological variables, including uncertainty concerning ones illness, hope, and the use of various emotion-centered, though not problem-centered coping strategies. Multiple regression analyses revealed that none of these psychological correlates mediated or moderated the relationship between disability and depression. Instead, disability, uncertainty, hope, and emotion-centered coping were significant independent predictors of depression, together accounting for approximately 40% of the variance in patients' self-reported depression. The relationship between disability and depression in MS is usually interpreted as evidence that depression is psychogenic and reactive to the demands and limitations of this disease. The demonstration that this relationship is not diminished when an array of potentially intervening psychological variables are included in the analysis raises questions concerning the validity of this interpretation.

Fatigue in multiple sclerosis: relationship to depression, disability, and disease pattern.

In order to investigate the associations between fatigue and depression, disability, and disease subtype, 207 individuals with clinically definite Multiple Sclerosis (MS) were administered the Fatigue Severity Scale and the Zung Self-rating Depression Scale during a regular clinic appointment. Their current level of disability was established using the Expanded Disability Status Scale. Fatigue and depression were highly correlated (r=0.58), even when the depression measure was corrected for items overlapping with fatigue and other symptoms or consequences of MS (r=0.44). Fatigue and disability were also correlated (r=0.33). Multiple regression revealed that both depressed mood and disability were significant predictors of fatigue, together accounting for approximately 23% of the variance in patients' self-reported fatigue. The combined groups of primary progressive (n=45) and secondary progressive patients (n=25) appeared to have higher fatigue scores than relapsing-remitting patients (n=137). However, an analysis of covariance revealed that this apparent difference was in fact attributable almost exclusively to differences in disability among the three subtypes of MS. Other reports of differences in fatigue between subtypes of MS should be re-examined in light of this finding.

Stress and coping in multiple sclerosis: exacerbation, remission and chronic subgroups.

Individuals with multiple sclerosis were asked to identify stressors they had experienced over a 6-month period and the coping mechanisms employed to deal with those stressors. Subjects who reported currently experiencing an exacerbation of symptoms were compared with those in remission or in the chronic phase of their illness. Of the 61 subjects who completed the Hassles Scale, Uplifts Scale, and Ways of Coping, those classified as being in an exacerbation phase of MS were found to have significantly higher hassles scores than those in the chronic phase. A significant difference was also found in terms of a factor labeled 'passive avoidant and aggressive coping' derived through a factor analysis of the Ways of Coping subscales. Subjects in the exacerbation subgroup had higher scores on this coping factor than those in the chronic subgroup. The difference in hassles scores remained significant after between-group differences in length of illness and reported number of symptoms were controlled through an analysis of covariance, although the difference in coping fell short of significance in this covariance analysis.

Theorizing about nurses' work lives: the personal and professional aftermath of living with healthcare 'reform'.

In this paper we discuss the impact of healthcare 'reform' on nurses' personal and professional lives. Using a thematic analysis, we interviewed 38 nurses in Nova Scotia, Canada regarding their experiences of job displacement, inability to find full-time employment and job losses. Their stories reflect how they lived day by day and the effects this had on their children, partners, friends and leisure, as well as their financial burdens. We theorize about the relationship between nurses' work and women's work, and particularly about women working in unstable conditions and the impact on their lives and that of the clients with whom they work.

A cross-cultural study of reactivation of posttraumatic stress disorder symptoms: American and Cambodian psychophysiological response to viewing traumatic video scenes.

A physiological hyperarousal state, which can be reactivated by traumatic stimuli, occurs frequently in patients with posttraumatic stress disorder (PTSD). The goals of this study were to determine whether physiological hyperarousal measured by increased heart rate is a specific response to reminders of a patient's own traumatic events or a more generalized hyperarousal state. Five brief videotape scenes of traumatic events (hurricane, auto accident, Cambodian refugee camp, domestic violence, and Vietnam War) were shown to two patient groups with PTSD (Vietnam veterans and Cambodian refugees) and three control groups (Vietnam veterans, Cambodian refugees, and nonpatient Americans). Observations of subjects' behavior, subjective ratings of distress, and heart rate change were recorded and evaluated. The results indicated that Cambodians with PTSD had the most reactions as measured by behavior and heart rate changes. These tended to occur during all scenes, not just the specific Cambodian scene, indicating a general nonspecific arousal. The Vietnam veterans had the fewest changes implying an inhibition of response. The control groups were intermediate in physiological response. The response in PTSD patients to reactivation scenes is complex and probably relates to type and degree of trauma, as well as to culture.

Sensitization with clozapine: beyond the dopamine hypothesis.

Clozapine elicits dose-dependent myoclonic jerks in partially restrained rats and induces paroxysmal electroencephalographic changes, myoclonus, and convulsive seizures in a small but significant percentage of patients. With the hypothesis that the central excitatory effects of clozapine may relate to the unique therapeutic activity of this agent, rats were administered repeated alternate day or weekly very low dose (1 mg/kg) injections of clozapine in an attempt to induce the central excitatory effect through sensitization or kindling. Although initial administrations of this dose elicited no motor response or other behavioral change, repeated administration of the same low dose on either the alternate-day or weekly schedule caused increasing numbers of myoclonic seizure-like jerks (MJs) reaching 75-110 MJs/hour by the sixth clozapine injection. Clozapine-sensitized animals exhibited a significantly different pattern of early gene expression in two subcortical sites compared with vehicle-treated controls. These findings may have importance for the treatment of psychosis.

TCR vaccines for active immunotherapy of T cell malignancies.

We have developed a TCR-based vaccine approach for the treatment of T cell malignancies. TCR genes were isolated from C6VL, a T cell tumor of C57BL/Ka origin. The transmembrane encoding domains of the TCR genes were replaced by sequences encoding for phosphatidylinositol-linked cell surface expression. A high expressing cell line was produced by transfection and amplification of the TCR genes. Large quantities of soluble native C6VL TCR-alphabeta protein was obtained by treating the high-expressing cells with a specific phospholipase and purifying the released TCR by affinity chromatography. Following vaccination with the TCR linked to keyhole limpet hemocyanin, specific anti-TCR humoral responses were induced. Both the carrier protein and an adjuvant were required for optimal responses. Hyperimmune serum from vaccinated mice reacted specifically with C6VL cells, and the immunizations did not affect the TCR repertoire, which suggested that the immune response was Id specific. The TCR-vaccinated mice were specifically protected from a lethal number of C6VL tumor cells. B cell-deficient mice were not protected by TCR vaccinations. Similarly, TCR-immunized mice depleted of CD8+ cells prior to tumor challenge were not protected. Thus, C6VL TCR vaccine effectively stimulated tumor protection, which depends on the presence of both B cells and CD8+ T cells.

Kindling with clozapine: behavioral and molecular consequences.

Clozapine is an 'atypical' neuroleptic that improves symptoms of many patients with schizophrenia whose illness is resistant to treatment with other neuroleptics. Unlike the 'typical neuroleptics (chlorpromazine, haloperidol), clozapine does not induce extrapyramidal symptoms such as Parkinsonism and tardive dyskinesia in humans or catalepsy in the rat. However, clozapine frequently causes epileptiform EEG changes and causes seizures in 3-5% of patients treated with this drug in therapeutic doses. Clozapine also induces dose dependent myoclonus in the partially restrained rat. In the experiments reported here, partially restrained rats were administered repeated alternate day or weekly low, fixed doses of clozapine (1 mg/kg). This dose initially caused no behavioral change. Following the third and subsequent administrations, the same dose elicited an increasing number of myoclonic seizure-like jerks reaching 140/h following the 15th injection in rats receiving the same low dose of clozapine on alternate days and 160/h following the 9th injection in animals that received the same dose once weekly. These effects are consistent with kindling, i.e. a progressive increase of brain excitability following repeated administration of a fixed subconvulsive dose of an excitatory agent. Clozapine kindled animals exhibited a significantly different pattern of early gene expression in ventral tegmental area, origin of the mesolimbic-mesocortical dopamine system and in the anterior thalamic nuclei, compared with saline treated controls subjected to exactly the same recording conditions. The evidence of central nervous system excitation with clozapine may be important to the unique therapeutic effect of this atypical antipsychotic in the treatment of symptoms, especially the deficit symptoms, of schizophrenia.

Lymphocyte subsets and interleukin-2 receptors in autistic children.

Blood samples were obtained from 10 male autistic children ages 7-15 years and 10 age-matched, male, healthy controls. Lymphocyte subsets (helper-inducer, suppressor-cytotoxic, total T, and total B cells) were enumerated using monoclonal antibodies and flow cytometry. Bound and soluble interleukin-2 receptors were assayed in unstimulated blood samples and in cell cultures following 72-hour stimulation with phytohemagglutinin. The children with autism had a lower percentage of helper-inducer cells and a lower helper:suppressor ratio, with both measures inversely related to the severity of autistic symptoms (r = - .56 and - .68, respectively). A lower percentage of lymphocytes expressing bound interleukin-2 receptors following mitogenic stimulation was also noted, and this too was inversely related to the severity of autistic symptoms.

Kinetics of the reaction of a myelin basic protein peptide with soluble IAu.

The kinetics of formation and dissociation of IAu-peptide complexes have been examined in the absence of detergent, using a glycosylphosphatidylinositol (GPI)-linked form of IAu. The GPI-linked form contains a lipid membrane anchor which can be specifically cleaved by phosphatidylinositol-specific phospholipase C to yield a water-soluble form of IAu. We find rapid binding of the myelin basic protein (MBP) peptide analogue Ac(1-14)A4C15 to IAu, as well as rapid dissociation of IAu-MBP peptide complexes at neutral pH in the absence of detergent. The reaction kinetics of the water-soluble and detergent-solubilized complexes are the same to within experiment error. In the presence of this MBP peptide, Ac(1-14)A4C15, cells transfected with native IAu as well as cells transfected with a GPI-linked form of IAu are functional in stimulating T-helper hybridoma cells.

Myelin basic protein peptide complexes with the class II MHC molecules I-Au and I-Ak form and dissociate rapidly at neutral pH.

The acetylated N-terminal peptide of myelin basic protein (MBP) is the immunodominant T cell epitope in the induction of experimental autoimmune encephalomyelitis in the I-Au- and I-Ak-expressing mouse strains. We used a direct binding assay to examine the kinetics of binding and dissociation of a series of MBP peptide analogues with the affinity-purified class II MHC molecules I-Au and I-Ak. We observe much faster in vitro rates of binding and dissociation than has been reported previously for other immunogenic peptides at neutral pH. The kinetics also reveal inactivation of the peptide-free class II MHC molecules. These results are consistent with previously proposed mechanisms for tolerance escape and autoimmune disease.

Clozapine and seizures.

Epileptiform EEG changes, myoclonus, and seizures are reported in some patients treated with clozapine. Although these are undesirable side effects, the excitation of specific neuronal networks by clozapine and other neuroleptics may be important for the therapeutic effect of this class of agents. In these experiments, intraperitoneal clozapine 2-16 mg/kg produced dose-related myoclonic jerks in partially restrained rats. Paroxysmal slow waves and spike activity were recorded from implanted electrodes in amygdala, hippocampus, and cortex following higher doses of clozapine, but the EEG abnormalities were not correlated with the myoclonic jerks. Single doses of chlorpromazine (8 and 16 mg/kg) rarely produced myoclonic jerks but provoked generalized tonic seizures in two animals preceded by multiple myoclonic jerks in one. Myoclonus and seizures reflect increased excitability of the central nervous system. It is possible that clozapine and other neuroleptics exert a therapeutic effect by increasing excitability in critical subcortical areas of the brain.

Formation of phenol and thiocatechol metabolites from bromobenzene premercapturic acids through pyridoxal phosphate-dependent C-S lyase activity.

When N-acetyl-S-(2-hydroxy-4-bromocyclohexa-3,5-dienyl)-L-cystein e (4-S-premercapturic acid) and N-acetyl-S-(2-hydroxy-5-bromocyclohexa-3,5-dienyl)-L-cystein e (3-S-premercapturic acid) were used as substrates in incubations with Hartley guinea pig kidney 9000 g supernatant preparations, the major products were the corresponding S-(2-hydroxy-4-bromocyclohexa-3,5-dienyl)-L-cysteine and S-(2-hydroxy-5-bromocyclohexa-3,5-dienyl)-L-cysteine. At the end of the incubation period, the percentage recovery of these N-deacetylate cysteine conjugates accounted for 77 +/- 2% of the substrates, 3-S- and 4-S-premercapturic acids. Removal of the N-acetyl group from premercapturic acids to form the corresponding cysteine conjugates by kidney N-deacetylase(s) showed no preference with respect to the 3-S- and 4-S-positional isomeric conjugates. Other metabolites which included the known sulfur-containing acids, mercaptolactate and mercaptoacetate, were also detected. 3- and 4-Bromophenol and 3- and 4-bromothioanisole were also formed. The addition of pyridoxal-5'-phosphate to the kidney incubation mixture resulted in a 5-fold increase in the formation of phenols and thioanisoles, along with four different isomeric O- and S-methylated 3-S-and 4-S-bromothiocatechols and two S-methylated 3-S- and 4-S-bromodihydrobenzene thiolols. This result indicated that a pyridoxal phosphate-dependent C-S lyase(s) is involved in the formation of both phenol and thiophenolic metabolites from S-(2-hydroxy-4-bromocyclohexa-3,5-dienyl)-L-cysteine and S-(2-hydroxy-5-bromocyclohexa-3,5-dienyl)-L-cysteine. Guinea pig liver 9000 g supernatant preparations did not N-deacetylate the 3-S- and 4-S-premercapturic acids to the same extent as kidney preparations, and this may account for decreased conversion of 3-S- and 4-S-premercapturic acids to 3- and 4-bromophenol and to thiophenolic products by liver preparations.

Speed of scanning in primary memory in persons with dementia of the Alzheimer type.

The nature of the search of primary memory by persons with a presumptive diagnosis of mild Alzheimer's disease (DAT) was compared with that of normal elderly and young persons using the Sternberg (1966) paradigm. DAT subjects evidenced a substantial deficit in the speed of scanning in primary memory and a deficit in at least one other stage of processing. These findings are consistent with the hypothesis that the decline in memory performance evidenced by persons with DAT may be attributable to an increase in the time requirements of mnemonic processing.

Immune variables, depression, and plasma cortisol over time in suddenly bereaved parents.

Bereavement has been associated with increased morbidity and mortality. The authors monitored a previously unstudied population, parents who had experienced sudden death of a formerly healthy child, for immunological changes, plasma cortisol level, and depression. Nine bereaved parents were matched case-for-case with nonbereaved controls and concurrently monitored 2, 4, 6, and 8 months post-bereavement. The bereaved parents showed significantly decreased T-suppressor cells, significantly increased T-helper cells, and depression compared with controls, but no difference in cortisol levels. The bereaved also showed nonsignificant but elevated blastogenesis in mitogen-stimulated cells. These changes persisted throughout the first 8 months of bereavement.

Subtractive hybridization system using single-stranded phagemids with directional inserts.

We describe a subtractive hybridization protocol which is designed to permit subtractions between cDNA libraries. The method uses single-stranded phagemids with directional inserts as both the driver and the target. We modified the M13 phagemid vector pBluescript for the directional cDNA cloning and subtractive hybridization. Two simplified methods for efficient construction of directional cDNA libraries are also described. Using a model system, we found that one round of subtractive hybridization results in a 5,000-fold specific subtraction of abundant molecules. We used two methods to quantify the efficiency and verify the specificity of the subtraction. In order to obtain these subtraction efficiencies, it was necessary to develop a method to purify the single-stranded DNA to homogeneity. The single-stranded purification involved using potassium iodide (KI) density centrifugation, restriction endonuclease digestion and phenol extraction in the presence of magnesium. We describe the several advantages of using directional inserts for the subtraction procedure.