Clonal expansion of myelin basic protein-reactive T cells in patients with multiple sclerosis: Restricted T cell receptor V gene rearrangements and CDR3 sequence

Myelin basic protein (MBP)-reactive T cells are thought to play an important role in the pathogenesis of multiple sclerosis (MS). In some patients with MS, these autoreactive T cells display a limited heterogeneity in their epitope recognition and T cell receptor (TCR) variable (V) gene usage. These individual-dependent properties of MBP-reactive T cells have led to the speculation that they may represent clonal expansion in vivo in some MS patients. In the present study, 51 MBP-reactive T cell clones derived from patients with MS and healthy individuals were examined for their epitope recognition and the TCR Vα and Vβ gene rearrangements. The V gene junctional region sequences of identified α and β genes were further analyzed to probe their clonal origins, as the sequences are unique for individual clones. Our data showed that 26 clones derived from nine patients with MS shared a predominant reactivity to the immunodominant regions of MBP, 84–102, 110–129 and 143–168, and used various TCR Vα and Vβ rearrangements. The V gene usage of the clones was restricted to certain Vα Vβ combination(s) in a given MS patient, but varied among different patients. The sequence analysis revealed that the clones generated from a given patient shared a limited or a single junctional region sequence pattern(s), indicating their oligoclonal or monoclonal origin(s). In contrast, 25 MBP-reactive T cell clones derived from normal individuals exhibited unfocused epitope recognition and V gene usage. Thus, the limited heterogeneity of MBP-reactive T cells in their structural and functional charactertistics reflects their clonal expansion in vivo in some patients with MS.     

Job satisfaction and occupational stress among general practitioners — a pilot study

Questionnaires assessing levels of job satisfaction and possible sources of stress were distributed to a random sample of general practitioners in the Greater Manchester area. The highest levels of job satisfaction were reported for `intrinsic' job factors such as freedom to choose method of working, amount of responsibility and amount of variety, rather than `extrinsic' factors such as rate of pay and hours of work. The main causes of stress appeared to be interruptions of various kinds. Factor analysis revealed four major sources of stress: interruptions; emotional involvement; administrative workload and work/home interface; and routine medical work. Of these, all but routine medical work were associated with job satisfaction. It is concluded that the major sources of stress for the general practitioner are not medical, but social.     

A simple and inexpensive metabolic cage for mice

An inexpensive and easily constructed metabolic cage for mice is presented. This apparatus can reliably monitor food and fluid consumption, as well as urine and fecal output, in a relatively non-intrusive manner.     

The Development of the Pathologic Changes of Alzheimer''s Disease and Senile Dementia in Patients with Down''s Syndrome

Senile plaques, neurofibrillary change and granulovacuolar degeneration characterize Alzheimer's disease (presenile dementia) and senile dementia and are also seen in the aged human brain. The development of these lesions was studied in 13 patients with Down's syndrome, ages 12 to 65, with the purpose of defining similarities and dissimilarities, if any, between their morphologies in these four conditions. Evaluation by light and, when applied, electron microscopy established apparent identities. The findings suggest that Down's syndrome, with its partially characterized genotypic and phenotypic abnormalities, is an appropriate model for the study of the pathogenesis of these lesions.     

Mechanisms of respiration and phosphorylation in Ascaris muscle mitochondria

In Ascaris muscle mitochondria the major respiratory chain-linked phosphorylation activity is accomplished by a NADH-linked reduction of fumarate to succinate. Oxygen can also be employed as a terminal electron acceptor via a cyanide- and salicyl-hydroxamate-resistant terminal oxidase. As in fumarate-dependent electron transport this process appears to be coupled to energy conservation at phosphorylation site I. The branchpoint from which electrons are taken from the main respiratory chain to either the alternative oxidase or fumarate reductase is likely to be on the oxygen side of the NADH dehydrogenase segment.Malate and succinate are the only substrates which appreciably support respiration in the mitochondrion of the nematode. Regardless of the presence or absence of oxygen malate is utilized by an oxidation-reduction reaction resulting in the formation of pyruvate, acetate, succinate, propionate and CO₂. In addition, aerobically, hydrogen peroxide is formed as the product of oxygen reduction. Succinate accumulation was found to be significantly higher in the anaerobic as compared to the aerobic incubation mixtures. This effect was accompanied by an increase in anaerobic malate consumption. ATP generation and the formation of pyruvate, acetate and propionate were found to be similar in the presence and absence of oxygen.In malate-supported respiration of intact Ascaris mitochondria reducing equivalents (NADH) are produced exclusively through pyruvate and acetate formation. These enzymatic reactions are functionally coupled to the electron transport-linked reductions of fumarate to succinate and oxygen to hydrogen peroxide, respectively. In accordance with the position of the redox potentials of the fumarate/succinate and O₂/H₂O₂ couples, anaerobic and aerobic respiration was found to be associated with relatively low energy conservation efficiencies. Thus one molecule of ATP was conserved per 2e^? transferred to fumarate or oxygen, respectively. No evidence could be obtained for a significant activity of energy conservation sites II and III and electron transfer through the alternative oxidase pathway was shown not to be coupled to phosphorylation.     

Human peripheral Null lymphocytes I. Isolation,immunological and functional characterization

A Null lymphocyte-enriched population was isolated from human peripheral blood of healthy donors using a combination of Ficoll density gradient centrifugation followed by elimination of mononuclear phagocytes, passage through Ig-anti-Ig columns and sedimentation of E rosettes. After each separation step lymphocyte fractions were examined for morphology, cell surface markers, mitogen responsiveness and effector functions in antibody-dependent (ADCC) and spontaneous cellular cytotoxicity (SCMC) reactions against an allogeneic melanoma cell line. The final Null lymphocyte preparation was recovered at a rate of 1 to 3 % from the population passed through Ig-anti-Ig columns (fraction FFF-C). The marker analysis revealed over 99 %of surface Ig-negative lymphoid cells; 50 to 60 % of these cells were ‘real Null’ cells lacking immunological cell surface markers, 7 % formed EA, 13 % EAC and 24 %E rosettes. Regarding the mitogen responses, passage through Ig-anti-Ig columns drastically reduced concanavalin A (Con A), pokeweed mitogen (PWM)and tuberculin (PPD) responses, whereas the phytohemagglutinin (PHA) response was reduced in absolute counts but not in the stimulation index. Compared to the T cell-enriched lymphocyte fraction, the Null cells showed significantly diminished proliferative responses to PHA and Con A and slightly increased reactivity to PWM and PPD. Although depleted of high affinity Fc receptor lymphocytes, the Null cell fractions exhibited good ADCC and SCMC activities being about 4 to 6 times higher than in the T cell fraction.     

Interaction of Angiotensin with Disseminated Intravascular Coagulation: A Possible Mechanism in the Genesis of Acute Renal Failure

Because of the importance of the renin-angiotensin system in renal homeostatic mechanisms, the effect of angiotensin administration upon disseminated intravascular coagulation has been studied in rabbits. An infusion of angiotensin II (0.1 μg/kg/min for 2 hours) produced neither histologic abnormalities in the kidneys nor an elevation of creatinine. After an infusion of thrombin (2.0 units/kg/min for 2 hours) only 3 of 10 rabbits, when sacrified 24 hours later, showed histologic lesions comprised of occasional fibrin thrombi and foci of tubular necrosis. Creatinine levels did not rise. In contrast, the combination of angiotensin and thrombin resulted in renal lesions in 12 of 14 animals. Four had frank cortical necrosis, while combinations of tubular necrosis, glomerular thrombosis and segmental glomerular infarction occurred in the others, together with elevated creatinine levels. Blockade of α-adrenoreceptors with phenoxybenzamine in 12 animals did not prevent either these histologic changes or creatinine elevation, showing that the effect of angiotensin was independent of α-adrenoreceptor stimulation. The synergistic interaction between angiotensin and disseminated intravascular coagulation was not explained by differences in the consumption of plasma fibrinogen but apparently was due to localization of fibrin thrombi within glomerular capillaries by the vasomotor actions of angiotensin, as has previously been shown to occur with α-adrenoreceptor simulation. Such a mechanism might contribute to renal glomerular deposition of fibrin in acute ischemic renal failure.     

Tacrine restores cholinergic nicotinic receptors and glucose metabolism in alzheimer patients as visualized by positron emission tomography

Three patients with Alzheimer's disease, a 68-year-old woman with mild dementia and 2 men (aged 64 and 72 years) with moderate dementia were treated orally with the cholinesterase inhibitor tacrine (tetrahydroaminoacridine), 80 mg daily, for several months. The patients were investigated using positron emission tomography (PET) prior to, and after 3 weeks and 3 months of treatment. The PET studies involved a multi-tracer system consisting of [¹⁸F]-fluoro-deoxy-glucose (¹⁸F-FDG) (tracer for glucose metabolism); ¹¹C-butanol (cerebral blood flow) and (S)(−)- and (R)(+)-[N-¹¹C-methyl]-nicotine (nicotinic receptors; cholinergic neural activity). Tacrine treatment increased the uptake of ¹¹C-nicotine to the brain. Significant reduced difference in uptake between the two enantiomers (S)(−)- and (R)(+)¹¹C-nicotine was observed in the frontal and temporal cortices after tacrine treatment in all three patients. The kinetic analysis indicated increased binding of (S)(−)¹¹C-nicotine in brain compatible with a restoration of nicotinic cholinergic receptors. The most pronounced effect was observed after 3 weeks and 3 months treatment in the patient with mild dementia. An increase in cerebral glucose utilization was found in the 68-year-old patient with mild dementia but also slightly in the 64-year-old man with moderate dementia when treated with tacrine for 3 months. Tacrine administration did not affect cerebral blood flow. The PET data obtained after 3 weeks of tacrine treatment was paralleled by improvement in neuropsychological performance. This study shows in vivo by PET neurochemical effects induced in brain by treatment with tacrine to Alzheimer patients. Intervention with tacrine in the early course of the disease might be necessary for clinical improvement.     

Use of soluble recombinant TNF receptor to improve detection of TNF secretion in cultures of tumor infiltrating lymphocytes.

The accurate quantitation of picogram amounts of TNF is possible by ELISA and is useful in many areas of biomedical research, including studies of TNF release in vitro by stimulated lymphocytes and macrophages, and of serum levels in patients with cancer and sepsis. However, we show in this report that the detection of recombinant TNF standards by ELISA falls over time with incubation at 37 degrees C, and is further decreased when incubated with tumor infiltrating lymphocytes (TIL), making accurate quantitation difficult. We demonstrate that the soluble dimeric form of the TNF receptor can prevent this decrease, both in the presence and absence of TIL. In contrast, the soluble monomeric TNF receptor was much less effective in preventing this decrease. In addition, the dimeric but not the monomeric TNF receptor was found to inhibit bioactivity of TNF as measured by L929 cytotoxicity. The dimeric TNF receptor does not interfere with the detection of recombinant TNF standards by ELISA, and entirely stabilizes TNF levels incubated over 48 h at 37 degrees C in the presence and absence of TIL. This protection is specific, and the TNF receptor does not stabilize interferon-gamma. The dimeric form of the soluble TNF receptor has proven useful in detecting TNF released by TIL transduced with the TNF cDNA that are currently being used in studies of the gene therapy of cancer with TIL. The dimeric TNF receptor may also prove useful in the accurate quantitation of TNF released by stimulated lymphocytes and macrophages in vitro, and in the quantitation of serum TNF levels in patients.