The binding and processing of monoclonal human IgG1 by cells of a human macrophage-like cell line (U937)

The goal of these experiments was to assess the relationship between the binding and processing of IgG by Fc-receptor-bearing cells. Cells of the U937 human macrophage-like cell line were incubated with 125I- labeled monomers, dimers, oligomers (composed of 2-4 IgG1 subunits), and HP (heavy polymers composed of 5 or more subunits per polymer) of monoclonal human IgG1 in vitro. Binding was assessed by spinning cells through a layer of phthalate oils. Internalization of IgG1 was assessed by quantitating residual binding to cells after surface-bound IgG was removed by a brief treatment with a solution containing 0.25 M acetic acid and 0.5 M sodium chloride. Catabolism was assessed by measuring the release of radioactive fragments of IgG1, which were not precipitated by 10% trichloroacetic acid. Unstimulated U937 bound about 10,000 molecules per cell of IgG1 monomer, with an equilibrium binding constant (Ka) of 5 X 10(8) M-1. After stimulation with a conditioned medium in vitro, binding per cell was increased 3-7--fold, and the Ka was decreased 2-4--fold. Both unstimulated and stimulated cells internalized and catabolized labeled IgG1 HP, but stimulated cells internalized and digested much more IgG1 HP per cell than unstimulated cells. Both monomers and dimers of IgG1 were internalized and degraded very slowly by stimulated cells, even though both preparations readily bound to cells. In contrast, oligomers and (to an even greater extent) IgG1 HP were internalized and degraded much more rapidly. Internalization of IgG1 HP was markedly inhibited by incubation at 4 degrees C, but not by incubation with a variety of metabolic inhibitors. Catabolism was inhibited by chloroquine and monensin (inhibitors of lysosomal acidification) and by cytochalasin (an inhibitor of microfilament polymerization). Binding to the surface of cells was not markedly inhibited by any agent tested. The capacity of cells to bind labeled IgG1 was markedly reduced by prior incubation in the presence of unlabeled IgG1. This reduction was in part due to the steric blockade of receptors caused by the avid, but reversible, binding of IgG1. In addition, IgG1 oligomers or HP (but not IgG1 monomers or dimers) also caused an irreversible reduction in the number of Fc receptors by a process analogous to receptor down-regulation, as observed in other receptor--ligand systems.     

Tear calcium levels and contact lens wear.

Hydrophilic contact lens spoliation can be associated with the deposition of calcium salts. The relationship between the concentration of calcium in tear fluid and contact lens use has been studied in healthy volunteers and in patients using various forms of hard and soft contact lenses, tear samples being collected with Schirmer strips and with lenses in situ. Methods are suggested for correcting data on calcium per strip to concentration when this cannot be directly determined. Further data were obtained by direct extraction of freshly removed soft contact lenses. The collected results indicate some variation in tear calcium concentrations, but values are unrelated to use of contact lens and type of lens fitted, and to the rates of tear flow with our procedures for collection. Local calcium concentrations are unlikely therefore to be a significant primary factor in soft contact lens spoliation, but the enlargement of the tear pool associated with the use of a soft contact lens does greatly increase the amount of calcium present, and this may be a factor in secondary deposition.     

Possibility of selection against mtDNA mutations in tumors

Several studies of tumors have revealed substantial numbers of clonally expanded somatic mutations in mitochondrial DNA (mtDNA), not observed in adjacent intact tissues. These findings were interpreted as indicating the involvement of mtDNA mutations in tumorigenesis. Such comparisons, however, ignore an important confounding factor: the monoclonal origin of tumors as opposed to the highly polyclonal nature of normal tissues. Analysis of recently published data on the incidence of somatic mutations in nontumor monoclonal cells suggests that, contrary to the prevailing view, the process of tumorigenesis may be accompanied by active selection against detrimental mtDNA mutations.     

Effects of periadolescent versus adult cocaine exposure on cocaine conditioned place preference and motor sensitization in mice

Rationale Age of initial exposure to addictive substances is inversely proportional to risk of developing drug dependence. There is debate, however, as to whether intake at a young age causes dependency or whether young people who experiment with addictive substances are predisposed to dependency by other factors.Objectives We tested the relationship between cocaine exposure at two different ages in mice and the development of subsequent drug-seeking behavior to test for age-specific exposure effects.Methods We performed dose-response analysis of cocaine conditioned place preference (CPP) and locomotor activity in periadolescent and adult C57Bl/6J mice. In addition, we pretreated periadolescent and adult C57Bl/6J mice with cocaine or saline in the home cage or a drug-associated context, and then examined their behavior in a biased CPP procedure in adulthood.Results Dose-response relationships were similar between the two age groups. In the pretreatment experiments, we observed locomotor sensitization during the pretreatment in periadolescent but not adult mice. We also observed an enhanced aversion to the non-preferred side of the chamber in periadolescent mice compared to adult mice, which was alleviated by cocaine association with that side. Third, we observed that after further conditioning in adulthood, there were no pretreatment-specific effects.Conclusions Our results are consistent with a vulnerable brain hypothesis for responses to cocaine based on our findings that periadolescent mice exhibit greater locomotor sensitization to cocaine, and greater baseline anxiety responses that are alleviated by cocaine exposure compared to adult mice.     

Dorsal and ventral distribution of excitable and synaptic properties of neurons of the bed nucleus of the stria terminalis

The bed nucleus of the stria terminalis (BNST) is a structure uniquely positioned to integrate stress information and regulate both stress and reward systems. Consistent with this arrangement, evidence suggests that the BNST, and in particular the noradrenergic input to this structure, is a key component of affective responses to drugs of abuse. We have utilized an in vitro slice preparation from adult mice to determine synaptic and membrane properties of these cells, focusing on the dorsal and ventral subdivisions of the anterolateral BNST (dBNST and vBNST) because of the differential noradrenergic input to these two regions. We find that while resting membrane potential and input resistance are comparable between these subdivisions, excitable properties, including a low-threshold spike (LTS) likely mediated by T-type calcium channels and an Ih-dependent potential, are differentially distributed. Inhibitory and excitatory postsynaptic potentials (IPSPs and EPSPs, respectively) are readily evoked in both dBNST and vBNST. The fast IPSP is predominantly GABAA-receptor mediated and is partially blocked by the AMPA/kainate-receptor antagonist CNQX. In the presence of the GABAA-receptor antagonist picrotoxin, cells in dBNST but not vBNST are more depolarized and have a higher input resistance, suggesting tonic GABAergic inhibition of these cells. The EPSPs elicited in BNST are monosynaptic, exhibit paired pulse facilitation, and contain both an AMPA- and an N-methyl-d-aspartate (NMDA) receptor-mediated component. These data support the hypothesis that neurons of the dorsal and ventral BNST differentially integrate synaptic input, which is likely of behavioral significance. The data also suggest mechanisms by which information may flow through stress and reward circuits.     

Teaching ISC technique

Intermittent self-catheterisation (ISC) is an extremely intimate technique and those teaching it must be aware of both verbal and non-verbal behaviour by patients throughout the procedure. The instructor needs to be empathetic, supportive and skilled in the technique, as the patient faces a change in both lifestyle and body image. Psychosexual awareness means being alert to any sexual anxieties the patient may have.     

Attentional demands for static postural control after stroke

OBJECTIVE: To assess the attentional demands associated with postural control among people who have had a stroke. DESIGN: Nonrandomized matched case-control study. SETTING: University research laboratory in Canada. PARTICIPANTS: Six individuals who had suffered a left or right cerebral ischemic attack in the past year and a sample of 6 age- and gender-matched controls. Participants in the stroke group had a mean age of 64.17+/-13.14 years; control participants had a mean age of 64.00+/-13.91 years. Mean National Institute of Health Stroke Scale scores for these patients were 7.67+/-4.92 at the time of stroke and 1.66+/-1.36 at the time of testing. None of the patients were taking medications that would alter cognitive status or balance abilities. INTERVENTION: Participants performed a verbal reaction-time test while engaged in 3 postural tasks (sitting, standing, standing with feet together). MAIN OUTCOME MEASURE: Reaction time: latency between visual stimulus and verbal response. RESULTS: Reaction times in the stroke group differed significantly in all conditions from the controls (410+/-72 ms vs 320+/-54 ms, P<.01). A significant interaction was found between group and postural task (P=.05), with reaction-time scores showing a progressive increase in postural task difficulty among participants who had suffered a stroke. Post hoc comparisons revealed that sitting reaction-time scores were significantly slower than reaction-time scores for feet together standing (P=.008) among participants in the stroke group. CONCLUSION: Individuals who have suffered a stroke showed increased attentional demands for tasks of static postural control compared with healthy, age-matched participants.