Trophic effects of insulin-like growth factor-I on fetal rat hypothalamic cells in culture

The hypothesis that insulin-like growth factor-I is a trophic factor for primary fetal rat hypothalamic cells was tested, since we previously reported a potent mitogenic effect of this peptide on virally-transformed hypothalamic cells. It was found that insulin-like growth factor-I produced significant and dose-dependent increases in the survival of fetal hypothalamic neurons in primary mixed glial/neuronal cultures. By 48 h in vitro, cultures treated with insulin-like growth factor-I (6 nM) had twice as many neurite-bearing cells as controls, while by day 15 a five-fold difference was present. The peptide was similarly active in promoting neuronal survival in neuron-enriched (98% neurons) hypothalamic cultures. Mixed hypothalamic cultures had specific binding sites for insulin-like growth factor-I. In addition, the neurons grown in the presence of insulin-like growth factor-I had a more differentiated morphology and had significantly higher levels of protein kinase C, an enzyme that increases during neurite formation and synaptogenesis. Finally, glial-enriched cultures (greater than 99% glial cells) obtained from the fetal hypothalamus showed increased [3H]thymidine incorporation in response to insulin-like growth factor-I. These results further support the contention that insulin-like growth factor-I is a neurotrophic factor and suggest that it may participate in the normal development of the hypothalamus by increasing neuronal survival/differentiation and stimulating glial growth.     

Identification ofCorynebacterium jeikeium andCorynebacterium CDC group D 2 with the API 20 strep system

A total of 170 strains ofCorynebacterium jeikeium and 23 strains ofCorynebacterium group D2 were examined in three British laboratories using the API 20 Strep identification system and three supplementary tests (catalase production, urease production and nitrate reduction). The isolates were collected from clinical specimens in various laboratories over a three-year period. The two species produced consistent reactions in these tests after 24 h. Two tests were highly discriminatory, with positive reactions for ribose fermentation seen forCorynebacterium jeikeium while urease production was observed withCorynebacterium group D2. This method allows routine clinical laboratories to rapidly identify these emerging pathogens.     

Direct sequencing of SSP-PCR-amplified cDNA to identify new alleles in the DR52-associated DRB1 group: identification of DRB1*1115, DRB1*1117 and DRB1*1319

Abstract: Low and high resolution sequence specific oligonucleotide probe hybridization patterns were used to design an approach to direct sequencing of allele specific amplified cDNA. Several PCR amplifications were used to derive overlapping sequence fragments to define complete first domain sequences for a single allele. This method has been used to characterize three new DRB1 alleles in the DR52 family, DRB1*1115, DRB1* 1117, and DRB1*1319. All three alleles carry polymorphisms previously observed in other DRB alleles and underscore the importance of utilizing a directed sequencing approach for obtaining unambiguous typing results in matching for bone marrow transplantation between unrelated donor and recipient.     

Dissociable effects of lesions to the dorsal or ventral noradrenergic bundle on the acquisition, performance, and extinction of aversive conditioning

In six experiments we studied the effects of lesions to either the dorsal or ventral noradrenergic bundle on the acquisition and extinction of the conditioned emotional response (CER) as measured in a conditioned suppression paradigm. Infusions of the neurotoxin 6-hydroxydopamine (6-OHDA) into the trajectory of the dorsal noradrenergic ascending bundle (DNAB) impaired the acquisition of on-the-baseline and off-the-baseline conditioned suppression. The acquisition impairment for on-the-baseline conditioning was also shown to still be present when training did not commence until 8 weeks following central noradrenergic depletion. However, in rats previously trained on the CER, DNAB lesions did not affect performance. There was also a small resistance to extinction following on-, but not off-the-baseline conditioning. The acquisition impairment was shown not to be because of an altered sensitivity to the footshock. In contrast, infusions of 6-OHDA into the ventral noradrenergic ascending bundle (VNAB) had no effect upon the acquisition of the CER in an on-the-baseline procedure, but retarded its extinction to a much greater extent. The results here are discussed in terms of other acquisition deficits shown by rats with DNAB lesions, and with reference to Gray's "anxiety" and Mason's "selective attention" theories of locus coeruleus function.     

Concomitant airway expression of granulocyte-macrophage colony-stimulating factor and decorin, a natural inhibitor of transforming growth factor-beta, breaks established inhalation tolerance

We previously showed that granulocyte-macrophage colony-stimulating factor (GM-CSF) breaks tolerance induction. The objective of this study was to determine whether GM-CSF breaks established inhalation tolerance. To induce tolerance, BALB/c mice were exposed to aerosolized ovalbumin (OVA) for 10 consecutive days. A control group was exposed to saline. Subsequently, tolerant and control animals were exposed to OVA in a GM-CSF-enriched airway microenvironment. Tolerant animals, unlike control animals, did not develop airway and peripheral blood eosinophilia, had diminished levels of OVA-specific IgE, and reduced airway hyper-responsiveness. While tolerant animals did not express IL-4, IL-5 and IL-13, levels of the regulatory cytokines IL-10, IFN-gamma and transfoming growth factor (TGF)-beta were similar between tolerant and non-tolerant animals. Lung CD4+ T cells were activated according to CD69, CD25 and T1/ST2 expression, but systemic responses characterized by splenocyte proliferation and Th2 effector function were dramatically reduced. Concurrent expression of GM-CSF and decorin, a natural inhibitor of TGF-beta, reversed eosinophilic unresponsiveness. Our study suggests that the breakdown of tolerance and, by extension, the emergence of eosinophilic inflammation, requires two signals: one that triggers sensitization and one that interferes with negative regulation. Moreover, our study shows that dysregulated expression of an extracellular matrix protein may break established tolerance and lead to eosinophilic airway inflammation.     

Comparative effects of quisqualic and ibotenic acid-induced lesions of the substantia innominata and globus pallidus on the acquisition of a conditional visual discrimination: differential effects on cholinergic mechanisms

Two experiments tested the hypothesis that the deficits in conditional discrimination learning produced by ibotenic acid-induced lesions of the ventral pallidum and substantia innominata are produced by loss of the magnocellular cholinergic cells in the nucleus basalis and adjacent regions. Experiment 1 replicated the previously reported deficit in conditional learning produced by ibotenate-induced lesions of the ventral pallidum/substantia innominata, but failed to demonstrate any restoration of learning by a subchronic regimen of the acetylcholinesterase inhibitor physostigmine sufficient to produce significant (30%), but equivalent, degrees of inhibition in the frontal cortex of ventral pallidum/substantia innominata-lesioned or sham-operated rats. Experiment 2 examined the effects of quisqualic acid-induced lesions of the ventral pallidum/substantia innominata. According to most of the measures of learning employed, the quisqualic acid-induced lesion of the ventral pallidum/substantia innominata failed to impair conditional learning, even though the quisqualate-induced lesion produced greater degrees of cholinergic neuron destruction than the ibotenate-induced lesion, as measured in terms of reductions in cortical choline acetyltransferase activity (44% vs 27%). Although consideration of individual data suggested that very high (60%) levels of choline acetyltransferase reduction in Experiment 2 might have detrimental effects of conditional learning, the overall failure of the quisqualate-induced lesions of the ventral pallidum/substantia innominata to impair learning is to be contrasted with the significant behavioural effects of ibotenate-induced lesions. Histological and immunocytochemical analysis showed that the quisqualate-induced lesion, unlike that produced by ibotenate, tended to produce less damage to the overlying dorsal globus pallidus and to parvocellular neurons of the ventral pallidum/substantia innominata, thus implicating these nonspecific effects of ibotenate-induced lesions in their behavioural effects. The present results question previous interpretations of the behavioural effects of ibotenate-induced lesions of the ventral pallidum/substantia innominata in terms of damage inflicted on the cortically-projecting cholinergic cells of the nucleus basalis, and suggest that quisqualic acid, although also nonspecific in its excitotoxic effects, is nevertheless more selective for producing damage to cholinergic neurons in the ventral pallidum/substantia innominata than ibotenic acid.     

Comparison between radioimmunoassay and direct and indirect enzyme-linked immunosorbent assays for determination of antibodies against Haemophilus influenzae type b capsular polysaccharide.

Levels of antibodies against Haemophilus influenzae type b capsular polysaccharide were determined in acute-phase and convalescent-phase serum samples obtained from 21 children with invasive H. influenzae type b infections and from 44 children vaccinated with two H. influenzae type b vaccines. Amounts of immunoglobulin G (IgG), IgM, and IgA antibodies were measured by direct and indirect enzyme-linked immunosorbent assay (ELISA), and the total amount of antibodies was measured by radioimmunoassay (RIA). Results obtained by ELISA were calculated by multiple-point parallel-line comparison and by endpoint analysis. A very good correlation was obtained between direct and indirect ELISA values. In the lower range of antibody concentrations, the correlation between ELISA values obtained by endpoint analysis and those obtained by multiple-point parallel-line comparison was poor, since the latter method of calculation yielded values of up to 1 microgram/ml in sera that were negative according to endpoint analysis. These sera with negative endpoint titers also had undetectable or very low antibody concentrations as measured by RIA. Consistent with this finding, in acute-phase and prevaccination sera with undetectable or low antibody concentrations as measured by RIA, ELISA values calculated by multiple-point parallel-line comparison were much higher. In sera with higher antibody concentrations, however, parallel-line comparisons showed good correlation between RIA and ELISA values. Although no reference method for measuring true antibody concentrations is available, ELISA values as calculated by multiple-point parallel-line comparison appear to overestimate antibody concentrations in sera containing low antibody concentrations, whereas ELISA values obtained by endpoint analysis are less well correlated with RIA values at higher concentrations.