Systemic Administration of Immunostimulatory DNA Sequences Mediates Reversible Inhibition of Th2 Responses in a Mouse Model of Asthma

This study investigated whether immunostimulatory DNA sequences (ISS) induce a transient or sustained inhibition of Th2 responses to inhaled antigen. We sensitized mice with subcutaneous injections to develop a Th2 response to ovalbumin (ova) and then administered a dose of ISS prior to ova inhalation challenge. Mice were then rechallenged with ova by inhalation a second time at varying time points after the first ova inhalation (1 to 8 weeks later) to determine whether the ISS dose administered prior to the first ova inhalation protected against a subsequent second ova inhalation challenge. A single dose of ISS inhibited the Th2 response to the first inhalation of ova antigen, as well as 4 weeks later to the second inhalation of ova. However, ISS did not inhibit a Th2 response to the second inhalation of ova 8 weeks later. The reversible inhibition of Th2 responses at 8 weeks suggests the need for repeated ISS administration at monthly intervals.     

The influence of age on neurotransmitter turnover in the rat's superior colliculus.

Measurements of the turnover of dopamine, noradrenaline and serotonin and their metabolites have been performed in the superior colliculus of adult and aged rats. The turnover of dopamine, noradrenaline and their metabolites after pargyline treatment was significantly lower in aged rats than in adults. On the contrary, the synthesis rate of serotonin (measured by accumulation of 5-hydroxytryptophan after decarboxylase blockade) and the turnover rate of serotonin (after pargyline treatment) did not change during aging. These findings suggest that aging has a different effect on catecholamines and serotonin turnover in the superior colliculus of the aged rats.     

Single-column thalamocortical network model exhibiting gamma oscillations, sleep spindles, and epileptogenic bursts

To better understand population phenomena in thalamocortical neuronal ensembles, we have constructed a preliminary network model with 3,560 multicompartment neurons (containing soma, branching dendrites, and a portion of axon). Types of neurons included superficial pyramids (with regular spiking [RS] and fast rhythmic bursting [FRB] firing behaviors); RS spiny stellates; fast spiking (FS) interneurons, with basket-type and axoaxonic types of connectivity, and located in superficial and deep cortical layers; low threshold spiking (LTS) interneurons, which contacted principal cell dendrites; deep pyramids, which could have RS or intrinsic bursting (IB) firing behaviors, and endowed either with nontufted apical dendrites or with long tufted apical dendrites; thalamocortical relay (TCR) cells; and nucleus reticularis (nRT) cells. To the extent possible, both electrophysiology and synaptic connectivity were based on published data, although many arbitrary choices were necessary. In addition to synaptic connectivity (by AMPA/kainate, NMDA, and GABA(A) receptors), we also included electrical coupling between dendrites of interneurons, nRT cells, and TCR cells, and--in various combinations--electrical coupling between the proximal axons of certain cortical principal neurons. Our network model replicates several observed population phenomena, including 1) persistent gamma oscillations; 2) thalamocortical sleep spindles; 3) series of synchronized population bursts, resembling electrographic seizures; 4) isolated double population bursts with superimposed very fast oscillations (>100 Hz, "VFO"); 5) spike-wave, polyspike-wave, and fast runs (about 10 Hz). We show that epileptiform bursts, including double and multiple bursts, containing VFO occur in rat auditory cortex in vitro, in the presence of kainate, when both GABA(A) and GABA(B) receptors are blocked. Electrical coupling between axons appears necessary (as reported previously) for persistent gamma and additionally plays a role in the detailed shaping of epileptogenic events. The degree of recurrent synaptic excitation between spiny stellate cells, and their tendency to fire throughout multiple bursts, also appears critical in shaping epileptogenic events.     

CXCR4 and CCR5 expression by H9 T-cells is downregulated by a peptide-nucleic acid immunomodulator

Product R (Reticulose(TM)) is a peptide-nucleic acid immunomodulator with broad-spectrum antiviral activity that was recently shown to increase expression of mRNAs encoding the proinflammatory cytokines, IFN-gamma, IL-1beta, IL-6 and TNF-alpha. Since these cytokines induce expression of the chemokines, MIP-1alpha, MIP-1beta, RANTES, and SDF-1, all of which inhibit viral infectivity, we were interested to determine if Product R also alters chemokine expression. In addition, the finding, that Product R decreases HIV-1 RNA and extracellular p24 antigen in H9 T-lymphoma cells, suggested to us that this drug may block viral infection by reducing the expression of chemokine receptors on target cells. We have therefore utilized H9 cells to test the effects of Product R on expression of mRNAs encoding the chemokine receptors, CD4, CXCR4 and CCR5, as well as their ligands, IL-16, SDF-1, MIP-1alpha, MIP-1beta, and RANTES, by RT-PCR. We also assayed the effect of Product R on surface receptor expression by flow cytometry, and on the chemotactic activity of these cells towards the CXCR4 ligand, SDF-1, and the CCR5 ligands, MIP-1alpha and RANTES. H9 cells were cultured for 3-21 days in medium containing 5% or 10% Product R, or 5% or 10% PBS. We found that, compared to control cultures, cells cultured in media containing Product R expressed lower amounts of CXCR4 and CCR5 mRNA and surface antigen at all time points. Culture for 3 days in media containing Product R also reduced the ability of cells to migrate towards 10-20 ng/ml SDF-1 and 100-250 ng/ml RANTES. In contrast, Product R had no effect on the expression of CD4 mRNA and receptor protein, or on expression of IL-16 mRNA. These findings suggest that Product R may have clinical efficacy in HIV-1-infected patients by downregulating viral coreceptors on target T-cells.     

Temporal-spatial ablation of neural crest in the mouse results in cardiovascular defects.

Neural crest cells are thought to play a critical role in human conotruncal morphogenesis and dysmorphogenesis. Much of our understanding of the contribution of neural crest to cardiovascular patterning comes from ablation and transplantation experiments in avian species. Although fate mapping experiments in mice suggests a conservation of function, the functional requirement for neural crest in cardiovascular development in mammals has not been formally tested. We used a novel two component genetic system for the temporal-spatial ablation of neural crest in the mouse. Affected embryos displayed a spectrum of cardiovascular outflow tract defects and aortic arch patterning abnormalities. We show that the severity of the cardiovascular phenotype is directly related to the level and extent of neural crest ablation. This is the first report of cardiac neural crest ablation in mammals, and it provides important insight into the role of the mammalian neural crest during cardiovascular development.     

Differential Effects of Thalidomide on Angiogenesis and Tumor Growth in Mice

Thalidomide, clinically used as an antiinflammatory and antitumoral drug, inhibited sponge-induced angiogenesis when administered systemically (100 mg/kg^–1) in mice. However, it failed to inhibit solid Ehrlich tumor in the same mouse strain. We have used functional, biochemical and histological parameters to assess neovascularization and fibrovascular tissue infiltration of the mice sponge granuloma. The neovascularization growth as detected by development of blood flow and hemoglobin content extracted from the implants showed that thalidomide inhibited fibrovascular tissue formation by 40%. The functional and biochemical parameters correlated well with the histological study. Thalidomide had no inhibitory effect in the development of Ehrlich tumor. The detection of this selective action using the same animal strain bearing two different processes, supports the hypothesis that rather than species specificity, thalidomide is tissue specific. This approach may be used to identify the specificity of other therapeutic agents against distinct angiogenesis-dependent diseases.     

Increased Bronchial Hypersensitivity after Early and Late Bronchial Reactions Provoked by Allergen Inhalation

The non-specific bronchial reactivity following bronchial allergen challenge was studied in 40 patients with allergic bronchial asthma, particularly in subjects without definite late reactions 6 h after the provocations (reduction in peak expiratory flow or forced expiratory volume in 1 s of less than 15% of the control value at this time). Among a group of 21 patients submitted to bronchial provocation tests, 13 carried out maximal exercise tests 6 and 1 week after the allergen challenge. In another group of 19 patients, the bronchial hyperreactivity to methacholine was assessed before and 6 h and 1 week after challenge. Two patients with a dual response (early & late) reacted with bronchial obstruction to the exercise. Exercise tests performed after 1 week did not provoke asthma in any patient. In the methacholine group a marked increase in responsiveness to methacholine 6 h after the provocation was observed in those patients with a dual response who were tested and in those with equivocal late reactions and even in three patients with an isolated immediate reaction. The increases responsiveness was still present in many patients 1 week after challenge. The airway caliber did not influence the degree of responsiveness to methacholine. Nor did the degree of responsiveness have any influence on the patterns of reactions observed after allergen exposure. It was concluded that in some individuals exposure to the relevant allergen predisposes them to exercise-inducible bronchial obstruction. Further, it was confirmed that non-specific bronchial reactivity can be increased not only in patients with late responses - both definite and equivocal--but also in some patients with immediate reactions alone.(ABSTRACT TRUNCATED AT 250 WORDS)     

Intracellular lectin-binding sites in symbiont-bearingCrithidia species

Crithidia oncopelti, C. deanei, andC. desouzai are flagellates of the Trypanosomatidae family that present bacterium-like endosymbionts in their cytoplasm. Direct and indirect lectin-gold labeling techniques were used at the electron microscopic level in Lowicryl K4M-embedded cells to demonstrate the presence of intracellular lectin-binding sites. We used the lectinsUlex europaeus I, Griffonia simplicifolia II, Ricinus communis I, Arachis hypogaea, G. simplicifolia I, Wistaria floribunda, Limulus polyphemus, andCanavalia ensiformis, which recognize -l-fucose, - and -N-acetylglucosamine, -galactose and -N-acetylgalactosamine, -galactose, -galactose, -N-acetylgalactosamine, sialic acid and -d-mannose, and -d-glucose residues, respectively. The nucleus was the cellular structure most frequently labeled by the lectins. The Golgi complex was seldom labeled, whereas the endoplasmic reticulum and the flagellar pocket presented a large number of binding sites. Symbionts had their two unit membranes weakly labeled by the different lectins but displayed no labeling of the space between the membranes.     

Dynamics of excitation and inhibition underlying stimulus selectivity in rat somatosensory cortex

Neurons in sensory systems respond to stimuli within their receptive fields, but the magnitude of the response depends on specific stimulus features. In the rodent whisker system, the response magnitude to the deflection of a particular whisker is, in most cells, dependent on the direction of deflection. Here we use in vivo intracellular recordings from thalamorecipient neurons in layers 3 and 4 of the rat barrel cortex to elucidate the dynamics of the synaptic inputs underlying direction selectivity. We show that cells are direction selective despite a broadly tuned excitatory and inhibitory synaptic input. Selectivity emerges from a direction-dependent temporal shift of excitation relative to inhibition. For preferred direction deflections, excitation precedes inhibition, but as the direction diverges from the preferred, this separation decreases. Our results illustrate a mechanism by which the timing of the synaptic inputs, and not their relative peak amplitudes, primarily determine feature selectivity.