Dendritic cell populations in Leishmania major-infected skin and draining lymph nodes

Using a metacyclic promastigote ear infection model of cutaneous leishmaniasis, we examined the phenotype, parasite load, and cytokine production of dendritic cells in the skin and draining lymph nodes of resistant C57BL/6J and susceptible BALB/c mice. Five dendritic cell populations were isolated from the skin and lymph nodes, and the main difference between the groups of mice was an increased number of plasmacytoid dendritic cells in the lymph nodes of the susceptible mice. Although similar cell types were present in the skin emigrants of both strains, there was a 10-fold larger number of cells in BALB/c mouse skin early in infection than in C57BL/6J mouse skin. None of the dendritic cells in the lymph nodes harbored parasites until 3 weeks after infection, with the Langerhans cells having the largest load and the plasmacytoid dendritic cells having the smallest load but the longest lasting infection. Although parasites could be detected in the lymph nodes a few hours after infection, none of the skin emigrants harbored parasites, indicating that they are not the vehicle that ferries the parasites from the skin to the lymph nodes. The presence of larger numbers of plasmacytoid cells in infected BALB/c mice, the more protracted infection of these cells, and their production of alpha interferon point to a complex and important role for the plasmacytoid cells in leishmaniasis.     

Effect of temperature on growth, hemagglutination, and protease activity of Porphyromonas gingivalis

Bacteria persisting in periodontal pockets are exposed to elevated temperatures during periods of inflammation. Temperature is an environmental factor that can modulate gene expression. Consequently, in the present study we examined the effect of temperature on the expression of virulence determinants by the periodontopathogen, Porphyromonas gingivalis. P. gingivalis W50 was grown in a complex medium under hemin excess at pH 7.0 and at a constant temperature of either 37, 39, or 41 degrees C; cultures were monitored for protease and hemagglutinin activity. P. gingivalis grew well at all three temperatures. An increase in growth temperature from 37 to 39 degrees C resulted in a 65% reduction in both total arginine- and lysine-specific activities (P < 0.01). A further rise in growth temperature to 41 degrees C led to even greater reductions in arginine-specific (82%; P < 0.001) and lysine-specific (73%; P < 0. 01) activities. These reductions were also associated with an altered distribution of individual arginine-specific enzyme isoforms. At 41 degrees C, there was a disproportionate reduction in the level of the heterodimeric RI protease, which also contains adhesin domains. The reduction also correlated with a markedly diminished hemagglutination activity of cells, especially in those grown at 41 degrees C, and a reduced immunoreactivity with a monoclonal antibody which recognizes gene products involved in hemagglutination. Thus, as the environmental temperature increased, P. gingivalis adopted a less aggressive phenotype, while retaining cell population levels. The coordinate down-regulation of virulence gene expression in response to an environmental cue linked to the intensity of the host inflammatory response is consistent with the clinically observed cyclical nature of disease progression in periodontal diseases.     

Identification of a mutation in synapsin I, a synaptic vesicle protein, in a family with epilepsy

A four generation family is described in which some men of normal intelligence have epilepsy and others have various combinations of epilepsy, learning difficulties, macrocephaly, and aggressive behaviour. As the phenotype in this family is distinct from other X linked recessive disorders linkage studies were carried out. Linkage analysis was done using X chromosome microsatellite polymorphisms to define the interval containing the causative gene. Genes from within the region were considered possible candidates and one of these, SYN1, was screened for mutations by direct DNA sequencing of amplified products. Microsatellite analysis showed that the region between MAOB (Xp11.3) and DXS1275 (Xq12) segregated with the disease. Two point linkage analysis demonstrated linkage with DXS1039, lod score 4.06 at theta = 0, and DXS991, 3.63 at theta = 0. Candidate gene analysis led to identification of a nonsense mutation in the gene encoding synapsin I that was present in all affected family members and female carriers and was not present in 287 control chromosomes. Synapsin I is a synaptic vesicle associated protein involved in the regulation of synaptogenesis and neurotransmitter release. The SYN1 nonsense mutation that was identified is the likely cause of the phenotype in this family.     

Semi-automated analysis of manually reconstructed tracks of progressively motile human spermatozoa

Several alternative algorithms for computer-assisted derivationof measurements of movement characteristics from manually reconstructedtracks of progressively motile human spermatozoa were compared.Fifty tracks were reconstructed at 30 Hz from video recordingsand analysed using traditional manual methods and by four combinationsof computer algo rithms. The best algorithm set was identified(‘Video-mot.mdpt’) and the values for the curvilinear,average path and linear velocities (V_CVL, V_AVE and V_LIN respectively),the amplitude of lateral displacement of the sperm head aboutthe axis of progression (A_H) and the number of times the spermhead crossed the average path (the ‘beat/cross frequency’,BXF) obtained using it were compared to those obtained by manualanalysis. There was a considerable time saving when the computer-assistedmethod was used and the values it gave for the various movementcharacteristics showed good correspondence with those obtainedmanually. In addition, repeated data entry and analysis wasfound to be highly reproducible. When the tracks were re-plottedat 6 Hz (as used by the multiple-exposure photomicrography methodfor sperm motility analysis) insufficient information remainedin the tracks for reliable determination of anything other thanV_LIN We conclude that the Videomot.mdpt program provides reliablevalues for the movement characteristics of progressively motilehuman spermatozoa, and believe it will be of great value inthe validation of commercial systems providing automated spermmovement analysis and in laboratories which do not have accessto such costly equipment.     

Dual effects of hemicholinium-3 at central cholinergic synapses.

1. Hemicholinium-3 (HC-3) has at least two presynaptic effects at cholinergic synapses between motor acon collaterals and spinal Renshaw cells, neither of which influenced non-cholinergic excitatory synapses on dorsal horn interneurones. 2. One, which occurred independently of repetitive activity of the synapses and which resulted in increased latency of synaptic excitation, presumably involved changes in the properties of presynaptic fibres and terminals which influenced transmitter release. 3. The other, which depended on continuous repetitive synaptic activity, was consistent with depletion by HC-3 of presynaptic stores of acetylcholine (ACh).     

A contribution to sonograph image quality estimation using point spread function

In medical sonography, sonograph image quality is an essential aspect for the safety of both patient and doctor. Its evaluation therefore requires an accurate and objective method for measurement. In this regard, a number of methods are in current use. Most of these are based on tissue mimicking phantom imaging. In contrast, we have used another principle based on Point Spread Function (PSF) analysis which is a product of the measuring system we have developed. In this case, the measured sonograph scans a small metallic ball target that moves in a water bath on a specified trajectory. The Region Of Interest (ROI) of the sonogram containing the ball target picture is digitised and the amplitude of the pixels analysed. The result is the PSF from which we calculate the lateral resolution (LR). For this purpose, we use our own original software. Using this method, we have to date been able to plot LR characteristics over the scanning plane. The method allows us to differentiate separate scanning lines and even multiple focal areas for dynamic focussing systems. It can detect malfunctions in dynamic focussing, size of aperture, time gain compensation function and/or transducer element failure. The procedure itself is not as easy or as fast to use as tissue mimicking phantoms or 3D signal to noise ratio evaluation, but it provides accurate and objective numeric parameters corresponding to the quality of image at any specified point over the whole scanning area. It is also a very powerful tool when used in combination with the other methods mentioned above.     

A pharmacological study of group I muscle afferent terminals and synaptic excitation in the intermediate nucleus and Clarke's column of the cat spinal cord

Summary When administered microelectrophoretically GABA and piperidine-4-sulphonic acid depolarized the central terminations of muscle group Ia and Ib afferent fibres in the lumbar intermediate nucleus and Clarke's column of cats anaesthetised with pentobarbitone sodium. Both this depolarization, and primary afferent depolarization, generated by impulses in other primary afferent fibres which produce prolonged bicuculline-sensitive inhibition of the firing of group I afferent fibre-excited interneurones in the intermediate nucleus and cells in Clarke's column, are reduced by microelectrophoretic bicuculline methochloride. Systemically administered (±)-baclofen hydrochloride (maximum dose 8 mg kg^–1) depressed the monosynaptic excitation of Clarke's column neurones by impulses in muscle and cutaneous afferent fibres. Microelectrophoretically administered (–)-baclofen reduced the bicuculline-sensitive primary afferent depolarization of group I terminations without, however, reducing the depolarizing action of GABA or piperidine-4-sulphonic acid. The depression by (–)-baclofen of the group I monosynaptic excitation of intermediate nucleus neurones is not reduced by concentrations of bicuculline methochloride adequate to suppress prolonged inhibition of these neurones     

A critical evaluation of current views regarding eye movement desensitization and reprocessing (EMDR): clarifying points of confusion.

EMDR is an active psychological treatment for PTSD that has received widely divergent reactions from the scientific and professional community. This article examines points of confusion in the published literature on EMDR, including the theoretical, empirical, and historical issues around EMDR and placebo effects, exposure procedures, the eye movement component, treatment fidelity issues, and outcome studies. It also examines historical information relevant to the scientific process and charges of "pseudoscience" regarding EMDR. We conclude that the confusion in the literature is due to (a) the lack of an empirically validated model capable of convincingly explaining the effects of the EMDR method, (b) inaccurate and selective reporting of research, (c) some poorly designed empirical studies, (d) inadequate treatment fidelity in some outcome research, and (e) multiple biased or inaccurate reviews by a relatively small group of authors. Reading the original research articles frequently helps to reduce the confusion arising from the research review literature.     

Targeted disruption of fibronectin-integrin interactions in human gingival fibroblasts by the RI protease of Porphyromonas gingivalis W50

Cell surface integrins mediate interactions between cells and their extracellular matrix and are frequently exploited by a range of bacterial pathogens to facilitate adherence and/or invasion. In this study we examined the effects of Porphyromonas gingivalis proteases on human gingival fibroblast (HGF) integrins and their fibronectin matrix. Culture supernatant from the virulent strain W50 caused considerably greater loss of the beta1 integrin subunit from HGF in vitro than did that of the beige-pigmented strain W50/BE1. Prior treatment of the W50 culture supernatant with the protease inhibitor Nalpha-p-tosyl-L-lysine chloromethyl ketone (TLCK) blocked its effects on cultured cells, indicating that this process is proteolytically mediated. Purified arginine-specific proteases from P. gingivalis W50 were able to mimic the effects of the whole-culture supernatant on loss of beta1 integrin expression. However purified RI, an alpha/beta heterodimer in which the catalytic chain is associated with an adhesin chain, was 12 times more active than RIA, the catalytic monomer, in causing loss of the alpha5beta1 integrin (fibronectin receptor) from HGF. No effect was observed on the alphaVbeta3 integrin (vitronectin receptor). The sites of action of RI and RIA were investigated in cells exposed to proteases pretreated with TLCK to inactivate the catalytic component. Use of both monoclonal antibody 1A1, which recognizes only the adhesin chain of RI, and a rabbit antibody against P. gingivalis whole cells indicated localization of RI on the fibroblasts in a clear, linear pattern typical of that seen with fibronectin and alpha5beta1 integrin. Exact colocalization of RI with fibronectin and its alpha5beta1 receptor was confirmed by double labeling and multiple-exposure photomicroscopy. In contrast, RIA bound to fibroblasts in a weak, patchy manner, showing only fine linear or granular staining. It is concluded that the adhesin component of RI targets the P. gingivalis arginine-protease to sites of fibronectin deposition on HGF, contributing to the rapid loss of both fibronectin and its main alpha5beta1 integrin receptor. Given the importance of integrin-ligand interactions in fibroblast function, their targeted disruption by RI may represent a novel mechanism of damage in periodontal disease.