Cyanide produced by human isolates of Pseudomonas aeruginosa contributes to lethality in Drosophila melanogaster

Some Pseudomonas aeruginosa strains are cyanogenic, and cyanide may contribute to the bacterium's virulence. Using human isolates of P. aeruginosa, we have shown that Drosophila melanogaster suspended above cyanogenic strains become motionless and develop bradycardia and that flies injected with cyanogenic bacterial strains die more rapidly than those injected with noncyanogenic strains. Flies exposed to cyanogenic strains had high cyanide and low adenosine triphosphate (ATP) concentrations in body extracts, and treatment with a cyanide antidote equalized survival of flies injected with cyanogenic and noncyanogenic strains. P. aeruginosa PAO1 strain with a mutation in the hydrogen cyanide synthase gene cluster was much less toxic to flies than the parental cyanogenic strain or 2 knock-in strains. Transgenic flies overexpressing rhodanese, which detoxifies cyanide by converting it to thiocyanate, were resistant to cyanide and the increased virulence of cyanogenic strains. We conclude that D. melanogaster is a good model for studying cyanide produced by P. aeruginosa.     

Impaired local microvascular vasodilatory effects of insulin and reduced skin microvascular vasomotion in obese women

Our study aim is to investigate whether obesity is characterized by an impairment of insulin-mediated vasodilatory effects and by a modification of basal vasomotion in the skin microvasculature. Forty healthy obese and forty healthy lean women were included. Microvascular effects of insulin as compared to a control substance were measured by cathodal iontophoresis combined with laser Doppler flowmetry. Vasomotion was examined by Fourier transform analyses of skin laser Doppler flow at rest. Locally administered insulin, as compared to the control substance, induced a microvascular vasodilatory response in lean (median (interquartile range): 31.6 (17.1-43.9) vs. 22.9 (16.4-36.7) perfusion units, P=0.04), but not in obese women (28.1 (14.4-47.1) vs. 27.5 (17.5-48.2) perfusion units, P=0.7). The relative insulin-induced increase in blood flow corrected for the control substance was higher in lean than obese women (ANOVA for repeated measures F=3.93, P=0.05). The contribution of the total frequency spectrum 0.01-1.6 Hz and of the frequency intervals 0.01-0.02 Hz and 0.02-0.06 Hz (representative of endothelial and neurogenic activity, respectively) to basal microvascular vasomotion was lower in obese than in lean women (P<0.05 for all). These findings show that obesity is characterized by an impaired direct microvascular vasodilatory effect of insulin and by decreased skin microvascular vasomotion in a way that is suggestive for alterations of endothelial and neurogenic activity.     

Antinociceptive effects of tumor necrosis factor alpha neutralization in a rat model of antigen-induced arthritis: evidence of a neuronal target

OBJECTIVE: The reduction of pain in the course of antiinflammatory therapy can result from an attenuation of the inflammatory process and/or from the neutralization of endogenous mediators of inflammation that act directly on nociceptive neurons. The purpose of this study was to investigate whether analgesic effects of the neutralization of tumor necrosis factor alpha (TNFalpha) are due to an attenuation of inflammation or whether direct neuronal effects significantly contribute to pain relief in the course of therapy. METHODS: Locomotor and pain-related behavior and histology were assessed in rats with chronic antigen-induced arthritis (AIA) in the knee joint, and the rats were treated with systemic saline, etanercept, or infliximab. The expression of TNF receptors (TNFRs) in dorsal root ganglia was measured using immunohistochemical analysis and polymerase chain reaction. Action potentials were recorded from afferent Adelta fibers and C fibers of the medial knee joint nerve, and etanercept and infliximab were injected intraarticularly into normal or inflamed knee joints (AIA or kaolin/carrageenan-induced inflammation). RESULTS: In rats with AIA, both etanercept and infliximab significantly decreased inflammation-induced locomotor and pain-related behavior, while joint swelling was only weakly attenuated and histomorphology still revealed pronounced inflammation. A large proportion of dorsal root ganglion neurons showed TNFRI- and TNFRII-like immunoreactivity. Intraarticular injection of etanercept reduced the responses of joint afferents to mechanical stimulation of the inflamed joint starting 30 minutes after injection, but had no effect on responses to mechanical stimulation of the uninflamed joint. CONCLUSION: Overall, these data show the pronounced antinociceptive effects of TNFalpha neutralization, thus suggesting that reduction of the effects of TNFalpha on pain fibers themselves significantly contributes to pain relief.     

Assessment of alphavbeta3 integrin expression after myocardial infarction by positron emission tomography

AIMS: The purpose of this study was to determine the feasibility of a new positron emission tomography (PET) imaging approach using an (18)F-labelled alpha(v)beta(3) integrin antagonist ((18)F-Galacto-RGD) to monitor the integrin expression after myocardial infarction. METHODS AND RESULTS: Male Wister rats were subjected to 20 min transient left coronary artery occlusion followed by reperfusion. Autoradiographic analysis and in vivo PET imaging were used to determine myocardial (18)F-Galacto-RGD uptake at different time points following reperfusion. RESULTS: PET imaging and autoradiography demonstrated no significant focal myocardial (18)F-Galacto-RGD uptake in non-operated control rats and at day 1 after reperfusion. However, focal accumulation in the infarct area started at day 3 (uptake ratio = 1.91 +/- 0.22 vs. remote myocardium), peaked between 1 (3.43 +/- 0.57) and 3 weeks (3.43 +/- 0.95), and decreased to 1.96 +/- 0.40 at 6 months after reperfusion. Pretreatment with alpha(v)beta(3) integrin antagonist c(-RGDfV-) significantly decreased tracer uptake, indicating the specificity of tracer uptake. The time course of focal tracer uptake paralleled vascular density as measured by CD31 immunohistochemical analysis. CONCLUSION: Regional (18)F-Galacto-RGD accumulation suggests up-regulation of alpha(v)beta(3) integrin expression after myocardial infarction, which peaks between 1 and 3 weeks and remains detectable until 6 months after reperfusion. This new PET tracer is promising for the monitoring of myocardial repair processes.     

Direct assessment of thymic reactivation after autologous stem cell transplantation

Methods to quantify Th cell reconstitution after immunosuppressive therapies such as hematopoietic stem cell transplantation are becoming a key issue since persistent Th cell deficiencies may result in severe complications and adverse events. We employed here cytometric monitoring of CD31+ thymus-naive Th cells for the direct assessment of human thymic function in 10 patients undergoing autologous stem cell transplantation for severe autoimmune diseases. High frequencies of posttransplant recurring naive Th cells coexpressed CD31 and stable long-term reconstitution with elevated absolute counts of CD31+ thymus-naive Th cells that were enriched with T cell receptor excision circles was demonstrated. Cytometric monitoring of CD31+ thymus-naive Th cells enables to directly evaluate human thymic function ex vivo.