A comparative study of ex vivo skin optical clearing using two‐photon microscopy

Multiphoton tomography (MPT) is a prospective tool for imaging the skin structure. Aiming to increase the probing depth, a comparative ex vivo study of optical clearing of porcine ear skin was performed by using two optical clearing agents (OCAs), i.e., glycerol and iohexol (Omnipaque^TM) at different concentrations, which exhibit different osmotic properties. The results show that a topical application of glycerol or Omnipaque^TM solutions onto the skin for 60 min significantly improved the depth and contrast of the MPT signals. By utilizing 40%, 60% and 100% glycerol, and 60% and 100% Omnipaque^TM it was demonstrated that both agents improve autofluorescence and SHG (second harmonic generation) signals from the skin. At the applied concentrations and agent time exposure, glycerol is more effective than Omnipaque^TM. However, tissue shrinkage and cell morphology changes were found for highly concentrated glycerol solutions. Omnipaque^TM, on the contrary, increases the safety and has no or minimal tissue shrinkage during the optical clearing process. Moreover Omnipaque^TM allows for robust multimodal optical/X‐ray imaging with automatically matched optically cleared and X‐ray contrasted tissue volumes. These findings make Omnipaque^TM more prospective than glycerol for some particular application.

     

Characterization of cell lines stably expressing human normal or mutated EGFP-tagged MC4R

The melanocortin receptor type 4 (MC4-R) is involved in food intake and represents a potential target for the treatment of some forms of obesity. The fluorescent protein EGFP was fused to the wild-type or mutated coding sequence of the human MC4-R. After transfection in HEK 293, clones stably expressing hMC4-R-EGFP were selected. Wild-type chimeric hMC4-R was well addressed to the cell membrane as demonstrated using confocal microscopy and displayed the same pharmacological characteristics as native hMC4R. NDP-alpha MSH induced a time-dependent internalization of MC4-R that was partially prevented by AgRP. The two mutated chimeric receptors studied here (CTCT-deleted and C271A) showed a high alteration of their response to ligand and were retained inside the cells. In conclusion, we have developed a model of clones stably expressing EGFP-tagged-hMC4-R. This is the only such model available to date and it provides a useful tool to follow the trafficking of MC4-R inside living cells.     

Bacillus Probiotic Supplementations Improve Laying Performance,Egg Quality,Hatching of Laying Hens,and Sperm Quality of Roosters

Probiotics and Antimicrobial Proteins - The study aims at elucidating the effect of bacilli probiotic preparations on the physiology of laying hens and roosters. Probiotic formulations were...     

Syntheses and preliminary pharmacological evaluation of the two epimers of the 5-F2t-isoprostane.

The total synthesis of the 5-F2t-isoprostane 1 and its 5-epimer 2 from diacetone-D-glucose is described. We report preliminary data on the vascular properties of these compounds.     

Identifying and Tracking Simulated Synaptic Inputs from Neuronal Firing: Insights from In Vitro Experiments

Accurately describing synaptic interactions between neurons and how interactions change over time are key challenges for systems neuroscience. Although intracellular electrophysiology is a powerful tool for studying synaptic integration and plasticity, it is limited by the small number of neurons that can be recorded simultaneously in vitro and by the technical difficulty of intracellular recording in vivo. One way around these difficulties may be to use large-scale extracellular recording of spike trains and apply statistical methods to model and infer functional connections between neurons. These techniques have the potential to reveal large-scale connectivity structure based on the spike timing alone. However, the interpretation of functional connectivity is often approximate, since only a small fraction of presynaptic inputs are typically observed. Here we use in vitro current injection in layer 2/3 pyramidal neurons to validate methods for inferring functional connectivity in a setting where input to the neuron is controlled. In experiments with partially-defined input, we inject a single simulated input with known amplitude on a background of fluctuating noise. In a fully-defined input paradigm, we then control the synaptic weights and timing of many simulated presynaptic neurons. By analyzing the firing of neurons in response to these artificial inputs, we ask 1) How does functional connectivity inferred from spikes relate to simulated synaptic input? and 2) What are the limitations of connectivity inference? We find that individual current-based synaptic inputs are detectable over a broad range of amplitudes and conditions. Detectability depends on input amplitude and output firing rate, and excitatory inputs are detected more readily than inhibitory. Moreover, as we model increasing numbers of presynaptic inputs, we are able to estimate connection strengths more accurately and detect the presence of connections more quickly. These results illustrate the possibilities and outline the limits of inferring synaptic input from spikes.     

Bluetongue virus serotype 8: abortion and transplacental transmission in cattle in the Burgundy region, France, 2008-2009

During the incursion of bluetongue virus (BTV) serotype 8 in France in 2007, an increase in the number of abortions in cattle was observed, but the cause was not clearly established. A survey of all the reported cases of abortion in cattle from November 2008 to April 2009 was conducted in the Nièvre district (Burgundy region) to determine the percentage of abortions as a result of BTV-8 and to study factors that could have played a role in BTV-8 transplacental transmission. BTV-8 was present in 16% of the fetuses or newborn calves that died within 48 h, from 780 dams. Dams inseminated before the BTV epizootic peak recorded from July to September 2008 were more likely to have BTV-positive abortions (OR=5.7, P<0.001) and those vaccinated in May or June 2008 were less likely to have BTV-positive abortions (OR=0.3, P=0.01 and OR=0.4, P=0.001, respectively). The gestational month was not a predictor of BTV abortion. In blood or spleen, fetuses/calves from RT-PCR-positive dams had significantly higher RNA concentrations than fetuses/calves from RT-PCR-negative dams. Of the 128 dams that had BTV-positive fetuses or calves, 60% were RT-PCR-negative. BTV-8-positive fetuses/calves were significantly more frequent (n=42 vs n=21, P=0.082) amongst those showing clinical signs or lesions suggestive of cerebral damage.     

Age-related impairment of HDL-mediated cholesterol efflux

Our aim in this study was to investigate the effect of aging on the capacity of HDLs to promote reverse cholesterol transport. HDLs were isolated from plasma of young (Y-HDL) and elderly (E-HDL) subjects. HDL-mediated cholesterol efflux was studied using THP-1 and J774 macrophages. Our results show that E-HDLs present a lower capacity to promote cholesterol efflux than Y-HDLs (41.7 +/- 1.4% vs. 49.0 +/- 2.2%, respectively; P = 0.013). Reduction in the HDL-mediated cholesterol efflux capacity with aging was more significant with HDL(3) than HDL(2) (Y-HDL(3), 57.3 +/- 1% vs. E-HDL(3), 50.9 +/- 2%; P = 0.012). Moreover, our results show that ABCA1-mediated cholesterol efflux is the more affected pathway in terms of cholesterol-removing capacity. Interestingly, the composition and structure of HDL revealed a reduction in the phosphatidylcholine-sphingomyelin ratio (E-HDL, 32.7 +/- 2.7 vs. Y-HDL, 40.0 +/- 1.9; P = 0.029) and in the phospholipidic layer membrane fluidity in E-HDL compared with Y-HDL as well as an alteration in the apolipoprotein A-I structure and charge. In conclusion, our results shown that E-HDLs present a reduced capacity to promote cholesterol efflux, principally through the ABCA1 pathway, and this may explain the increase of the incidence of cardiovascular diseases observed during aging.