Correlated fluorescence lifetime and spectral measurements in living cells

Studies of proteins' interaction in cells by FRET can take benefit from two important photo-physical properties describing fluorescent proteins: fluorescence emission spectrum and fluorescence lifetime. These properties provide specific and complementary information about the tagged proteins and their environment. However, none of them taken individually can completely quantify the involved fluorophore characteristics due to their multiparametric dependency with molecular environment, experimental conditions, and interpretation complexity. A solution to get a better understanding of the biological process implied at the cellular level is to combine the spectral and temporal fluorescence data acquired simultaneously at every cell region under investigation. We present the SLiM-SPRC160, an original temporal/spectral acquisition system for simultaneous lifetime measurements in 16 spectral channels directly attached to the descanned port of a confocal microscope with two-photon excitation. It features improved light throughput, enabling low-level excitation and minimum invasivity in living cells studies. To guarantee a fairly good level of accuracy and reproducibility in the measurements of fluorescence lifetime and spectra on living cells, we propose a rigorous protocol for running experiments with this new equipment that preserves cell viability. The usefulness of SLiM approach for the precise determination of overlapping fluorophores is illustrated with the study of known solutions of rhodamine. Then, we describe reliable FRET experiments in imaging mode realized in living cells using this protocol. We also demonstrate the benefit of localized fluorescence spectrum-lifetime acquisitions for the dynamic study of fluorescent proteins. proteins.     

An integer goal programming model for determining capital expenditures for correctional facilities

The selection of capital expenditure projects for the construction of state correctional facilities is often complicated by the existence of multiple and conflicting objectives on the part of the various interest groups involved in the decision-making process. While some groups view the limited availability of state funds to construct such facilities as the paramount consideration, others might consider having adequate capacity to house prisoners in a satisfactory manner, and the effect of prison overcrowding on prisoner sentencing as the primary factors in the decision to construct new facilities. As such. it is imperative that the limited funds available for constructing correctional facilities be allocated in the most efficient and satisfactory manner possible. In this paper, integer goal programming is demonstrated via a case example as a means for allocating funds for capital expenditures for new and renovated correctional facilities. Sensitivity analysis is performed using the model in order to demonstrate its capability for testing various planning scenarios including alternative priority structures, goal constraints and goal levels.     

Constipation Mitigation by Rhubarb Extract in Middle-Aged Adults Is Linked to Gut Microbiome Modulation: A Double-Blind Randomized Placebo-Controlled Trial

Gut microbiota alterations are intimately linked to chronic constipation upon aging. We investigated the role of targeted changes in the gut microbiota composition in the relief of constipation symptoms after rhubarb extract (RE) supplementation in middle-aged volunteers. Subjects (95% women, average 58 years old) were randomized to three groups treated with RE at two different doses determined by its content of rhein (supplementation of 12.5 mg and 25 mg per day) vs. placebo (maltodextrin) for 30 days. We demonstrated that daily oral supplementation of RE for 30 days was safe even at the higher dose. Stool frequency and consistency, and perceived change in transit problem, transit speed and difficulty in evacuating, investigated by validated questionnaires, were improved in both groups of RE-treated volunteers compared to placebo. Higher abundance of Lachnospiraceae (mainly Roseburia and Agathobacter) only occurred after RE treatment when present at low levels at baseline, whereas an opposite shift in short-chain fatty acid (SCFA) levels was observed in both RE-treated groups (increase) and placebo (decrease). Fecal Lachnospiraceae and SCFA were positively correlated with stool consistency. This study demonstrates that RE supplementation promotes butyrate-producing bacteria and SCFA, an effect that could contribute to relieving chronic constipation in middle-aged persons.     

Mucosal Adjuvants Delivered by a Mucoadhesive Patch for Sublingual Administration of Subunit Vaccines

Among mucosal administration routes for vaccines, the sublingual route has been proven capable of inducing a potent systemic and mucosal immune response. However, the absence of a simple and compliant delivery system and the lack of robust mucosal adjuvants impede the development of sublingual vaccines. Here, we describe a mucoadhesive patch made of a layer-by-layer assembly of polysaccharides, chitosan, and hyaluronic acid. The mucoadhesive patch was covered by adjuvanted nanoparticles carrying viral proteins. We showed that the nanoparticles effectively cross the outer layers of the sublingual mucosa to reach the epithelium. Furthermore, the encapsulated adjuvants, 3M-052 and mifamurtide, targeting toll-like receptor (TLR) 7/8 and nucleotide-binding oligomerization domain-2 (NOD2), respectively, remain fully active after encapsulation into nanoparticles and exhibit a cytokine/chemokine signature similar to the mucosal gold-standard adjuvant, the cholera toxin. However, the particulate adjuvants induced more moderate levels of proinflammatory interleukin (IL)-6 and keratinocyte chemoattractant (KC), suggesting a controlled activation of the innate immune response.     

The Size-Dependent Effects of Silver Nanoparticles on Germination,Early Seedling Development and Polar Metabolite Profile of Wheat (Triticum aestivum L.)

The phytotoxicity of silver nanoparticles (Ag NPs) to plant seeds germination and seedlings development depends on nanoparticles properties and concentration, as well as plant species and stress tolerance degrees. In the present study, the effect of citrate-stabilized spherical Ag NPs (20 mg/L) in sizes of 10, 20, 40, 60, and 100 nm, on wheat grain germination, early seedlings development, and polar metabolite profile in 3-day-old seedlings were analyzed. Ag NPs, regardless of their sizes, did not affect the germination of wheat grains. However, the smaller nanoparticles (10 and 20 nm in size) decreased the growth of seedling roots. Although the concentrations of total polar metabolites in roots, coleoptile, and endosperm of seedlings were not affected by Ag NPs, significant re-arrangements of carbohydrates profiles in seedlings were noted. In roots and coleoptile of 3-day-old seedlings, the concentration of sucrose increased, which was accompanied by a decrease in glucose and fructose. The concentrations of most other polar metabolites (amino acids, organic acids, and phosphate) were not affected by Ag NPs. Thus, an unknown signal is released by small-sized Ag NPs that triggers affection of sugars metabolism and/or distribution.     

Searching for multimedia: analysis of audio,video and image Web queries

The development of digital libraries has enhanced the integration of textual and multimedia information in many document collections. The World Wide Web provides the connectivity for many digital library users. Studies exploring the searching characteristics of Web users are an important and a growing area of research. Most Web user studies have focused on general Web searching, regardless of subject matter or format. Little research has examined how Web users search for multimedia information. Our study examines users' multimedia searching on a major Web search service. The data set examined consisted of 1,025,908 queries from 211,058 users of Excite ®, a major Web search service. From this data set, we identified and analyzed queries for audio, image, and video queries. Our findings were compared to results from general Web searching studies. Implications for the design of Web searching services and interfaces are discussed.     

Multi-heuristic scheduling: three approaches to tune compromises

Most of the available industrial schedulers are based on a simulation approach using dispatching rules. These rules are often dedicated to the satisfaction of a single performance criterion, and are used whatever the characteristics of the workshop or of the set of jobs. An approach which allows one to bring in compromises between rules is set out in this paper. These compromises can be parametered in accordance with the objectives of the workshop and the characteristics of the jobs in order to introduce some reactivity in the decision system. Three ways to set up the parameters are compared: experimental design, fuzzy expert system and neural network. The method allowing one to define compromises can be implemented on each scheduler that uses a simulation approach. Tests have been made with an industrial scheduler called SIPAPLUS, the results of which are developed in this paper.