Combined FCS and PCH analysis to quantify protein dimerization in living cells

Protein dimerization plays a crucial role in the regulation of numerous biological processes. However, detecting protein dimers in a cellular environment is still a challenge. Here we present a methodology to measure the extent of dimerization of GFP-tagged proteins in living cells, using a combination of fluorescence correlation spectroscopy (FCS) and photon counting histogram (PCH) analysis of single-color fluorescence fluctuation data. We named this analysis method brightness and diffusion global analysis (BDGA) and adapted it for biological purposes. Using cell lysates containing different ratios of GFP and tandem-dimer GFP (diGFP), we show that the average brightness per particle is proportional to the fraction of dimer present. We further adapted this methodology for its application in living cells, and we were able to distinguish GFP, diGFP, as well as ligand-induced dimerization of FKBP12 (FK506 binding protein 12)-GFP. While other analysis methods have only sporadically been used to study dimerization in living cells and may be prone to errors, this paper provides a robust approach for the investigation of any cytosolic protein using single-color fluorescence fluctuation spectroscopy

Major Differences in Diet across Three Linguistic Regions of Switzerland: Results from the First National Nutrition Survey menuCH.

Switzerland is a multilingual country located between Germany, France and Italy, which differ by dietary habits and related outcomes. We explored differences in food consumption as well as compliance to the Swiss food-based dietary guidelines (FBDG) across the German-, French-, and Italian-speaking regions. The 2014-2015 nationwide cross-sectional survey was conducted among a stratified random sample of 2057 adults aged 18 to 75 years. Trained dietitians assessed food consumption via two non-consecutive 24-h dietary recalls using the international validated software GloboDiet <sup>®</sup> . Recorded foods and beverages were classified into six groups and 31 subgroups relevant for assessing compliance to the FBDG (Swiss Food Pyramid). Usual daily intake distributions were modelled and weighted for sampling design, non-response, weekdays and season. Participation rate was 38%. Significant differences across regions were observed in 18 of 31 food subgroups ( <i>p</i> ≤ 0.01). Weighted mean daily intakes in the German-, French- and Italian-speaking regions were, respectively, 245 g, 155 g, 140 g for soft drinks, 273 g, 214 g, 135 g for coffee, 127 g, 72 g, 109 g for milk, 32 g, 45 g, 43 g for red meat, 18 g, 29 g, 34 g for fish/seafood, 8.1 g, 6.4 g, 3.7 g for butter, and 206 g, 214 g, 168 g for vegetables. The seven FBDGs were followed by <1% of the population. Four in 10 participants met ≥3 FBDG. Eighteen percent of participants ate ≥5 portions of fruit and vegetables a day, without regional differences. Food consumption substantially differed across the three linguistic regions of Switzerland. Adherence to FBDG was uniformly low. This highlights the potential influence of culture on diet. Nutritional education along with public health interventions are needed and may be most efficient if regionally targeted

An embedded cohesive crack model for finite element analysis of quasi-brittle materials

This paper presents a numerical implementation of the cohesive crack model for the anal-ysis of quasibrittle materials based on the strong discontinuity approach in the framework of the finite element method. A simple central force model is used for the stress versus crack opening curve. The additional degrees of freedom defining the crack opening are determined at the crack level, thus avoiding the need for performing a static condensation at the element level. The need for a tracking algorithm is avoided by using a consistent pro-cedure for the selection of the separated nodes. Such a model is then implemented into a commercial program by means of a user subroutine, consequently being contrasted with the experimental results. The model takes into account the anisotropy of the material. Numerical simulations of well-known experiments are presented to show the ability of the proposed model to simulate the fracture of quasibrittle materials such as mortar, concrete and masonry

Mast cell distribution in human carotid atherosclerotic plaque differs significantly by histological segment

Mast cells (MCs) are important contributors to atherosclerotic plaque progression. For prospective studies on mast cell contributions to plaque instability, the distribution of intraplaque MCs needs to be elucidated. Plaque stability is generally histologically assessed by dividing the plaque specimen into segments to be scored on an ordinal scale. However, owing to competitive use, studies may have to deviate to adjacent segments, yet intersegment differences of plaque characteristics, especially MCs, are largely unknown. Therefore, the hypothesis that there is no segment to segment difference in MC distribution between atherosclerotic plaque segments was tested, and intersegment associations between MCs and other plaque characteristics was investigated.\nTwenty-six carotid atherosclerotic plaques from patients undergoing carotid endarterectomy included in the Athero-Express Biobank were analysed. The plaque was divided in 5 mm segments, differentiating between the culprit lesion (segment 0), adjacent segments (-1/+1) and more distant segments (-2/+2) for the presence of MCs. The associations between the intersegment distribution of MCs and smooth muscle cells, macrophage content, and microvessel density in the culprit lesion were studied.\nA statistically significant difference in MCs/mm2 between the different plaque segments (p 2 between the culprit and adjacent segment (p = .037) and between the culprit lesion and the more distant segment (p 2 in multiple different segments were positively correlated with microvessel density and macrophage content in the culprit lesion.\nMC numbers reveal significant intersegment differences in human carotid plaques. Future histological studies on MCs should use a standardised segment for plaque characterisation as plaque segments cannot be used interchangeably for histological MC analyses.Biopharmaceutic

Effects of Potassium or Sodium Supplementation on Mineral Homeostasis: A Controlled Dietary Intervention Study

CONTEXT: Although dietary potassium and sodium intake may influence calcium-phosphate metabolism and bone health, the effects on bone mineral parameters, including fibroblast growth factor 23 (FGF23), are unclear. OBJECTIVE: Here, we investigated the effects of potassium or sodium supplementation on bone mineral parameters. DESIGN, SETTING, PARTICIPANTS: We performed a post hoc analysis of a dietary controlled randomized, blinded, placebo-controlled crossover trial. Prehypertensive individuals not using antihypertensive medication (n = 36) received capsules containing potassium chloride (3 g/d), sodium chloride (3 g/d), or placebo. Linear mixed-effect models were used to estimate treatment effects. RESULTS: Potassium supplementation increased plasma phosphate (from 1.10 ± 0.19 to 1.15 ± 0.19 mmol/L, P = 0.004), in line with an increase in tubular maximum of phosphate reabsorption (from 0.93 ± 0.21 to 1.01 ± 0.20 mmol/L, P < 0.001). FGF23 decreased (114.3 [96.8-135.0] to 108.5 [93.5-125.9] RU/mL, P = 0.01), without change in parathyroid hormone and 25-hydroxy vitamin D3. Fractional calcium excretion decreased (from 1.25 ± 0.50 to 1.11 ± 0.46 %, P = 0.03) without change in plasma calcium. Sodium supplementation decreased both plasma phosphate (from 1.10 ± 0.19 to 1.06 ± 0.21 mmol/L, P = 0.03) and FGF23 (from 114.3 [96.8-135.0] to 108.7 [92.3-128.1] RU/mL, P = 0.02). Urinary and fractional calcium excretion increased (from 4.28 ± 1.91 to 5.45 ± 2.51 mmol/24 hours, P < 0.001, and from 1.25 ± 0.50 to 1.44 ± 0.54 %, P = 0.004, respectively). CONCLUSIONS: Potassium supplementation led to a decrease in FGF23, which was accompanied by increase in plasma phosphate and decreased calcium excretion. Sodium supplementation reduced FGF23, but this was accompanied by dec

A Virtual Testing Approach for Laminated Composites Based on Micromechanics

International audienceThe chapter deals with a crucial question for the design of composite structures: how can one predict the evolution of damage up to and including final fracture? Virtual testing, whose goal is to drastically reduce the huge number of industrial tests involved in current characterization procedures, constitutes one of today’s main industrial challenges. In this work, one revisits our multiscale modeling answer through its practical aspects. Some complements regarding identification, kinking, and crack initiation are also given. Finally, the current capabilities and limits of this approach are discussed, as well as the computational challenges that are inherent to “Virtual Structural Testing.