SELECTIVE MEASUREMENT OF α SMOOTH MUSCLE ACTIN: WHY β-ACTIN CAN NOT BE USED AS A HOUSEKEEPING GENE WHEN TISSUE FIBROSIS OCCURS

Abstract Background Prevalence of fibroproliferative diseases, including chronic kidney disease is rapidly increasing and has become a major public health problem worldwide. Fibroproliferative diseases are characterized by increased expression of α smooth muscle actin (α-SMA) that belongs to the family of the six conserved actin isoforms showing high degree homology. The aim of the present study was to develop real-time PCRs that clearly discriminate α-SMA and ß-actin from other actin isoforms. Results Real-time PCRs using self-designed mouse, human and rat specific α-SMA or ß-actin primer pairs resulted in the specific amplification of the artificial DNA templates corresponding to mouse, human or rat α-SMA or ß-actin, however ß-actin showed cross-reaction with the housekeeping γ-cyto-actin. We have shown that the use of improperly designed literary primer pairs significantly affects the results of PCRs measuring mRNA expression of α-SMA or ß-actin in the kidney of mice underwent UUO. Conclusion We developed a set of carefully designed primer pairs and PCR conditions to selectively determine the expression of mouse, human or rat α-SMA and ß-actin isoforms. We demonstrated the importance of primer specificity in experiments where the results are normalized to the expression of ß-actin especially when fibrosis and thus increased expression of α-SMA is occur

Lattice QCD at the physical point: Simulation and analysis details

We give details of our precise determination of the light quark masses m_{ud}=(m_u+m_d)/2 and m_s in 2+1 flavor QCD, with simulated pion masses down to 120 MeV, at five lattice spacings, and in large volumes. The details concern the action and algorithm employed, the HMC force with HEX smeared clover fermions, the choice of the scale setting procedure and of the input masses. After an overview of the simulation parameters, extensive checks of algorithmic stability, autocorrelation and (practical) ergodicity are reported. To corroborate the good scaling properties of our action, explicit tests of the scaling of hadron masses in N_f=3 QCD are carried out. Details of how we control finite volume effects through dedicated finite volume scaling runs are reported. To check consistency with SU(2) Chiral Perturbation Theory the behavior of M_\pi^2/m_{ud} and F_\pi as a function of m_{ud} is investigated. Details of how we use the RI/MOM procedure with a separate continuum limit of the running of the scalar density R_S(\mu,\mu') are given. This procedure is shown to reproduce the known value of r_0m_s in quenched QCD. Input from dispersion theory is used to split our value of m_{ud} into separate values of m_u and m_d. Finally, our procedure to quantify both systematic and statistical uncertainties is discussed.Comment: 45 page

Multimode fibre:Light-sheet microscopy at the tip of a needle

We also thank the UK Engineering and Physics Sciences Research Council for funding under grant EP/J01771X/1. Finally, we would like to thank EXCELLENT TEAMS (CZ.1.07/2.3.00/30.0005) from European Social Fund and CEITEC - Central European Institute of Technology (CZ.1.05/1.1.00/02.0068) from European Regional Development Fund for support.Light-sheet fluorescence microscopy has emerged as a powerful platform for 3-D volumetric imaging in the life sciences. Here, we introduce an important step towards its use deep inside biological tissue. Our new technique, based on digital holography, enables delivery of the light-sheet through a multimode optical fibre - an optical element with extremely small footprint, yet permitting complex control of light transport processes within. We show that this approach supports some of the most advanced methods in light-sheet microscopy: by taking advantage of the cylindrical symmetry of the fibre, we facilitate the wavefront engineering methods for generation of both Bessel and structured Bessel beam plane illumination. Finally, we assess the quality of imaging on a sample of fluorescent beads fixed in agarose gel and we conclude with a proof-of-principle imaging of a biological sample, namely the regenerating operculum prongs of Spirobranchus lamarcki.Publisher PDFPeer reviewe

Test System Stability and Natural Variability of a Lemna Gibba L. Bioassay

BACKGROUND: In ecotoxicological and environmental studies Lemna spp. are used as test organisms due to their small size, rapid predominantly vegetative reproduction, easy handling and high sensitivity to various chemicals. However, there is not much information available concerning spatial and temporal stability of experimental set-ups used for Lemna bioassays, though this is essential for interpretation and reliability of results. We therefore investigated stability and natural variability of a Lemna gibba bioassay assessing area-related and frond number-related growth rates under controlled laboratory conditions over about one year. METHODOLOGY/PRINCIPAL FINDINGS: Lemna gibba L. was grown in beakers with Steinberg medium for one week. Area-related and frond number-related growth rates (r(area) and r(num)) were determined with a non-destructive image processing system. To assess inter-experimental stability, 35 independent experiments were performed with 10 beakers each in the course of one year. We observed changes in growth rates by a factor of two over time. These did not correlate well with temperature or relative humidity in the growth chamber. In order to assess intra-experimental stability, we analysed six systematic negative control experiments (nontoxicant tests) with 96 replicate beakers each. Evaluation showed that the chosen experimental set-up was stable and did not produce false positive results. The coefficient of variation was lower for r(area) (2.99%) than for r(num) (4.27%). CONCLUSIONS/SIGNIFICANCE: It is hypothesised that the variations in growth rates over time under controlled conditions are partly due to endogenic periodicities in Lemna gibba. The relevance of these variations for toxicity investigations should be investigated more closely. Area-related growth rate seems to be more precise as non-destructive calculation parameter than number-related growth rate. Furthermore, we propose two new validity criteria for Lemna gibba bioassays: variability of average specific and section-by-section segmented growth rate, complementary to average specific growth rate as the only validity criterion existing in guidelines for duckweed bioassays

Improved Xenobiotic Metabolism and Reduced Susceptibility to Cancer in Gluten-Sensitive Macaques upon Introduction of a Gluten-Free Diet

A non-human primate (NHP) model of gluten sensitivity was employed to study the gene perturbations associated with dietary gluten changes in small intestinal tissues from gluten-sensitive rhesus macaques (Macaca mulatta).Stages of remission and relapse were accomplished in gluten-sensitive animals by administration of gluten-free (GFD) and gluten-containing (GD) diets, as described previously. Pin-head-sized biopsies, obtained non-invasively by pediatric endoscope from duodenum while on GFD or GD, were used for preparation of total RNA and gene profiling, using the commercial Rhesus Macaque Microarray (Agilent Technologies),targeting expression of over 20,000 genes.When compared with normal healthy control, gluten-sensitive macaques showed differential gene expressions induced by GD. While observed gene perturbations were classified into one of 12 overlapping categories--cancer, metabolism, digestive tract function, immune response, cell growth, signal transduction, autoimmunity, detoxification of xenobiotics, apoptosis, actin-collagen deposition, neuronal and unknown function--this study focused on cancer-related gene networks such as cytochrome P450 family (detoxification function) and actin-collagen-matrix metalloproteinases (MMP) genes.A loss of detoxification function paralleled with necessity to metabolize carcinogens was revealed in gluten-sensitive animals while on GD. An increase in cancer-promoting factors and a simultaneous decrease in cancer-preventing factors associated with altered expression of actin-collagen-MMP gene network were noted. In addition, gluten-sensitive macaques showed reduced number of differentially expressed genes including the cancer-associated ones upon withdrawal of dietary gluten. Taken together, these findings indicate potentially expanded utility of gluten-sensitive rhesus macaques in cancer research

Why do you drink caffeine? The development of the Motives for Caffeine Consumption Questionnaire (MCCQ) and its relationship with gender, age and the types of caffeinated beverages

Caffeine is the most popular psychoactive substance that is consumed worldwide. As motives influence behavior, investigation of the motivational background of caffeine consumption should help provide a better understanding of the popularity of caffeinated products. The present study aimed (i) to explore and operationalize the motives of caffeine consumption and (ii) to reveal possible differences in the motives regarding gender, age and the type of caffeinated products consumed. Motives for caffeine consumption were collected from regular caffeine consumers (N = 26) and were informed by a review of the relevant literature. Following this, a cross-sectional study was conducted on a convenience sample of Hungarian university students and working adults (N = 598). The participants completed the Motives for Caffeine Consumption Questionnaire and the Caffeine Consumption Questionnaire. Six motivational factors were identified: Alertness, Habit, Mood, Social, Taste and Symptom Management. Women had higher scores on Habit, Social, Taste and Symptom Management. Younger participants had higher scores on Alertness than the older group, and the older group had higher scores on Habit and Symptom Management. Five types of caffeine users were identified. Those who consumed (i) coffee, (ii) tea, (iii) energy drinks, (iv) coffee and tea and (v) mixed drinks. Several differences between the five groups were revealed across all motives except for Taste. The present study developed a robust psychometric instrument for assessing caffeine consumption motives. The factors varied in importance in relation to gender, age and caffeine consumption habits

Target Identification for Stereotactic Thalamotomy Using Diffusion Tractography

BACKGROUND: Stereotactic targets for thalamotomy are usually derived from population-based coordinates. Individual anatomy is used only to scale the coordinates based on the location of some internal guide points. While on conventional MR imaging the thalamic nuclei are indistinguishable, recently it has become possible to identify individual thalamic nuclei using different connectivity profiles, as defined by MR diffusion tractography. METHODOLOGY AND PRINCIPAL FINDINGS: Here we investigated the inter-individual variation of the location of target nuclei for thalamotomy: the putative ventralis oralis posterior (Vop) and the ventral intermedius (Vim) nucleus as defined by probabilistic tractography. We showed that the mean inter-individual distance of the peak Vop location is 7.33 mm and 7.42 mm for Vim. The mean overlap between individual Vop nuclei was 40.2% and it was 31.8% for Vim nuclei. As a proof of concept, we also present a patient who underwent Vop thalamotomy for untreatable tremor caused by traumatic brain injury and another patient who underwent Vim thalamotomy for essential tremor. The probabilistic tractography indicated that the successful tremor control was achieved with lesions in the Vop and Vim respectively. CONCLUSIONS: Our data call attention to the need for a better appreciation of the individual anatomy when planning stereotactic functional neurosurgery

A brief report on the associations amongst social media use, gender, and body esteem in a UK student sample

Research into the effects of social media on personal wellbeing have been controversial in recent years, with recent research highlighting links between social media use and body esteem. This conceptual replication study aimed to explore relationships amongst social media use, body esteem and gender amongst UK university students (n=100). Participants completed measures of social media use and body image esteem. It was hypothesised that social media intensity and usage would negatively predict body esteem, with high social media intensity relating to lower body esteem. We find that gender was the only significant predictor of body esteem, with women having lower body esteem compared to men. We were unable to replicate previous findings, as our findings show no relationships amongst social media intensity, use, and body esteem