Determination of reference genes for circadian studies in different tissues and mouse strains

<p>Abstract</p> <p>Background</p> <p>Circadian rhythms have a profound effect on human health. Their disruption can lead to serious pathologies, such as cancer and obesity. Gene expression studies in these pathologies are often studied in different mouse strains by quantitative real time polymerase chain reaction (qPCR). Selection of reference genes is a crucial step of qPCR experiments. Recent studies show that reference gene stability can vary between species and tissues, but none has taken circadian experiments into consideration.</p> <p>Results</p> <p>In the present study the expression of ten candidate reference genes (<it>Actb</it>, <it>Eif2a</it>, <it>Gapdh</it>, <it>Hmbs</it>, <it>Hprt1</it>, <it>Ppib</it>, <it>Rn18s</it>, <it>Rplp0</it>, <it>Tbcc </it>and <it>Utp6c</it>) was measured in 131 liver and 97 adrenal gland samples taken from three mouse strains (C57BL/6JOlaHsd, 129Pas plus C57BL/6J and <it>Crem </it>KO on 129Pas plus C57BL/6J background) every 4 h in a 24 h period. Expression stability was evaluated by geNorm and NormFinder programs. Differences in ranking of the most stable reference genes were observed both between individual mouse strains as well as between tissues within each mouse strain. We show that selection of reference gene (<it>Actb</it>) that is often used for analyses in individual mouse strains leads to errors if used for normalization when different mouse strains are compared. We identified alternative reference genes that are stable in these comparisons.</p> <p>Conclusions</p> <p>Genetic background and circadian time influence the expression stability of reference genes. Differences between mouse strains and tissues should be taken into consideration to avoid false interpretations. We show that the use of a single reference gene can lead to false biological conclusions. This manuscript provides a useful reference point for researchers that search for stable reference genes in the field of circadian biology.</p

Tresholds of blood variables obtained by receiver operating characteristic analysis for indication of fat and glycogen content in the liver of postpartum dairy cows

The objective was to determine optimal thresholds of blood variables that are associated with liver lipid and glycogen content in dairy cows. Liver specimens of 15 Holstein dairy cows were taken 2 weeks postpartum (PP) and used for determination of lipid and glycogen content, respectively. Blood samples were collected weekly, starting from week 2 antepartum (AP) up to week 2 PP. Selected blood parameters and body condition scoring (BCS) were evaluated through receiver operating characteristic curve analysis as indication of fat and glycogen content in the liver. Postpartal low liver glycogen content (mode 20%, stereological quantification of fat content in the liver tissue samples) can be indicated if values are higher than 3.5 for BCS at week 2 AP. Optimal time for indicating both low liver glycogen/high fat content is 1 week PP, with LDL cut off point set at 0.88 mmol/L. Established cut off values in this study can be used for indication of fat and glycogen content in the liver of dairy cows

Transition-State Interactions in a Promiscuous Enzyme: Sulfate and Phosphate Monoester Hydrolysis by Pseudomonas aeruginosa Arylsulfatase.

Pseudomonas aeruginosa arylsulfatase (PAS) hydrolyzes sulfate and, promiscuously, phosphate monoesters. Enzyme-catalyzed sulfate transfer is crucial to a wide variety of biological processes, but detailed studies of the mechanistic contributions to its catalysis are lacking. We present linear free energy relationships (LFERs) and kinetic isotope effects (KIEs) of PAS and analyses of active site mutants that suggest a key role for leaving group (LG) stabilization. In LFERs PASWT has a much less negative Brønsted coefficient (βleaving groupobs-Enz = -0.33) than the uncatalyzed reaction (βleaving groupobs = -1.81). This situation is diminished when cationic active site groups are exchanged for alanine. The considerable degree of bond breaking during the transition state (TS) is evidenced by an 18Obridge KIE of 1.0088. LFER and KIE data for several active site mutants point to leaving group stabilization by active site K375, in cooperation with H211. 15N KIEs and the increased sensitivity to leaving group ability of the sulfatase activity in neat D2O (Δβleaving groupH-D = +0.06) suggest that the mechanism for S-Obridge bond fission shifts, with decreasing leaving group ability, from charge compensation via Lewis acid interactions toward direct proton donation. 18Ononbridge KIEs indicate that the TS for PAS-catalyzed sulfate monoester hydrolysis has a significantly more associative character compared to the uncatalyzed reaction, while PAS-catalyzed phosphate monoester hydrolysis does not show this shift. This difference in enzyme-catalyzed TSs appears to be the major factor favoring specificity toward sulfate over phosphate esters by this promiscuous hydrolase, since other features are either too similar (uncatalyzed TS) or inherently favor phosphate (charge).BBSRC BB/I004327/1 EPSRC EP/E019390/1

Geospatial analysis and living urban geometry

This essay outlines how to incorporate morphological rules within the exigencies of our technological age. We propose using the current evolution of GIS (Geographical Information Systems) technologies beyond their original representational domain, towards predictive and dynamic spatial models that help in constructing the new discipline of "urban seeding". We condemn the high-rise tower block as an unsuitable typology for a living city, and propose to re-establish human-scale urban fabric that resembles the traditional city. Pedestrian presence, density, and movement all reveal that open space between modernist buildings is not urban at all, but neither is the open space found in today's sprawling suburbs. True urban space contains and encourages pedestrian interactions, and has to be designed and built according to specific rules. The opposition between traditional self-organized versus modernist planned cities challenges the very core of the urban planning discipline. Planning has to be re-framed from being a tool creating a fixed future to become a visionary adaptive tool of dynamic states in evolution

Digital representation of park use and visual analysis of visitor activities

Urban public parks can serve an important function by contributing to urban citizens' quality of life. At the same time, they can be the location of processes of displacement and exclusion. Despite this ambiguous role, little is known about actual park use patterns. To learn more about park use in three parks in Zurich, Switzerland, extensive data on visitor activities was collected using a new method based on direct recording via a portable GIS solution. Then, the data was analyzed using qualitative and quantitative methods. This paper examines whether geographic visualization of these data can help domain experts like landscape designers and park managers to assess park use. To maximize accessibility, the visualizations are made available through a web-interface of a common, off-the-shelf GIS. The technical limitations imposed by this choice are critically assessed, before the available visualization techniques are evaluated in respect to the needs and tasks of practitioners with limited knowledge on spatial analysis and GIS. Key criteria are each technique's level of abstraction and graphical complexity. The utility and suitability of the visualization techniques is characterized for the distinct phases of exploration, analysis and synthesis. The findings suggest that for a target user group of practitioners, a combination of dot maps showing the raw data and surface maps showing derived density values for several attributes serves the purpose of knowledge generation best

Patterns of using places for recreation and relaxation in peri-urban areas: The case of Lake Podpec, Slovenia

The paper addresses the issues of place capacity for occupancy for rest and recreation in the context of peri-urban landscapes of high natural or cultural values. It sets these issues in the frame of bottom-up approaches, considering qualitative-, small- and slow-data-oriented methods and techniques, using observation & behaviour mapping. It focuses on designing the concept of a natural site’s carrying capacity and evaluating an area’s carrying capacity for specific use. It proceeds from a study conducted by Golicnik Marušic (2015), which showed the prudence of addressing the development and planning of green infrastructure from the perspective of user needs and habits. By including the concept of an area’s carrying capacity, it also incorporates the conservation aspect. Based on data obtained through observation and mapping, the article comments on current use and users in terms of their age, use of a place, frequency of activities, and type of activities in the case of Lake Podpec, a popular recreational site at the fringe of Ljubljana marsh, Slovenia

Cardiac Arrest and Inverted Takotsubo Cardiomyopathy Following Intramyometrial Vasopressin Injection During Myomectomy

Vasopressin is involved in cardiovascular homeostasis that can influence coronary perfusion. It is commonly used as a local vasoconstricting agent during gynaecological procedures.We present a case of cardiac arrest and inverse Takotsubo features following intramyometrial vasopressin administration during myomectomy. The patient was successfully resuscitated and recovered completely. Cardiac presentation was compatible with inverse Takotsubo cardiomyopathy that could have been triggered by high-dose vasopressin-induced coronary vasoconstriction. The patient’s cardiac function resolved with no long-term sequelae.Takotsubo cardiomyopathy usually results from an excessive catecholaminergic surge. High-dose vasopressin-induced coronary vasospasm could have been the mechanism underlying the clinical presentation in our patient

Vzorci zasedbe prostora za potrebe sprostitve in rekreacije v obmestnem prostoru: primer Jezero pri Podpeci, Slovenija

Članek obravnava vprašanja o nosilni sposobnosti prostora za sprostitev in rekreacijo v obmestnih krajinah visokih naravnih ali kulturnih vrednot. Temelji na konceptu pristopov od spodaj navzgor, pri čemer uporablja metodo opazovanja in vedenjskih zemljevidov, ki temelji na drobnih kvalitativnih podatkih. Osredotoča se na zasnovo koncepta nosilne sposobnosti naravno vrednega obmocja za rabo in na ugotavljanje ocene nosilne sposobnosti takega območja za rabo. Izhajali smo iz Goličnik Marušic (2015), ki nakazuje smisel obravnave razvoja in načrtovanja zelene infrastrukture z vidika uporabnikovih potreb in navad ter s konceptom nosilne sposobnosti prostora za zasedbo vključili tudi varstveni vidik. Na podlagi izvedenih podatkov, pridobljenih z opazovanjem in kartiranjem vedenja, smo komentirali dejansko rabo, uporabnike z vidika starostnih skupin ter način zasedanja prostora, pogostost pojavljanja dejavnosti in načine zadrževanja v prostoru na primeru Jezera pri Podpeči, priljubljene izletniško-rekreacijske tocke na robu Ljubljanskega barja

The Human Cities Observation Mission: Research Tool for Public Space Anlyses and Evaluation

info:eu-repo/semantics/publishe