Direct monitoring of the calcium concentration in the sarcoplasmic and endoplasmic reticulum of skeletal muscle myotubes

Direct monitoring of the free Ca2+ concentration in the sarcoplasmic reticulum (SR) was carried out in rat skeletal myotubes transfected with a specifically targeted aequorin chimera (srAEQ). Myotubes were also transfected with a chimeric aequorin (erAEQ) that we have demonstrated previously is retained in the endoplasmic reticulum (ER). Immunolocalization analysis showed that although both recombinant proteins are distributed in an endomembrane network identifiable with immature SR, the erAEQ protein was retained also in the perinuclear membrane. The difficulty of measuring [Ca2+] in 100-1000 microM range was overcome with the use of the synthetic coelenterazine analogue, coelenterazine n. We demonstrate that the steady state levels of [Ca2+] measured with srAEQ is around 300 microM, whereas that measured with erAEQ is significantly lower, i.e. around 200 microM. The effects of caffeine, high KCl, and nicotinic receptor stimulation, in the presence or absence of external calcium or after blockade of the Ca-ATPase, were investigated with both chimeras. The kinetics of [Ca2+] changes revealed by the erAEQ were similar, but not identical, neither quantitatively nor qualitatively, to those monitored with the srAEQ, indicating that at this stage of muscle development, differences exist between SR and ER in their mechanisms of Ca2+ handling. The functional implications of these findings are discussed.     

Vascular aging. Role of elastin receptor

A distinction is made between atheromatous plaque formation and arteriosclerosis, only this second process is strictly age-dependent. Interactions between lipids and constituents of the vascular wall are however involved in both processes, although by different mechanisms. The progressive increase with age of elastase activity is a second important factor in the age-dependent progression of arteriosclerosis. The fragmentation of elastic fibers produces elastin peptides, present in the circulating blood in microgram/ml conc-s, increasing in several arteriopathies. The constant presence of elastin peptides in the circulation maintains activated the elastin receptor (16) on endothelial cells producing NO* with vasorelaxing activity (18). The simultaneous production of superoxyde leads to the formation of peroxynitrate, neutralized by reduced glutathion. This process, maintained over decades may well impair the antiradical defense mechanisms of the cells and deprive the endothelium from the vasorelaxing activity of NO*. We propose therefore that the maintained activation of the elastin receptor with free radical and lytic enzyme production might well represent the initiating process underlying atherogenesis.     

Behavior of cylindrical tanks whose axes are horizontal

A cylindrical tank is mounted with its axis horizontal. Supports on either side run lengthwise. If the tank wall is thin, deformations significantly affect the stresses, so that standard stress formulae (which ignore deformations) may over-estimate stresses by a factor of ten or more. In this paper, deformations are accounted for by a nonlinear analysis. Stresses and deformed shapes are reported for various positions of the supports, radius to thickness ratios, and depths of fill.     

Use of the Extreme Groups Approach: A Critical Reexamination and New Recommendations.

Analysis of continuous variables sometimes proceeds by selecting individuals on the basis of extreme scores of a sample distribution and submitting only those extreme scores to further analysis. This sampling method is known as the extreme groups approach (EGA). EGA is often used to achieve greater statistical power in subsequent hypothesis tests. However, there are several largely unrecognized costs associated with EGA that must be considered. The authors illustrate the effects EGA can have on power, standardized effect size, reliability, model specification, and the interpretability of results. Finally, the authors discuss alternative procedures, as well as possible legitimate uses of EGA. The authors urge researchers, editors, reviewers, and consumers to carefully assess the extent to which EGA is an appropriate tool in their own research and in that of others. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Performance-Analyse und -Optimierung in der Softwareentwicklung

Nicht-funktionale Anforderungen an ein Softwaresystem beschreiben Aspekte, die nicht direkt die Funktionalit?t, wie sie der Benutzer sieht, betreffen.     

Quantitating Fluorescence Intensity from Fluorophore: The Definition of MESF Assignment

The quantitation of fluorescence radiance may at first suggest the need to obtain the number of fluorophore that are responsible for the measured fluorescence radiance. This goal is beset by many difficulties since the fluorescence radiance depends on three parameters 1) the probability of absorbing a photon (molar extinction), 2) the number of fluorophores, and 3) the probability of radiative decay of the excited state (quantum yield). If we use the same fluorophore in the reference solution and the analyte then, to a good approximation, the molar extinction drops out from the comparison of fluorescence radiance and we are left with the comparison of fluorescence yield which is defined as the product of fluorophore concentration and the molecular quantum yield. The equality of fluorescence yields from two solutions leads to the notion of equivalent number of fluorophores in the two solutions that is the basis for assignment of MESF (Molecules of Equivalent Soluble Fluorophore) values. We discuss how MESF values are assigned to labeled microbeads and by extension to labeled antibodies, and how these assignments can lead to the estimate of the number of bound antibodies in flow cytometer measurements.