Submitochondrial distribution of three key steroidogenic proteins (steroidogenic acute regulatory protein and cytochrome p450scc and 3beta-hydroxysteroid dehydrogenase isomerase enzymes) upon stimulation by intracellular calcium in adrenal glomerulosa cells

In adrenal glomerulosa cells, angiotensin II (Ang II) and potassium stimulate aldosterone synthesis through activation of the calcium messenger system. The rate-limiting step in steroidogenesis is the transfer of cholesterol to the inner mitochondrial membrane. This transfer is believed to depend upon the presence of the steroidogenic acute regulatory (StAR) protein. The aim of this study was 1) to examine the effect of changes in cytosolic free calcium concentration and of Ang II on intramitochondrial cholesterol and 2) to study the distribution of StAR protein in submitochondrial fractions during activation by Ca2+ and Ang II. To this end, freshly prepared bovine zona glomerulosa cells were submitted to a high cytosolic Ca2+ clamp (600 nM) or stimulated with Ang II (10 nM) for 2 h. Mitochondria were isolated and subfractionated into outer membranes, inner membranes (IM), and contact sites (CS). Stimulation of intact cells with Ca2+ or Ang II led to a marked, cycloheximide-sensitive increase in cholesterol in CS (to 143 +/- 3. 2 and 151.1 +/- 18.1% of controls, respectively) and in IM (to 119 +/- 5.1 and 124.5 +/- 6.5% of controls, respectively). Western blot analysis revealed a cycloheximide-sensitive increase in StAR protein in mitochondrial extracts of Ca2+-clamped glomerulosa cells (to 159 +/- 23% of controls). In submitochondrial fractions, there was a selective accumulation of StAR protein in IM following stimulation with Ca2+ (228 +/- 50%). Similarly, Ang II increased StAR protein in IM, and this effect was prevented by cycloheximide. In contrast, neither Ca2+ nor Ang II had any effect on the submitochondrial distribution of cytochrome P450scc and 3beta-hydroxysteroid dehydrogenase isomerase. The intramitochondrial presence of the latter enzyme was further confirmed by immunogold staining in rat adrenal fasciculata cells and by immunoblot analysis in MA-10 mouse testicular Leydig cells. These findings demonstrate that under acute stimulation with Ca2+-mobilizing agents, newly synthesized StAR protein accumulates in IM after transiting through CS. Moreover, our results suggest that the import of StAR protein into IM may be associated with cholesterol transfer, thus promoting precursor supply to the two first enzymes of the steroidogenic cascade within the mitochondria and thereby activating mineralocorticoid synthesis.     

Calcium stimulates intramitochondrial cholesterol transfer in bovine adrenal glomerulosa cells

In adrenal glomerulosa cells, angiotensin II (Ang II) stimulates aldosterone synthesis through rises of cytosolic calcium ([Ca2+]c). The rate-limiting step in this process is the transfer of cholesterol to the inner mitochondrial membrane, where it is converted to pregnenolone by the P450 side chain cleavage enzyme. The aim of the present study was to examine the effect of changes in [Ca2+]c and of Ang II on intramitochondrial cholesterol distribution. Freshly prepared bovine zona glomerulosa cells were submitted to a cytosolic Ca2+ clamp (600 nM) or stimulated with Ang II (10 nM). Mitochondria were isolated and subfractionated into outer membranes (OM), inner membranes (IM), and contact sites (CS). Cholesterol content was determined by the cholesterol oxidase assay. Stimulation of intact cells with Ca2+ led to a marked decrease in cholesterol content of OM (to 54 +/- 24% of controls, n = 5) and to a concomitant increase of cholesterol in CS and IM (to 145 +/- 14%, n = 5). When glomerulosa cells were exposed to Ang II, a marked increase of cholesterol in CS occurred (to 172 +/- 16% of controls, n = 5). No significant changes were detected in OM cholesterol, suggesting a stimulation of cholesterol supply to the mitochondria in response to Ang II. Cycloheximide specifically and significantly reduced Ca2+-activated cholesterol transfer to CS and IM. In conclusion, our data indicate that one of the main functions of the Ca2+ messenger is to increase cholesterol supply to the P450 side chain cleavage enzyme by enhancing endogenous intermembrane cholesterol transfer to a mitochondrial site containing the enzymes responsible for the initial steps of the steroidogenic cascade.     

Three phases in the system Hg1 − xCxBa2CuOy (0.00 ≤ x ≤ 1.00)

Samples of the [Hg_{1 − x}C_x]-Ba₂CuOy system [(Hg,C)-1201] with stoichiometries within the range 0.00≤x≤1.00 were synthesized by a highpressure, high-temperature technique. The sample with X = 0.00 [HgBa₂CuO_4+δ] was non-superconducting with lattice parameters A = 3.8676(2) Å, C = 9.470(1) Å. At low ranges of substitution (0.10≤x≤0.30), the predominant phase was found to be Hg-1201 with identical lattice parameters to those found for X = 0.00. At X = 0.40 the Hg-1201 phase coexists in similar proportions with the phase of mixed [Hg_{1 − x},C_x]-1201 stoichiometry. At X = 0.50, this second phase with lattice parameters A = 3.9271(3) Å, C = 8.676(2) Å is predominant and only traces of the Hg-1201 phase were found. At X = 0.75 the [Hg_{1 − x},C_x]-1201 phase found for X = 0.50 coexists with the CO₃Ba₂CuO_y phase, which is the extreme of the substitution range. This latter was predominant at X = 1.00 with lattice parameters A = 4.0044(3) Å, C = 7.909(1) Å, but at higher pressure than the previous substitutions. This system is constituted of three main phases, HgBa₂CuO_{4 + δ}, Hg_{1 − x}C_xBa₂CuO_y (x ≈ 0.50) and CO₃Ba₂CuO_y, which appear in different proportions according to the percentage of substitution and the applied synthesis pressure.     

Requirement-oriented risk management for incremental software development

In incremental software development (ISD) functionalities are delivered incrementally and requirements keep on evolving across iterations. The requirements evolution involves the addition of new dependencies and conflicts among functional and non-functional requirements along with changes in priorities and dependency weights. This, in turn, demands refactoring the order of development of system components to minimize the impact of these changes. Neglecting the non-functional constraints in the software development process exposes it to risks that may accumulate across several iterations. In this research work, we propose a risk management framework for ISD processes that provides an estimate of risk exposure for the project when functional features are frozen while ignoring the associations with non-functional requirements. Our framework proposes suitable risk reduction strategies that work in tandem with the risk assessment module. We also provide a tool interface for our risk management framework.

     

Time-domain Analysis of Laboratory Experiments on the Transient Pressure Damping in a Leaky Polymeric Pipe

Water Resources Management - The pressure damping occurring in pressurized pipes with a leak during transients has been examined as a diagnostic tool – the so-called transient damping method...     

A single pass algorithm for clustering evolving data streams based on swarm intelligence

Existing density-based data stream clustering algorithms use a two-phase scheme approach consisting of an online phase, in which raw data is processed to gather summary statistics, and an offline phase that generates the clusters by using the summary data. In this article we propose a data stream clustering method based on a multi-agent system that uses a decentralized bottom-up self-organizing strategy to group similar data points. Data points are associated with agents and deployed onto a 2D space, to work simultaneously by applying a heuristic strategy based on a bio-inspired model, known as flocking model. Agents move onto the space for a fixed time and, when they encounter other agents into a predefined visibility range, they can decide to form a flock if they are similar. Flocks can join to form swarms of similar groups. This strategy allows to merge the two phases of density-based approaches and thus to avoid the computing demanding offline cluster computation, since a swarm represents a cluster. Experimental results show that the bio-inspired approach can obtain very good results on real and synthetic data sets.     

Landscape-aware location-privacy protection in location-based services

Mobile network providers have developed a variety of location-based services (LBSs), such as friend-finder, point of interest services, emergency rescue and many other safety and security services. The protection of location-privacy has consequently become a key aspect to the success of LBSs, since users consider their own physical location and movements highly privacy-sensitive, and demand for solutions able to protect such an information in a variety of environments. The idea behind location-privacy protection is that the individual should be able to set the level at which the location information is released to avoid undesired exploitation by a potential attacker: one of the approaches to this problem is given by the application of spatial obfuscation techniques, actuated by a trusted agent, and consisting in artificial perturbations of the location information collected by sensing technologies, before its disclosure to third parties. In many situations, however, landscape/map information can help a third party to perform Bayesian inference over spatially obfuscated data and to refine the user’s location estimate up to a violation of the original user’s location-privacy requirements. The goal of this paper is to provide a map-dependent obfuscation procedure that enables the release of the maximum possible user’s location information, that does not lead to a violation of the original user’s location-privacy requirements, even when refined through map-based inference.     

Real time identification of discrete event systems using Petri nets

The paper defines the identification problem for Discrete Event Systems (DES) as the problem of inferring a Petri Net () model using the observation of the events and the available output vectors, that correspond to the markings of the measurable places. Two cases are studied considering different levels of the system knowledge. In the first case the place and transition sets are assumed known. Hence, an integer linear programming problem is defined in order to determine a modelling the DES. In the second case the transition and place sets are assumed unknown and only an upper bound of the number of places is given. Hence, the identification problem is solved by an identification algorithm that observes in real time the occurred events and the corresponding output vectors. The integer linear programming problem is defined at each observation so that the can be recursively identified. Some results and examples characterize the identified systems and show the flexibility and simplicity of the proposed technique. Moreover, an application to the synthesis of supervisory control of systems via monitor places is proposed.     

A convex optimization approach to filtering in jump linear systems with state dependent transitions

We introduce a new methodology to construct a Gaussian mixture approximation to the true filter density in hybrid Markovian switching systems. We relax the assumption that the mode transition process is a Markov chain and allow it to depend on the actual and unobservable state of the system. The main feature of the method is that the Gaussian densities used in the approximation are selected as the solution of a convex programming problem which trades off sparsity of the solution with goodness of fit. A meaningful example shows that the proposed method can outperform the widely used interacting multiple model (IMM) filter and GPB2 in terms of accuracy at the expenses of an increase in computational time.     

AC brushless drive with low-resolution Hall-effect sensors for surface-mounted PM Machines

An ac brushless drive in which Hall-effect sensors are used as rotor position sensors is presented in this paper. Three different methods to obtain a high-resolution position estimation from the low-resolution sensors are described and compared through simulation and experimental testing. The proposed control algorithm's most innovative feature is its adaptability to the entire speed range, including startup, when using any of the three estimation algorithms. The control algorithm has been implemented and tested in order to drive a slotless axial-flux permanent-magnet (PM) machine for domestic appliance applications.