Sven Bossuyt

Spatiotemporally Distinct Protein Kinase D Activation in Adult Cardiomyocytes in Response to Phenylephrine and Endothelin

Protein kinase D (PKD) is a nodal point in cardiac hypertrophic signaling. It triggers nuclear export of class II histone deacetylase (HDAC) and regulates transcription. Although this pathway is thought to be critical in cardiac hypertrophy and heart failure, little is known about spatiotemporal aspects of PKD activation at the myocyte level. Here, we demonstrate that in adult cardiomyocytes two important neurohumoral stimuli that induce hypertrophy, endothelin-1 (ET1) and phenylephrine (PE), trigger comparable global PKD activation and HDAC5 nuclear export, but via divergent spatiotemporal PKD signals. PE-induced HDAC5 export is entirely PKD-dependent, involving fleeting sarcolemmal PKD translocation (for activation) and very rapid subsequent nuclear import. In contrast, ET1 recruits and activates PKD that remains predominantly sarcolemmal. This explains why PE-induced nuclear HDAC5 export in myocytes is totally PKD-dependent, whereas ET1-induced HDAC5 export depends more prominently on InsP3 and CaMKII signaling. Thus α-adrenergic and ET-1 receptor signaling via PKD in adult myocytes feature dramatic differences in cellular localization and translocation in mediating hypertrophic signaling. This raises new opportunities for targeted therapeutic intervention into distinct limbs of this hypertrophic signaling pathway.

Optimized Patterns for Digital Image Correlation

This work presents theoretical background on a novel class of strain sensor patterns. A combination of morphological image processing and Fourier analysis is used to characterize gray-scale images, according to specific criteria, and to synthesize patterns that score particularly well on these criteria. The criteria are designed to evaluate, with a single digital image of a pattern, the suitability of a series of images of that pattern for full-field displacement measurements by digital image correlation (DIC). Firstly, morphological operations are used to flag large featureless areas and to remove from consideration features too small to be resolved. Secondly, the autocorrelation peak sharpness radius en the autocorrelation margin are introduced to quantify the sensitivity and robustness, respectively, expected when using these images in DIC algorithms. For simple patterns these characteristics vary in direct proportion to each other, but it is shown how to synthesize a range of patterns with wide autocorrelation margins even though the autocorrelation peaks are sharp. Such patterns are exceptionally well-suited for DIC measurements.

A combined arc-melting and tilt-casting furnace for the manufacture of high-purity bulk metallic glass materials.

An arc-melting furnace which includes a tilt-casting facility was designed and built, for the purpose of producing bulk metallic glass specimens. Tilt-casting was chosen because reportedly, in combination with high-purity processing, it produces the best fatigue endurance in Zr-based bulk metallic glasses. Incorporating the alloying and casting facilities in a single piece of equipment reduces the amount of laboratory space and capital investment needed. Eliminating the sample transfer step from the production process also saves time and reduces sample contamination. This is important because the glass forming ability in many alloy systems, such as Zr-based glass-forming alloys, deteriorates rapidly with increasing oxygen content of the specimen. The challenge was to create a versatile instrument, in which high purity conditions can be maintained throughout the process, even when melting alloys with high affinity for oxygen. Therefore, the design provides a high-vacuum chamber to be filled with a low-oxygen inert atmosphere, and takes special care to keep the system hermetically sealed throughout the process. In particular, movements of the arc-melting electrode and sample manipulator arm are accommodated by deformable metal bellows, rather than sliding O-ring seals, and the whole furnace is tilted for tilt-casting. This performance of the furnace is demonstrated by alloying and casting Zr(55)Cu(30)Al(10)Ni(5) directly into rods up to ø 10 mm which are verified to be amorphous by x-ray diffraction and differential scanning calorimetry, and to exhibit locally ductile fracture at liquid nitrogen temperature.

Enhanced temperature uniformity by tetrahedral laser heating

emperature profile on a spherical sample that is heated by laser beams in various geometries while processed in vacuum is analyzed. Sample heating by one or four laser beams was considered. An analytical expression was derived for directional sample heating cases. It suggests an enhanced temperature uniformity over the samples when heated with four diffuse laser beams arranged in a tetrahedral geometry. This was experimentally verified by heating a spherical stainless steel sample by laser beams. Both the calculated and experimentally determined temperature variations over the sample suggest that use of diffuse four beams arranged in tetrahedral geometry would be effective in reducing temperature variation to within 1 K. The enhancement in the temperature uniformity for four diffuse beams arranged in a tetrahedral geometry by a factor of 50 over a single focused beam is promising to accurately measure of thermophysical properties. This drastic improvement in temperature uniformity might even enable atomic diffusion measurements in the undercooled liquid states of the bulk glass forming alloys since Marangoni and gravity driven convection will be substantially reduced.

β-Adrenergic Signaling Inhibits Gq-Dependent Protein Kinase D Activation by Preventing Protein Kinase D Translocation

Rationale: Both &bgr;-adrenergic receptor (&bgr;-AR) and Gq-coupled receptor (GqR) agonist–driven signaling play key roles in the events, leading up to and during cardiac dysfunction. How these stimuli interact at the level of protein kinase D (PKD), a nodal point in cardiac hypertrophic signaling, remains unclear. Objective: To assess the spatiotemporal dynamics of PKD activation in response to &bgr;-AR signaling alone and on coactivation with GqR-agonists. This will test our hypothesis that compartmentalized PKD signaling reconciles disparate findings of PKA facilitation and inhibition of PKD activation. Methods and Results: We report on the spatial and temporal profiles of PKD activation using green fluorescent protein-tagged PKD (wildtype or mutant S427E) and targeted fluorescence resonance energy transfer–based biosensors (D-kinase activity reporters) in adult cardiomyocytes. We find that &bgr;-AR/PKA signaling drives local nuclear activation of PKD, without preceding sarcolemmal translocation. We also discover pronounced interference of &bgr;-AR/cAMP/PKA signaling on GqR-induced translocation and activation of PKD throughout the cardiomyocyte. We attribute these effects to direct, PKA-dependent phosphorylation of PKD-S427. We also show that phosphomimetic substitution of S427 likewise impedes GqR-induced PKD translocation and activation. In neonatal myocytes, S427E inhibits GqR-evoked cell growth and expression of hypertrophic markers. Finally, we show altered S427 phosphorylation in transverse aortic constriction–induced hypertrophy. Conclusions: &bgr;-AR signaling triggers local nuclear signaling and inhibits GqR-mediated PKD activation by preventing its intracellular translocation. PKA-dependent phosphorylation of PKD-S427 fine-tunes the PKD responsiveness to GqR-agonists, serving as a key integration point for &bgr;-adrenergic and Gq-coupled stimuli.

Minimizing convection effects to measure diffusion in liquid droplets during high-temperature electrostatic levitation

We present an approach to reduce the convective flow in an electrostatically levitated liquid droplet to such an extent that diffusion is the dominant mechanism for mass transport, thus enabling direct measurements of atomic diffusion in reactive liquids at elevated temperatures. Convection is minimized by containerless processing, and reducing temperature variations in the sample. The diffusion tracer is deposited in situ in the electrostatic levitation device used for containerless processing. Uniform noncontact heating of the sample is achieved by laser heating with multiple beams arranged symmetrically, e.g., in a tetrahedral geometry. The expected temperature variations and the resulting convection flows are estimated for a Zr-based glass-forming alloy. The analysis suggests that diffusion experiments are possible throughout the entire undercooled liquid temperature range of this alloy and, in microgravity, up to 50 K above the liquidus temperature.

Localisation of plastic deformation in friction stir and electron beam copper welds

Optical strain measurement through digital image correlation during tensile testing was utilised to study the effect of strain rate and welding defects on localisation of plastic deformation in friction stir welded and electron beam welded phosphorous-alloyed oxygen-free copper. Welds cut from full-size nuclear waste copper canisters and additionally made defective welds were studied. Optical strain measurement represents a new way of studying deformation of materials and obtaining local material data. The data was used to construct strain maps and estimate local true stress–strain curves. Using Gini coefficients to quantify strain localisation, it appears that the lower strain rate resulted in slightly faster localisation of plastic deformation with lower stress level.

Damage detection in CFRP components using DIC

Unidirectional carbon fiber reinforced polymer composites (UD CFRP) are high performance materials for structural components, but they are very sensitive to damage. Structural health monitoring is therefore required in safety-critical applications. Many non-destructive evaluation techniques are not suitable for in-service monitoring, which calls for new approaches. We investigate the use of full-field digital image correlation (DIC) for detecting damage in UD CFRP components. Stereo-DIC data is used to analyze changes in vibration modes due to artificial defects. Finally, the effect of the defects is assessed and the suitability of the DIC method is evaluated.

A priori error estimate of the finite element solution to a Poisson inverse source problem

We study an example of the class of inverse source problems: to determine the source field of the Poisson equation from a finite number of density field measurements. Such problems are not well-posed, hence we consider reconstructions obtained using Tikhonov regularization. The regularized problem can in general only be solved using a numerical method, which introduces an approximation error. We consider approximations obtained using the finite element method and derive a priori error estimates for the numerical error. Our estimates allow different regularization terms, e.g. L2 and H1 regularization, and are expressed with respect to the finite element mesh sizes used to discretize the forward and the inverse problems. The novelty of our approach is that we consider finite measurement data, whereas the existing work assumes infinite dimensional measurements. The obtained results are illustrated by three numerical experiments. These examples cover situations where the solution regularity is high, as well as situations where the regularity is low. In addition, measurements in the interior of the domain and measurements on the boundary of the domain are both investigated. The numerical tests verify that the derived a priori error estimates work well in these different situations.

Shear Banding Observed in Real-Time with a Laser Speckle Method

In many high-strength materials, such as in metallic glasses, localized deformations can lead to quasi-brittle behavior where the amount of energy dissipated by mechanical deformation prior to material failure is small, as only a small volume fraction of the material deforms inelastically. Thus, techniques that allow observations of localized deformations at relevant length and time scales are instrumental in order to engineer such materials. Here, a real-time optical system based on high-speed imaging of laser speckle is applied to the propagation of shear bands in metallic glass. The quality and accuracy of information obtainable by this technique as well as its applicability to study important aspects of shear banding in metallic glasses are discussed and a comparison with more conventional approaches are presented.