Integrins Increase Sarcoplasmic Reticulum Activity for Excitation—Contraction Coupling in Human Stem Cell-Derived Cardiomyocytes

Engagement of the sarcoplasmic reticulum (SR) Ca²⁺ stores for excitation–contraction (EC)-coupling is a fundamental feature of cardiac muscle cells. Extracellular matrix (ECM) proteins that form the extracellular scaffolding supporting cardiac contractile activity are thought to play an integral role in the modulation of EC-coupling. At baseline, human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) show poor utilisation of SR Ca²⁺ stores, leading to inefficient EC-coupling, like developing or human CMs in cardiac diseases such as heart failure. We hypothesised that integrin ligand–receptor interactions between ECM proteins and CMs recruit the SR to Ca²⁺ cycling during EC-coupling. hiPSC-CM monolayers were cultured on fibronectin-coated glass before 24 h treatment with fibril-forming peptides containing the integrin-binding tripeptide sequence arginine–glycine–aspartic acid (2 mM). Micropipette application of 40 mM caffeine in standard or Na⁺/Ca²⁺-free Tyrode’s solutions was used to assess the Ca²⁺ removal mechanisms. Microelectrode recordings were conducted to analyse action potentials in current-clamp. Confocal images of labelled hiPSC-CMs were analysed to investigate hiPSC-CM morphology and ultrastructural arrangements in Ca²⁺ release units. This study demonstrates that peptides containing the integrin-binding sequence arginine–glycine–aspartic acid (1) abbreviate hiPSC-CM Ca²⁺ transient and action potential duration, (2) increase co-localisation between L-type Ca²⁺ channels and ryanodine receptors involved in EC-coupling, and (3) increase the rate of SR-mediated Ca²⁺ cycling. We conclude that integrin-binding peptides induce recruitment of the SR for Ca²⁺ cycling in EC-coupling through functional and structural improvements and demonstrate the importance of the ECM in modulating cardiomyocyte function in physiology.     

Observations of restricted beam-induced specimen motion with small-spot illumination

When lattice images of monolamellar paraffin crystals are recorded with a beam of diameter on the order of 1000 A, image contrast in part of the image is found to be at least three times higher than contrast in images obtained with conventional, flood illumination. Variations in contrast within these images, observed in previous experiments, are now found to be the result of radiation damage caused by the beam, which causes the crystal lattice to expand. This residual mode of beam-induced specimen motion, as well as distortions which propagate to adjacent, unirradiated areas of the specimen, are detected in a number of different ways. It thus appears that beam-induced specimen motion can be virtually eliminated over at least part of the illuminated area. Deleterious effects of the remaining mode of motion can be minimized by appropriate choice of exposure, beam-spot diameter, and spacing between exposed spots.     

Strength and electrical conductivity of a deformation-processed Cu-5 Pct Nb composite

A Cu-5 pct Nb alloy was deformation processed by wire drawing to very large reductions (99.9993 pct) and the strength and electrical conductivity properties compared with similarly deformation processed Cu-20 pct Nb. The results showed that the Cu-5 pct Nb alloy was transformed into a composite material with the original Nb dendrites becoming ribbonlike filaments in a similar fashion to higher Nb-containing Cu-Nb alloys. The degree of strength increase with increasing deformation processing greatly exceeds rule-of-mixtures expectations at higher degrees of deformation processing, where the Nb becomes highly aligned with the wire axis. A 5 pct Nb addition appears to contain close to the minimum amount of Nb phase necessary to produce appreciable strengthening during deformation processing of Cu-Nb alloys. The strength-conductivity properties of the deformation-processed Cu-5 pct Nb alloy show significant improvements in strength over the best commercial alloys in the conductivity range of 80 to 90 pct international annealed copper standard (IACS). This article is based on a presentation made in the symposium “High Performance Copper-Base Materials” as part of the 1991 TMS Annual Meeting, February 17–21, 1991, New Orleans, LA, under the auspices of the TMS Structural Materials Committee.     

FASTBUS Demonstration Systems

This paper will provide a demonstration of basic FASTBUS hardware and test software. The systems will include single crate segments, simple computer I/O, a fast sequencer and memory, some simple diagnostic and display devices and a UNIBUS to FASTBUS processor interface. The equipment will be set up to show the basic FASTBUS protocols and timing transactions, as well as some of the general initialization software features.     

Electron-crystallographic refinement of the structure of bacteriorhodopsin

Using electron diffraction data corrected for diffuse scattering together with additional phase information from 30 new images of tilted specimens, an improved experimental density map has been calculated for bacteriorhodopsin. The atomic model has then been rebuilt into this new map with particular attention to the surface loops. All the residues from 7 to 227 as well as ten lipid molecules are now included, although a few amino acid residues in three of the six surface loops, about half of the lipid hydrophobic chains and all of the lipid head groups are disordered. The model has then been refined against the experimental diffraction amplitudes to an R-factor of 28% at 3.5 angstrom resolution with strict geometry (0.005 angstrom) bond length deviation) using the improvement of the "free" phase residual between calculated and experimental phases from images as an objective criterion of accuracy. For the refinement some new programs were developed to restrain the number of parameters, to be compatible with the limited resolution of our data. In the final refined model of the protein (2BRD), compared with earlier co-ordinates (1BRD), helix D has been moved towards the cytoplasm by almost 4 angstrom, and the overall accuracy of the co-ordinates of residues in the other six helices has been improved. As a result the positions of nearly all the important residues in bacteriorhodopsin are now well determined. In particular, the buried, protonated Asp115 is 7 angstrom from, and so not in contact with, the retinal and Met118 forms a cap on the pocket occupied by the beta-ionone ring. No clear density exists for the side-chain of Arg82, which forms a central part of the extracellular half-channel. The only arginine side-chain built into good density is that of Arg134 at the extracellular end of helix E, the others being disordered near one of the two surfaces. The interpretation of the end of helix F on the extracellular surface is now clearer; an extra loose helical turn has been built bringing the side-chain of Glu194 close to Arg134 to form a probable salt bridge. The model provides an improved framework for understanding the mechanism of the light-driven proton pumping. A number of cavities that could contain water molecules were found by searching the refined model, most of them above or below the Schiff base in the half-channels leading to the two surfaces. The ordered and disordered regions of the structure are described by the temperature factor distribution.     

Freezing in sealed capillaries for preparation of frozen hydratedsections

We have investigated the freezing of specimens in a confined volume for preparation of vitreous samples for cryosectioning. With 15% dextran as a cryoprotectant, a sample sealed in a copper tube begins to freeze into crystalline ice when plunged into liquid ethane. Crystallization rapidly causes an increase in the pressure to the point that much of the sample freezes in a vitreous state. We used synchrotron X‐ray diffraction of samples frozen with various amounts of dextran to characterize the ice phases and crystal orientation, providing insights on the freezing process. We have characterized cryosections obtained from these samples to explore the optimum amount of cryoprotectant. Images of cryosectioned bacteria frozen with various levels of cryoprotectant illustrate effects of cryoprotectant concentration.     

Effect of Severe Image Compression on Iris Recognition Performance

We investigate three schemes for severe compression of iris images in order to assess what their impact would be on recognition performance of the algorithms deployed today for identifying people by this biometric feature. Currently, standard iris images are 600 times larger than the IrisCode templates computed from them for database storage and search; but it is administratively desired that iris data should be stored, transmitted, and embedded in media in the form of images rather than as templates computed with proprietary algorithms. To reconcile that goal with its implications for bandwidth and storage, we present schemes that combine region-of-interest isolation with JPEG and JPEG2000 compression at severe levels, and we test them using a publicly available database of iris images. We show that it is possible to compress iris images to as little as 2000 bytes with minimal impact on recognition performance. Only some 2% to 3% of the bits in the IrisCode templates are changed by such severe image compression, and we calculate the entropy per code bit introduced by each compression scheme. Error tradeoff curve metrics document very good recognition performance despite this reduction in data size by a net factor of 150, approaching a convergence of image data size and template size.     

Simple rainflow counting algorithms

Two simple algorithms for performing rainflow counting are presented in this paper. The second algorithm is suitable for microcomputer devices that are placed in vehicles to record field data.     

Effect of varactor Q-factor on tuning sensitivity of microwave oscillators, including reverse tuning

It is shown how the effect of resistive losses in a varactor diode can drastically affect the tuning characteristics of millimetre-wave oscillators and can even cause the oscillator to tune in the reverse direction.