Flexibility in the receptor-binding domain of the enzymatic colicin E9 is required for toxicity against Escherichia coli cells

The events that occur after the binding of the enzymatic E colicins to Escherichia coli BtuB receptors that lead to translocation of the cytotoxic domain into the periplasmic space and, ultimately, cell killing are poorly understood. It has been suggested that unfolding of the coiled-coil Mull receptor binding domain of the E colicins may be an essential step that leads to the loss of immunity protein from the colicin and immunity protein complex and then triggers the events of translocation. We introduced pairs of cysteine mutations into the receptor binding domain of colicin E9 (ColE9) that resulted in the formation of a disulfide bond located near the middle or the top of the R domain. After dithiothreitol reduction, the ColE9 protein with the mutations L359C and F412C (ColE9 L359C-F412C) and the ColE9 protein with the mutations Y324C and L447C (ColE9 Y324C-L447C) were slightly less active than equivalent concentrations of ColE9. On oxidation with diamide, no significant biological activity was seen with the ColE9 L359C-F412C and the ColE9 Y324C-L447C mutant proteins; however diamide had no effect on the activity of ColE9. The presence of a disulfide bond was confirmed in both of the oxidized, mutant proteins by matrix-assisted laser desorption ionization-time of flight mass spectrometry. The loss of biological activity of the disulfide-containing mutant proteins was not due to an indirect effect on the properties of the translocation or DNase domains of the mutant colicins. The data are consistent with a requirement for the flexibility of the coiled-coil R domain after binding to BtuB

Fast catheter segmentation from echocardiographic sequences based on segmentation from corresponding X-ray fluoroscopy for cardiac catheterization interventions

© 2014 IEEE. Echocardiography is a potential alternative to X-ray fluoroscopy in cardiac catheterization given its richness in soft tissue information and its lack of ionizing radiation. However, its small field of view and acoustic artifacts make direct automatic segmentation of the catheters very challenging. In this study, a fast catheter segmentation framework for echocardiographic imaging guided by the segmentation of corresponding X-ray fluoroscopic imaging is proposed. The complete framework consists of: 1) catheter initialization in the first X-ray frame; 2) catheter tracking in the rest of the X-ray sequence; 3) fast registration of corresponding X-ray and ultrasound frames; and 4) catheter segmentation in ultrasound images guided by the results of both X-ray tracking and fast registration. The main contributions include: 1) a Kalman filter-based growing strategy with more clinical data evalution; 2) a SURF detector applied in a constrained search space for catheter segmentation in ultrasound images; 3) a two layer hierarchical graph model to integrate and smooth catheter fragments into a complete catheter; and 4) the integration of these components into a system for clinical applications. This framework is evaluated on five sequences of porcine data and four sequences of patient data comprising more than 3000 X-ray frames and more than 1000 ultrasound frames. The results show that our algorithm is able to track the catheter in ultrasound images at 1.3 s per frame, with an error of less than 2 mm. However, although this may satisfy the accuracy for visualization purposes and is also fast, the algorithm still needs to be further accelerated for real-time clinical applications

Design and integration of a parallel, soft robotic end-effector for extracorporeal ultrasound

Objective: In this work we address limitations in state-of-the-art ultrasound robots by designing and integrating a novel soft robotic system for ultrasound imaging. It employs the inherent qualities of soft fluidic actuators to establish safe, adaptable interaction between ultrasound probe and patient. Methods: We acquire clinical data to determine the movement ranges and force levels required in prenatal foetal ultrasound imaging and design the soft robotic end-effector accordingly. We verify its mechanical characteristics, derive and validate a kinetostatic model and demonstrate controllability and imaging capabilities on an ultrasound phantom. Results: The soft robot exhibits the desired stiffness characteristics and is able to reach 100% of the required workspace when no external force is present, and 95% of the workspace when considering its compliance. The model can accurately predict the end-effector pose with a mean error of 1.18+/-0.29mm in position and 0.92+/-0.47deg in orientation. The derived controller is, with an average position error of 0.39mm, able to track a target pose efficiently without and with externally applied loads. Ultrasound images acquired with the system are of equally good quality compared to a manual sonographer scan. Conclusion: The system is able to withstand loads commonly applied during foetal ultrasound scans and remains controllable with a motion range similar to manual scanning. Significance: The proposed soft robot presents a safe, cost-effective solution to offloading sonographers in day-to-day scanning routines. The design and modelling paradigms are greatly generalizable and particularly suitable for designing soft robots for physical interaction tasks

Three-Axis Fiber-Optic Body Force Sensor for Flexible Manipulators

This paper proposes a force/torque sensor structure that can be easily integrated with a flexible manipulator structure. The sensor’s ring-like structure with its hollow inner section provides ample space for auxiliary components, such as cables and tubes, to be passed through and, hence, is very suitable for integration with tendon-driven and fluid-actuated manipulators. The sensor structure can also accommodate the wiring for a distributed sensor system as well as for diagnostic instruments that may be incorporated in the manipulator. Employing a sensing approach based on optical fibers as done here allows for the creation of sensors that are free of electrical currents at the point of sensing and immune to magnetic fields. These sensors are inherently safe when used in the close vicinity of humans and their measuring performance is not impaired when they are operated in or nearby machines such as magnetic resonance imaging (MRI) scanners. This type of sensor concept is particularly suitable for inclusion in instruments and robotic tools for minimally invasive surgery (MIS). The paper summarizes the design, integration challenges and calibration of the proposed optical three-axis force sensor. The experimental results confirm the effectiveness of our optical sensing approach and show that after calibrating its stiffness matrix, force and momentum components can be determined accurately

Image-based view-angle independent cardiorespiratory motion gating and coronary sinus catheter tracking for x-ray-guided cardiac electrophysiology procedures

Determination of the cardiorespiratory phase of the heart has numerous applications during cardiac imaging. In this article we propose a novel view-angle independent near-real time cardiorespiratory motion gating and coronary sinus (CS) catheter tracking technique for x-ray fluoroscopy images that are used to guide cardiac electrophysiology procedures. The method is based on learning CS catheter motion using principal component analysis and then applying the derived motion model to unseen images taken at arbitrary projections, using the epipolar constraint. This method is also able to track the CS catheter throughout the x-ray images in any arbitrary subsequent view. We also demonstrate the clinical application of our model on rotational angiography sequences. We validated our technique in normal and very low dose phantom and clinical datasets. For the normal dose clinical images we established average systole, end-expiration and end-inspiration gating success rates of 100%, 85.7%, and 92.3%, respectively. For very low dose applications, the technique was able to track the CS catheter with median errors not exceeding 1 mm for all tracked electrodes. Average gating success rates of 80.3%, 71.4%, and 69.2% were established for the application of the technique on clinical datasets, even with a dose reduction of more than 10 times. In rotational sequences at normal dose, CS tracking median errors were within 1.2 mm for all electrodes, and the gating success rate was 100%, for view angles from RAO 90° to LAO 90°. This view-angle independent technique can extract clinically useful cardiorespiratory motion information using x-ray doses significantly lower than those currently used in clinical practice

Immunity protein release from a cell-bound nuclease colicin complex requires global conformational rearrangement

Nuclease colicins bind their target receptor BtuB in the outer membrane of sensitive Escherichia coli cells in the form of a high-affinity complex with their cognate immunity proteins. The release of the immunity protein from the colicin complex is a prerequisite for cell entry of the colicin and occurs via a process that is still relatively poorly understood. We have previously shown that an energy input in the form of the cytoplasmic membrane proton motive force is required to promote immunity protein (Im9) release from the colicin E9/Im9 complex and colicin cell entry. We report here that engineering rigidity in the structured part of the colicin translocation domain via the introduction of disulfide bonds prevents immunity protein release from the colicin complex. Reduction of the disulfide bond by the addition of DTT leads to immunity protein release and resumption of activity. Similarly, the introduction of a disulfide bond in the DNase domain previously shown to abolish channel formation in planar bilayers also prevented immunity protein release. Importantly, all disulfide bonds, in the translocation as well as the DNase domain, also abolished the biological activity of the Im9-free colicin E9, the reduction of which led to a resumption of activity. Our results show, for the first time, that conformational flexibility in the structured translocation and DNase domains of a nuclease colicin is essential for immunity protein release, providing further evidence for the hypothesis that global structural rearrangement of the colicin molecule is required for disassembly of this high-affinity toxin-immunity protein complex prior to outer membrane translocation

Surface flattening of the human left atrium and proof-of-concept clinical applications

Surface flattening in medical imaging has seen widespread use in neurology and more recently in cardiology to describe the left ventricle using the bull's-eye plot. The method is particularly useful to standardize the display of functional information derived from medical imaging and catheter-based measurements. We hypothesized that a similar approach could be possible for the more complex shape of the left atrium (LA) and that the surface flattening could be useful for the management of patients with atrial fibrillation (AF). We implemented an existing surface mesh parameterization approach to flatten and unfold 3D LA models. Mapping errors going from 2D to 3D and the inverse were investigated both qualitatively and quantitatively using synthetic data of regular shapes and computer tomography scans of an anthropomorphic phantom. Testing of the approach was carried out using data from 14 patients undergoing ablation treatment for AF. 3D LA meshes were obtained from magnetic resonance imaging and electroanatomical mapping systems. These were unfolded using the developed approach and used to demonstrate proof-of-concept applications, such as the display of scar information, electrical information and catheter position. The work carried out shows that the unfolding of complex cardiac structures, such as the LA, is feasible and has several potential clinical uses for the management of patients with AF.</p