Konstantin D. Bergmeister

Alanyl–glutamine dipeptide restores the cytoprotective stress proteome of mesothelial cells exposed to peritoneal dialysis fluids

BACKGROUND Exposure of mesothelial cells to peritoneal dialysis fluids (PDF) results in cytoprotective cellular stress responses (CSR) that counteract PDF-induced damage. In this study, we tested the hypothesis that the CSR may be inadequate in relevant models of peritoneal dialysis (PD) due to insufficient levels of glutamine, resulting in increased vulnerability against PDF cytotoxicity. We particularly investigated the role of alanyl-glutamine (Ala-Gln) dipeptide on the cytoprotective PDF stress proteome. METHODS Adequacy of CSR was investigated in two human in vitro models (immortalized cell line MeT-5A and mesothelial cells derived from peritoneal effluent of uraemic patients) following exposure to heat-sterilized glucose-based PDF (PD4-Dianeal, Baxter) diluted with medium and, in a comparative proteomics approach, at different levels of glutamine ranging from depletion (0 mM) via physiological (0.7 mM) to pharmacological levels (8 mM administered as Ala-Gln). RESULTS Despite severe cellular injury, expression of cytoprotective proteins was dampened upon PDF exposure at physiological glutamine levels, indicating an inadequate CSR. Depletion of glutamine aggravated cell injury and further reduced the CSR, whereas addition of Ala-Gln at pharmacological level restored an adequate CSR, decreasing cellular damage in both PDF exposure systems. Ala-Gln specifically stimulated chaperoning activity, and cytoprotective processes were markedly enhanced in the PDF stress proteome. CONCLUSIONS Taken together, this study demonstrates an inadequate CSR of mesothelial cells following PDF exposure associated with low and physiological levels of glutamine, indicating a new and potentially relevant pathomechanism. Supplementation of PDF with pharmacological doses of Ala-Gln restored the cytoprotective stress proteome, resulting in improved resistance of mesothelial cells to exposure to PDF. Future work will study the clinical relevance of CSR-mediated cytoprotection.

Prosthetic reconstruction in high amputations of the upper extremity

ZusammenfassungHintergrundDie Steuerung myoelektrischer Armprothesen erfolgte konventionellerweise über zwei Oberflächenelektroden, welche von zwei getrennt innervierten Muskelgruppen angesteuert werden. Zwischen den verschiedenen prothetischen Gelenken wird mittels Kokontraktion dieser Muskeln gewechselt und in der jeweiligen Ebene mit denselben Muskeln linear gesteuert. Ein harmonischer, dem natürlichen Bewegungsmuster entsprechender Bewegungsablauf ist mit diesem Steuerungsmechanismus nicht möglich.FragestellungÜbersicht über die chirurgischen, therapeutischen und prothetischen Möglichkeiten bei hohen Amputationen der oberen Extremität.Material und MethodeEs erfolgte eine selektive Literaturrecherche unter Berücksichtigung eigener Erfahrungen des klinischen Alltags und Durchsicht von Patientenakten.ErgebnisseDurch selektive Nerventransfers der amputierten Armnerven des Plexus brachialis auf verbliebene Stumpfmuskulatur können bis zu sechs Signalgeber geschaffen werden, welche intuitiv und simultan die verschiedenen prothetischen Gelenke steuern können. Auf diese Weise ist eine effiziente und harmonische Steuerung der Prothese gewährleistet ohne dass der Patient zwischen den verschiedenen Steuerungsebenen wechseln muss. Gleichzeitig werden etwaige Neurome behandelt und somit ein schmerzfreies Tragen der Prothese ermöglicht. Aufgrund der dadurch vermehrten Verwendung von myoelektrischen Prothesen steigen auch die Anforderungen an den Stumpf. Hier gilt es sowohl chirurgisch als auch orthopädietechnisch eine stabile Verbindung zwischen Stumpf und Prothese zu schaffen, um eine optimale Prothesenfunktion zu ermöglichen.AbstractBackgroundConventional upper arm prostheses are controlled via two surface electrodes that measure motor activity of two separately innervated muscle groups. The various prosthetic joints are chosen by co-contractions and controlled linearly by these two muscles. A harmonious and natural course of movements is not possible in this way.ObjectivesOverview regarding surgical, therapeutical and prosthetic options in high amputations of the upper extremity.MethodsSelective literature research including the authors’ own experience in everyday clinical practice as well as a review of medical records.ResultsSelective nerve transfers of the amputated nerves of the brachial plexus to the remaining stump muscles can create up to six myosignals for intuitive and simultaneous control of the different prosthetic joints. In this way, an efficient and harmonious control of the prosthetic device is possible without the need to change between the different control levels. At the same time, possible neuromas are treated and painless wear of the prosthesis is achieved. Due to the resulting extended use of the prosthetic device, the demands regarding stump quality are increased. Thus, both surgically and by the means of the orthopedic technician a stable stump-socket connection should be achieved to enable optimal prosthetic function.

Prosthesis Control with an Implantable Multichannel Wireless Electromyography System for High-Level Amputees: A Large-Animal Study.

Background: Myoelectric prostheses lack a strong human-machine interface, leading to high abandonment rates in upper limb amputees. Implantable wireless electromyography systems improve control by recording signals directly from muscle, compared with surface electromyography. These devices do not exist for high amputation levels. In this article, the authors present an implantable wireless electromyography system for these scenarios tested in Merino sheep for 4 months. Methods: In a pilot trial, the electrodes were implanted in the hind limbs of 24 Sprague-Dawley rats. After 8 or 12 weeks, impedance and histocompatibility were assessed. In the main trial, the system was tested in four Merino sheep for 4 months. Impedance of the electrodes was analyzed in two animals. Electromyographic data were analyzed in two freely moving animals repeatedly during forward and backward gait. Results: Device implantation was successful in all 28 animals. Histologic evaluation showed a tight encapsulation after 8 weeks of 78.2 ± 26.5 µm subcutaneously and 92.9 ± 31.3 µm on the muscular side. Electromyographic recordings show a distinct activation pattern of the triceps, brachialis, and latissimus dorsi muscles, with a low signal-to-noise ratio, representing specific patterns of agonist and antagonist activation. Average electrode impedance decreased over the whole frequency range, indicating an improved electrode-tissue interface during the implantation. All measurements taken over the 4 months of observation used identical settings and showed similar recordings despite changing environmental factors. Conclusion: This study shows the implantation of this electromyography device as a promising alternative to surface electromyography, providing a potentially powerful wireless interface for high-level amputees.

Automated muscle fiber type population analysis with ImageJ of whole rat muscles using rapid myosin heavy chain immunohistochemistry.

Introduction: Skeletal muscle consists of different fiber types which adapt to exercise, aging, disease, or trauma. Here we present a protocol for fast staining, automatic acquisition, and quantification of fiber populations with ImageJ. Methods: Biceps and lumbrical muscles were harvested from Sprague‐Dawley rats. Quadruple immunohistochemical staining was performed on single sections using antibodies against myosin heavy chains and secondary fluorescent antibodies. Slides were scanned automatically with a slide scanner. Manual and automatic analyses were performed and compared statistically. Results: The protocol provided rapid and reliable staining for automated image acquisition. Analyses between manual and automatic data indicated Pearson correlation coefficients for biceps of 0.645–0.841 and 0.564–0.673 for lumbrical muscles. Relative fiber populations were accurate to a degree of ± 4%. Conclusions: This protocol provides a reliable tool for quantification of muscle fiber populations. Using freely available software, it decreases the required time to analyze whole muscle sections. Muscle Nerve 54: 292–299, 2016

Broadband Prosthetic Interfaces: Combining Nerve Transfers and Implantable Multichannel EMG Technology to Decode Spinal Motor Neuron Activity

Modern robotic hands/upper limbs may replace multiple degrees of freedom of extremity function. However, their intuitive use requires a high number of control signals, which current man-machine interfaces do not provide. Here, we discuss a broadband control interface that combines targeted muscle reinnervation, implantable multichannel electromyographic sensors, and advanced decoding to address the increasing capabilities of modern robotic limbs. With targeted muscle reinnervation, nerves that have lost their targets due to an amputation are surgically transferred to residual stump muscles to increase the number of intuitive prosthetic control signals. This surgery re-establishes a nerve-muscle connection that is used for sensing nerve activity with myoelectric interfaces. Moreover, the nerve transfer determines neurophysiological effects, such as muscular hyper-reinnervation and cortical reafferentation that can be exploited by the myoelectric interface. Modern implantable multichannel EMG sensors provide signals from which it is possible to disentangle the behavior of single motor neurons. Recent studies have shown that the neural drive to muscles can be decoded from these signals and thereby the users intention can be reliably estimated. By combining these concepts in chronic implants and embedded electronics, we believe that it is in principle possible to establish a broadband man-machine interface, with specific applications in prosthesis control. This perspective illustrates this concept, based on combining advanced surgical techniques with recording hardware and processing algorithms. Here we describe the scientific evidence for this concept, current state of investigations, challenges, and alternative approaches to improve current prosthetic interfaces.

A Rapid Automated Protocol for Muscle Fiber Population Analysis in Rat Muscle Cross Sections Using Myosin Heavy Chain Immunohistochemistry

Quantification of muscle fiber populations provides a deeper insight into the effects of disease, trauma, and various other influences on skeletal muscle composition. Various time-consuming methods have traditionally been used to study fiber populations in many fields of research. However, recently developed immunohistochemical methods based on myosin heavy chain protein expression provide a quick alternative to identify multiple fiber types in a single section. Here, we present a rapid, reliable and reproducible protocol for improved staining quality, allowing automatic acquisition of whole cross sections and automatic quantification of fiber populations with ImageJ. For this purpose, embedded skeletal muscles are cut in cross sections, stained using myosin heavy chains antibodies with secondary fluorescent antibodies and DAPI for cell nuclei staining. Whole cross sections are then scanned automatically using a slide scanner to obtain high-resolution composite pictures of the entire specimen. Fiber population analyses are subsequently performed to quantify slow, intermediate and fast fibers using an automated macro for ImageJ. We have previously shown that this method can identify fiber populations reliably to a degree of ±4%. In addition, this method reduces inter-user variability and time per analyses significantly using the open source platform ImageJ.

Ultrasound and shock-wave stimulation to promote axonal regeneration following nerve surgery: a systematic review and meta-analysis of preclinical studies

Limited regeneration after nerve injury often leads to delayed or incomplete reinnervation and consequently insufficient muscle function. Following nerve surgery, application of low-intensity ultrasound or extracorporeal shock waves may promote nerve regeneration and improve functional outcomes. Because currently clinical data is unavailable, we performed a meta-analysis following the PRISMA-guidelines to investigate the therapeutic effect of ultrasound and shock wave therapies on motor nerve regeneration. Ten ultrasound-studies (N = 445 rats) and three shock-wave studies (N = 110 rats) were identified from multiple databases. We calculated the difference in means or standardized mean difference with 95% confidence intervals for motor function, nerve conduction velocity and histomorphological parameters of treated versus sham or non-treated animals. Ultrasound treatment showed significantly faster nerve conduction, increased axonal regeneration with thicker myelin and improved motor function on sciatic functional index scale (week two: DM[95%CI]: 19,03[13,2 to 25,6], 71 animals; week four: 7,4[5,4 to 9,5], 47 animals). Shock wave induced recovery improvements were temporarily significant. In conclusion, there is significant evidence for low-intensity ultrasound but not for extracorporeal shock wave treatment to improve nerve regeneration. Prospective clinical trials should therefore investigate available FDA-approved ultrasound devices as adjunct postoperative treatment following nerve surgery.

Prospects of Neurorehabilitation Technologies Based on Robust Decoding of the Neural Drive to Muscles Following Targeted Muscle Reinnervation

Latest advances in neurorehabilitation technologies provide users with reliable mechatronic devices. Nonetheless, the control capabilities of these systems are limited to techniques that rely on indirect measures of neural information using EMG signals. We foresee that the combination of targeted muscle reinnervation (TMR) and high-density EMG electrodes supported by advanced blind source separation techniques (BSS) can substantially enhance current neurorehabilitation solutions. TMR provides access to the nerve activity by connecting nerves to muscles, used as biological amplifiers. Control would benefit from richer information content directly related to spinal motor neuron activity. The motor neuron firing statistics is obtained by applying advanced decomposition algorithms on the multi-channel EMGs from the targeted reinnervated muscles. It is expected that the control of these systems will be more dexterous and precise.

Fascicular shifting: a novel technique to overcome large nerve defects

OBJECTIVE Over the last decade, a number of authors have investigated the utility of different biological and synthetic matrices as alternatives to conventional nerve grafts. However, the autologous nerve graft remains the gold standard, even though it often involves using a pure sensory nerve to reconstruct a mixed or even a pure motor nerve. Furthermore, limited donor sites often necessitate a significant mismatch of needed nerve tissue, especially for large proximal nerve defects such as brachial plexus lesions. Here, the authors present a new technique that overcomes these problems: the fascicular shift procedure (FSP). A fascicular group of the nerve distal to the injury is harvested in a sufficient length to bridge the nerve defect. METHODS The method of fascicular shifting was tested at the sciatic nerve in 45 Lewis rats. In the experimental group, a 15-mm nerve defect was created and reconstructed with a fascicular group that was harvested directly distal to the gap. This group was compared with 1 negative control group (defect without reconstruction) and 3 positive control groups (sensory, motor, and mixed graft). After 12 weeks of nerve regeneration, outcome was evaluated using retrograde labeling, histomorphometric analysis, and muscle force analysis. RESULTS All reconstructed groups showed successful regeneration with various levels of function. The negative control group showed minimal force measurements that were of no functional value. The fascicular shift provided sufficient guidance to overcome nerve defects, had higher (p < 0.1) motor neuron counts (1958.75 ± 657.21) than the sensory graft (1263.50 ± 538.90), and was equal to motor grafts (1490.43 ± 794.80) and mixed grafts (1720.00 ± 866.421). This tendency of improved motor regeneration was confirmed in all analyses. The mixed graft group was compared with the experimental group to investigate the influence of the potential damage induced by the fascicular shift distal to the repair site. However, none of the analyses revealed an impairment of nerve regeneration for both the tibial and common peroneal index muscles. CONCLUSIONS This study demonstrates that harvesting a transplant from the nerve segment distal to the injury site offers a mixed graft without causing additional donor-site morbidity. These grafts perform statistically better than a standard sensory graft in terms of motor recovery. The fascicular shift presents a novel method to reconstruct large proximal nerve defects, making it immensely attractive in brachial plexus reconstruction.

Acceptance of animal research in our science community

Animal research is debated highly controversial, as evident by the “Stop Vivi-section” initiative in 2015. Despite widespread protest to the initiative by researchers, no data is available on the European medical research community’s opinion towards animal research. In this single-center study, we investigated this question in a survey of students and staff members at the Medical University of Vienna. A total of 906 participants responded to the survey, of which 82.8% rated the relevance of animal research high and 62% would not accept a treatment without prior animals testing. Overall, animal research was considered important, but its communication to the public considered requiring improvement.