Non-equilibrium dynamics of biological matter in microfluidic environments - from red blood cell flickering to conformational transitions of actin filaments

Even the most basic and seemingly simple living biological systems exist far from thermodynamic equilibrium and studying their dynamic behavior represents a crucial step towards a better understanding of their fundamental properties. Here, we present the investigations of two different, out-of-equilibrium biological systems, namely single cell studies on the human red blood cell (RBC) and single macromolecule analysis of actin filaments, both by exploiting the exceptional physics at the micro-scale and the corresponding unique capabilities of experimental control. In particular, existing approaches to RBC analysis on the single-cell level usually rely on chemical or physical manipulations that often cause difficulties with preserving the RBC's integrity in a controlled microenvironment. We introduce a straightforward, self-filling microfluidic device that autonomously separates and isolates single RBCs directly from unprocessed human blood samples and confines them in diffusion-controlled microchambers by solely exploiting their unique intrinsic properties. Using bright-field microscopy, this noninvasive approach enables the time-resolved analysis of RBC flickering during the reversible shape evolution from the discocyte to the echinocyte morphology. A better understanding of this central shape transformation is especially relevant for blood storage applications as the formation of echinocytes can affect blood handling. We are further able to study the photo-induced oxygenation cycle of single functional RBCs by Raman microscopy without the limitations typically observed in optical tweezers based methods. Due to its specialized geometry, our device is particularly suited for studies on single RBCs under precise control of their environment. The provision of important insights into the RBC's biomedical and biophysical properties will improve the understanding of RBC microcirculation and can further contribute to advances in pathology diagnosis. Furthermore, we study the non-equilibrium conformational dynamics of semiflexible actin filaments experiencing hydrodynamic forces. Improving the knowledge about these dynamic processes of semiflexible polymers is of particular importance for the description of unusual transport in cellular flows and pattern formation processes in cytoplasmic streaming. The actin filaments are flowing through structured microchannels with alternating high- and low-velocity segments. These flow fields of spatially varying flow strength result in a compressive force on the filaments when they are entering the low-velocity regions and conversely an extensional force is acting on them when they are reentering the high-velocity segments. The semiflexible actin filaments undergo a length-dependent buckling transition under compression with a corresponding change in end-to-end distance and a rise in bending energy. However, the degree of increase of the length-normalized bending energy shows no evident dependence on the contour length. Increasing the fluid flow velocity results in a large rise of the compressive hydrodynamic force with a strong increase in storage of elastic energy due to the bending of the semiflexible filaments. At the passage from the low-velocity segments to the high-velocity ones, an extensional force is acting on the partially elastically relaxed filaments and a conformational transition from a coiled to a stretched state with a suppression of thermal fluctuations can be observed. Despite the symmetry of the microfluidic channels and therefore a similar rate of the absolute values of extension or compression in the specific channel segments, the observed stretch-coil and coil-stretch transitions distinctly differ in the evolution of the conformational changes and bending energies. This asymmetry of the non-equilibrium and non-stationary conformational transitions shows a strong dependence on the contour and persistence length, the degree of relaxation and the extensional or compressional rate. Many polymer solutions are non-Newtonian fluids and our studies may therefore have an impact on the analysis as well as sorting of polymers by elucidating the non-Newtonian flow behavior of semiflexible filaments in specific microflows, which may consequently lead to a better understanding of intercellular flows

Non-invasive removal of sandblasted and acid-etched titanium palatal implants, a retrospective study

BACKGROUND Short, rough-surfaced palatal implants are an established and reliable anchor for orthodontic treatment. Until recently, removal was only possible surgically using a hollow cylinder trephine. This standard method retrieves the implant combined with a larger bone volume and is therefore considered invasive and has known complications. Lately, an explantation tool which allows a sufficient force application to break the bone-implant-connection and unscrew the palatal implant was developed and, since its introduction, has been used as the method of choice in several orthodontic offices. OBJECTIVES The aim of this study was to assess the complications caused by removing rough-surfaced palatal implants simply by unscrewing them with an explantation tool in contrast to standard protocol by surgical removal with a trephine. MATERIAL AND METHODS The removal of 73 palatal implants using a customized explantation tool has been evaluated retrospectively and was compared to an existing sample of 44 conventional surgical explantations. RESULTS The new clinical procedure resulted in successful removal of 71 (97.3 per cent) palatal implants. In two cases, the new method failed but removal with the established surgical method was still possible with no further complications. The non-invasive palatal implant removal with a customized explantation tool had less medical complications compared to an existing sample of surgical explantations. CONCLUSIONS User's opinion was that the new method is more easily executed, less invasive, and also applicable without local anaesthesia. Therefore, it is considered to be beneficial for patients and the treatment approach of choice. However, further research is needed for verificatio

A self-filling microfluidic device for noninvasive and time-resolved single red blood cell experiments

Existing approaches to red blood cell (RBC) experiments on the single-cell level usually rely on chemical or physical manipulations that often cause difficulties with preserving the RBC's integrity in a controlled microenvironment. Here, we introduce a straightforward, self-filling microfluidic device that autonomously separates and isolates single RBCs directly from unprocessed human blood samples and confines them in diffusion-controlled microchambers by solely exploiting their unique intrinsic properties. We were able to study the photo-induced oxygenation cycle of single functional RBCs by Raman microscopy without the limitations typically observed in optical tweezers based methods. Using bright-field microscopy, our noninvasive approach further enabled the time-resolved analysis of RBC flickering during the reversible shape evolution from the discocyte to the echinocyte morphology. Due to its specialized geometry, our device is particularly suited for studying the temporal behavior of single RBCs under precise control of their environment that will provide important insights into the RBC's biomedical and biophysical properties

Live cell X-ray imaging of autophagic vacuoles formation and chromatin dynamics in fission yeast

Seeing physiological processes at the nanoscale in living organisms without labeling is an ultimate goal in life sciences. Using X-ray ptychography, we explored in situ the dynamics of unstained, living fission yeast Schizosaccharomyces pombe cells in natural, aqueous environment at the nanoscale. In contrast to previous X-ray imaging studies on biological matter, in this work the eukaryotic cells were alive even after several ptychographic X-ray scans, which allowed us to visualize the chromatin motion as well as the autophagic cell death induced by the ionizing radiation. The accumulated radiation of the sequential scans allowed for the determination of a characteristic dose of autophagic vacuole formation and the lethal dose for fission yeast. The presented results demonstrate a practical method that opens another way of looking at living biological specimens and processes in a time-resolved label-free setting

Event-Related Potentials Reveal Rapid Verification of Predicted Visual Input

Human information processing depends critically on continuous predictions about upcoming events, but the temporal convergence of expectancy-based top-down and input-driven bottom-up streams is poorly understood. We show that, during reading, event-related potentials differ between exposure to highly predictable and unpredictable words no later than 90 ms after visual input. This result suggests an extremely rapid comparison of expected and incoming visual information and gives an upper temporal bound for theories of top-down and bottom-up interactions in object recognition

Perspectives of teachers and pupils on the concert band as a picture puzzle: A qualitative interview study

Many schools have concert bands; to date, however, little research has been conducted on the experience of participants tin this kind of music lesson. In the frame-work of a qualitative interview study, I focused on accessing and contrasting the various perspectives of general music teachers, instrumental instructors and pupils on concert bands (“Bläserklassen”). The study adheres to grounded theory methodology, reconstructing both the conceptual assumptions of the teachers of three different concert bands and the discrepancies emerging in a comparison of the perspectives. Strikingly, participants’ perspectives on the concert bands proved to diverge significantly. This article argues for integrating this stated ambiguity within the theory of music pedagogy and using it to positively further the didactic development of what is offered in the classroom situation. (DIPF/Orig.

Adaptation of planning in music learning situations in light of qualitative interviews

How do teachers handle unexpected situations in music lessons, and what do pupils notice about their teachers’ planning? These questions were examined in qualitative interviews with teachers and pupils as part of a research project focusing on the adaptiveness of learning situations in the subject of music. This article outlines the main finding that teachers take into consideration learning goals, planning, their perception of the situation and the adaptation of planning. They constantly focus on their pupils’ learning process; their planning takes place in the situation. Based on their many observations, teachers try to create lessons as optimally as possible to achieve their learning goals. Often students hardly notice this cyclical process. In the course of the interviews, it became clear that flexible planning is part of adaptive teaching. (DIPF/Orig.

Noninvasive palatal implant removal

OBJECTIVE Orthodontic palatal implants are commonly used and do provide reliable absolute anchorage to assist orthodontic treatment. However, once treatment is completed, removal of these temporary implants is not considered easy or risk free. This short communication presents a clinical case in which a novel noninvasive procedure was applied to remove an osseointegrated palatal implant. MATERIAL AND METHODS A customized explantation tool, tightly fixed to the implant and precisely grasping the implant's head, was used in combination with a ratchet to unscrew the implant instead of the traditional removal by trephine. RESULTS Only a topical anesthetic was necessary before the implant-bone contact was broken by turning the ratchet counterclockwise. The implant was retrieved without any local anesthesia. The explanted palatal implant had no bone appending to it, except in its apical anti-rotational grooves, and the healing process thereafter was unproblematic. CONCLUSIONS Noninvasive palatal implant removal offers a simple and fast approach for explantation. Moreover, it might reduce the risk of adverse patient reactions, iatrogenic tooth and nerve injuries, and possible oro-antral communications

Editor’s Note

publishedVersio

Non-invasive removal of sandblasted and acid-etched titanium palatal implants, a retrospective study

BACKGROUND Short, rough-surfaced palatal implants are an established and reliable anchor for orthodontic treatment. Until recently, removal was only possible surgically using a hollow cylinder trephine. This standard method retrieves the implant combined with a larger bone volume and is therefore considered invasive and has known complications. Lately, an explantation tool which allows a sufficient force application to break the bone-implant-connection and unscrew the palatal implant was developed and, since its introduction, has been used as the method of choice in several orthodontic offices. OBJECTIVES The aim of this study was to assess the complications caused by removing rough-surfaced palatal implants simply by unscrewing them with an explantation tool in contrast to standard protocol by surgical removal with a trephine. MATERIAL AND METHODS The removal of 73 palatal implants using a customized explantation tool has been evaluated retrospectively and was compared to an existing sample of 44 conventional surgical explantations. RESULTS The new clinical procedure resulted in successful removal of 71 (97.3 per cent) palatal implants. In two cases, the new method failed but removal with the established surgical method was still possible with no further complications. The non-invasive palatal implant removal with a customized explantation tool had less medical complications compared to an existing sample of surgical explantations. CONCLUSIONS User's opinion was that the new method is more easily executed, less invasive, and also applicable without local anaesthesia. Therefore, it is considered to be beneficial for patients and the treatment approach of choice. However, further research is needed for verification