Cell motility in microfluidic environments

Trypanosomes are single-celled bloodstream parasites and causative agents of African Sleeping Sickness in humans and Nagana disease in domestic livestock. The pathogen is transmitted by the bite of the tsetse fly and lethal to the infected host if left untreated. Eponymous for the Trypanosoma genus name (Trypanosoma: drilling body ) is the striking nature of their movement which is often described by the spiral motion of a corkscrew. However, looking at trypanosomes at high spatiotemporal resolution, we find that the way the cells move is more complex than described before and changes over time and cell. Apart from the question how trypanosomes move, we find ourselves confronted with the question why trypanosomes move at all? In this context, M. Engstler et al. have shown that active movement is essential for cells that are exposed to hostile antibodies. Hydrodynamic flow induced by active movement of the cell leads to a delocalization of antibodies that have bound to the cell surface: Antibodies exposed to the flow around a forward swimming cell are driven backwards into the flagellar pocket where they are taken up by endocytosis and rendered harmless by subsequent digestion. In contrary, a backward swimming cell is accumulating antibodies at the tip of the flagellum and gets digested itself by the host s immune system. If the described mechanism of hydrodynamic protein sorting is a ubiquitous feature in nature, it has to be proven in more detailed studies of cell motility as well as the involved hydrodynamic condition.The aim of this thesis is to study and quantify the movement of trypanosomes in their microfluidic environments in order to help understanding the mechanisms and reasons of their motility. To achieve this goal we constructed an optical trapping fluorescence microscope optimized for high spatiotemporal resolution and reduced phototoxicity. In combination with advanced microfluidic methods we were not only able to control hydrodynamic flow conditions and spatial confinement, but also to position, manipulate and measure forces on the single cell level, as well as to specifically label single living cells in the microflow.In this work we could show for the first time that using strongly focussed diode lasers it is possible to optically trap living trypanosomes over time scales of t < 15 min, without inducing significant photodamage. The optical stall forces acting on trypanosomes were determined and used to measure the propagation forces of single and dividing trypanosomes.In combination with automated image processing routines we also analyzed the positioning of trypanosomes within the optical trap and found distinct trapping loci which could be correlated to structural features of trypanosomes

Photo-Controlled Dynamics and Transport in Entangled Wormlike Micellar Nanocomposites Studied by XPCS

Dynamics of nanoparticles (NPs) in microscopic networks, in particular, localization and transport, play a key role in designing new functional nanocomposites and drug delivery systems. To this aim, it is crucial to understand the interplay between the network structure and dynamics on the microscopic scale which determines NP diffusion. Here, we study the localization and transport of spherical NPs in photorheological wormlike micellar nanocomposites where the mobility of the NPs is controlled by the network mesh size and the micelle length, which can be tuned by UV-illumination. The macroscopic viscoelastic properties are measured by classical rheology, while X-ray photon correlation spectroscopy and nanorheology provide information on the microscopic NP dynamics on length scales on the order of the network mesh size. On long time scales, the data reveal that transport through the network is determined by the ratio between the NP size and the network mesh size, while upon UV illumination, the NP mobility is drastically enhanced. On shorter time scales, the influence of the dynamical and structural micelle properties on the NP dynamics under confinement is explored and indicates an anomalous speed-up of the dynamics, which is discussed in the context of changes in the local structure and non-linear phenomena such as strain stiffening and hopping motion

Trypanosome motion represents an adaptation to the crowded environment of the vertebrate bloodstream

Blood is a remarkable habitat: it is highly viscous, contains a dense packaging of cells and perpetually flows at velocities varying over three orders of magnitude. Only few pathogens endure the harsh physical conditions within the vertebrate bloodstream and prosper despite being constantly attacked by host antibodies. African trypanosomes are strictly extracellular blood parasites, which evade the immune response through a system of antigenic variation and incessant motility. How the flagellates actually swim in blood remains to be elucidated. Here, we show that the mode and dynamics of trypanosome locomotion are a trait of life within a crowded environment. Using high-speed fluorescence microscopy and ordered micro-pillar arrays we show that the parasites mode of motility is adapted to the density of cells in blood. Trypanosomes are pulled forward by the planar beat of the single flagellum. Hydrodynamic flow across the asymmetrically shaped cell body translates into its rotational movement. Importantly, the presence of particles with the shape, size and spacing of blood cells is required and sufficient for trypanosomes to reach maximum forward velocity. If the density of obstacles, however, is further increased to resemble collagen networks or tissue spaces, the parasites reverse their flagellar beat and consequently swim backwards, in this way avoiding getting trapped. In the absence of obstacles, this flagellar beat reversal occurs randomly resulting in irregular waveforms and apparent cell tumbling. Thus, the swimming behavior of trypanosomes is a surprising example of micro-adaptation to life at low Reynolds numbers. For a precise physical interpretation, we compare our high-resolution microscopic data to results from a simulation technique that combines the method of multi-particle collision dynamics with a triangulated surface model. The simulation produces a rotating cell body and a helical swimming path, providing a functioning simulation method for microorganism with a complex swimming strategy