Biomedical sciences increasingly recognise the importance of biophysics in health and disease. For the most part, this is due to an emerging new perspective of the broad impact of mechanobiology on the immune response. While most mechanisms of the immune response are adequately explained by cell-biology, biochemistry, and genetics, many of its features profoundly depend on biomechanical aspects.
The Biophysical Immunology (BPI) laboratory aims to unravel the impact of these mechanobiological aspect on the immune system in health and disease. Especially, we are interested in the biophysical functioning of the vital cellular cytoskeleton, the primary determinant of immune cell biophysics.
Having expertise across multiple disciplines from theoretical-physics to immunology, the BPI group is determined to carve out a niche and investigate the contentious biomedical research.
Mechanobiology in health and disease
Cellular interactions are often ruled by physical changes due to motion, so-called mechanical force. Such forces are involved during molecular rearrangements and most obviously when cells form close contacts, avoid each other, or reshape to change morphology. Mechanobiology focuses on the quantitative study of these mechanical forces in living cells.
A new perspective of mechanobiology is currently emerging across multiple disciplines in health and disease. In contrast to conventional beliefs, recent evidence
indicates that cells regulate their cell mechanics not only downstream of signalling events triggered by external stimuli, but that cells employ a diversity of feedback mechanisms enabling them to dynamically adjust their mechanics to meet physiological needs. Consequently, this provides an previously unforeseen picture wherein cells actively exert and resist biomechanical force to tune their mechano-biology and thus facilitate their function, which is particularly important during the
profound three-dimensional interactions in between cells or with their environment.
Quantifying cellular forces has therefore become an important mission across multiple disciplines at the interface of bioengineering, physics, and biomedical sciences. However, progress in measuring cellular forces has been hindered by the profound lack of a broadly-applicable force probing technology with the right spatio-temporal sensitivity. Historically, advances in understanding mechanisms of mechano-biology underlying key events of cytoskeletal actin organisation and their impact on immune cell function have been driven by innovations in technologies, that provide greater spatial and temporal sensitivity coupled to appropriate cell-biological, immunochemical, and genetic tools. Importantly, state-of-the-art microscopy has not been informative about real-time spatiotemporal dynamics of the actin cytoskeleton until recently.
To uncover the importance and broad impact of mechanobiology in health and disease, we apply and develop novel biophysical tools, force probing, and advanced live-cell super-resolution microscopies.
Research aims and objectives
The Biophysical Immunology (BPI) group led by Dr Marco Fritzsche is located within the MRC Human Immunology Unit (HIU) at the Weatherall Institute of Molecular Medicine (WIMM) and the Kennedy Institute for Rheumatology (KIR) at the University of Oxford, UK.
The BPI group is associated to the Micron Oxford Advanced Bioimaging Unit and works in close collaboration with the Nano-immunology-group of Professor Christian Eggeling located within the HIU and the Immunological-synapse-group of Professor Michael Dustin located at KIR.
Primary research aim is the identification and characterisation of mechanobiological mechanisms controlling immune cell activation in health and diease. Our objectives therefore involve the application and development of novel ultrasensitive, live-cell super-resolution microscopy techniques with spatiotemporal sensitivity down to the nanoscale, superior to conventional optical microscopes.
Research project: Nanoscale mechanical forces in the immune response
To-date the methodology of traction force microscopy (TFM) remains the leading force probing technology in the biomedical sciences. In such TFM experiments, mechanical force production is quantified by monitoring surface tractions produced by cells onto an elastic substrate of a given elasticity. Comprehensive mechanical measurements can only be achieved by very few methodologies, because mechanical quantifications depend on how they are executed. Especially, cell rheology, time-dependent mechanical properties, vastly differs at different time- and length-scales. On short time-scales (milli-seconds) and large length-scales (micro-meters) cells show poroelastic properties, and at long time-scales (minutes) they exhibit a power law behaviour in response to application of external forces. Hence, parameters such as displacements, cell tractions, and turnover rates must be monitored at a multitude of time- and length-scales in order to characterise biomechanical forces.
The greatest shortcoming of classical TFM is its limited sensitivity due to the finite density at which the displacement field can be sampled within the gel, which must be high enough to reflect the complexity of the traction field that is applied by the cell.
To overcome these challenges of classical TFM, we recently improved the spatial resolution and accuracy of TFM using optical super-resolution stimulated emission depletion (STED) microscopy. The increased spatial resolution of STED-TFM (STFM) allows a greater than five-fold higher sampling of the forces generated by the cell than conventional TFM, leading to more accurate quantification of cellular tractions. Please see detailed information for the development and application of the STFM.
Research project: Cytoskeletal actin dynamics in mechanobiology
Little is also known about the mechanisms controlling the organisation and dynamics of cytoskeletal actin structures in space and time in living immune cells.
Two fundamentally different mechanisms exist to generate actin networks: self-assembly and self-organisation. Self-assembly involves the physical association of motor proteins to actin, for instance, to assemble actin filaments into equilibrium networks. Self-organised actin networks are at steady state kept far from equilibrium by the constant input of energy. They arise solely through the local physical interactions of actin filaments initiated by a spontaneous instability such as symmetry breaking events, for example, when actin-polymerisation dynamics or intrinsic mechanical stress of actin networks spontaneously change.
Conclusive understanding of these rearrangements involves the tight coupling between biochemical and physical properties of three components of the actin cytoskeleton: (i) actin kinetics: actin protomer association/dissociation into filamentous actin (F-actin); (ii) actin dynamics: the assembly of filaments into actin structures; and (iii) mechanics: the cytoskeleton generates, senses, and transmits mechanical forces.
Cell-free studies involving computer-simulations and experiments in vitro have previously demonstrated how collective action of actin-associated proteins can organise actin filaments into dynamic patterns, such as vortices, asters and stars.
We have recently shown evidence of such self-organisation of the actin cortex in living cells. During cell adhesion, an active multistage process naturally leads to pattern transitions from actin vortices over stars into asters. The organisaiton of actin filaments (green in the video) is primarily driven by Arp2/3 complex (orange in the video) nucleation, but not by myosin motors, which is in contrast to what has been theoretically predicted and observed in vitro.
Research project: Calcium signalling in the immune response
As a second messenger, calcium plays a vital role in relaying information from cell-surface receptors into changes in cellular mechanisms, including cytoskeletal rearrangement. In T cells, calcium signalling is a key aspect in its development, survival, and activation. Given the transient, dynamic nature of calcium, a high temporal resolution is crucial in measuring calcium levels over time. And, while such studies have been done, they are typically done with only a few cells at a time. Because calcium responses can be heterogeneous between , it is imperative to measure many cells at once while still depicting individual calcium levels. CalQuo2 is recently updated user-friendly, MATLAB based software that can robustly analyses calcium patterns of individual cells within hundreds to thousands of cells that have been captured using fluorescence microscopy. Please see detailed information for the development and application of the calcium analysis software CalQuo.