MicroRobotic toolkit to deliver spatiotemporally resolved physicochemical signals and control cell sociology

Periodic Reporting for period 4 - ROBOCHIP (MicroRobotic toolkit to deliver spatiotemporally resolved physicochemical signals and control cell sociology)

Berichtszeitraum: 2021-09-01 bis 2022-08-31

The primary scientific objective of the project is the discovery of physical principles behind multicellular organization that takes place during morphogenesis and self-healing. Understanding how external mechanical loads are intertwined with the cell-mediated forces to jointly control the formation and maintenance of tissue shape informs tissue engineering strategies and creation of artificial biological machines. The distinguishing features of our technological approach are the fine control, extreme dexterity, high-throughput, and multi-dimensionality. We have developed microrobotic technologies integrated with state-of-the-art microscopes that allow application of spatiotemporally resolved dynamic stimuli and real-time imaging. From the scientific perspective, our work serves as a test-bed for some of the most intriguing questions in development biology, mechanobiology, and regenerative medicine. What are the governing equations that relate the magnitude, timing and direction of force generation to the tissue mechanical properties, such as fibre alignment, viscosity and stiffness? How far, over what time, and in what geometry locally generated forces are transmitted? How cells maintain tissue boundaries, when they decide to move and expand the tissue and how the equilibrium achieved after an injury or change in boundary conditions are other important questions we are trying to address. Our aim was to quantitatively describe how matrix architecture and mechanics influence transmission of forces, and elucidate principles of physical organization instantiated by remodeling of a fibrillar substrate. Answering this question would allow us to solve the inverse problem: setting the initial conditions and applying controlled dynamic mechanical signals so that, at the equilibrium state, the engineered tissue would attain the desired morphology. Regulation of mechanical signals and cues has implications in many aspects of physiology and pathology including cancer, immunology, cardiology, and orthopaedics. Our technology is important to study cells and tissues associated with all these systems and develop new therapies to address malfunctioning in mechanotransduction pathways. In addition to the contributions to healthcare, our project has important educational value. A synergistic collaboration among different disciplines including mechanics, microtechnology, materials science, robotics and biomedical engineering is essential to ensure the success of this project. The principal investigator created two new unique masters courses at the host institution, one on microrobotic technologies and another on mechanobiology. The lecture notes and exercises are freely accessible by researchers at other institutes and public.

We have built compact, dexterous and modular robotic micromanipulation systems to provide the ability to manipulate cells within tissues. The first platform consists of a custom-design six-degrees-of-freedom robotic system that has submicron positioning precision. We used microelectromechanical systems technology to form arrays of free-standing 3D fibrous microtissues. We then performed robot-assisted microsurgery, such as local incisions and implantation, to study how the tissues change shape over time. We showed both experimentally and by using computer simulations that cells at the tissue boundary develop surface stresses and, together with contractile activity of cells residing in the core, drive macroscale deformation and tissue shaping. We discovered a way of stabilizing surface stresses as energy minimization at the surfaces leads to deviation from the prescribed shape. The process involves solid to fluid transitions induced by mechanical perturbations, generates spatially distributed surface stresses at interfaces, and is amenable to both additive and subtractive manufacturing. Second, we combined the rapid and wireless response of hybrid optomechanical nanomaterials with state-of-the-art microengineering techniques to develop machinery that can apply physiologically relevant mechanical loading at the cell scale. Our data provided novel insights on the characteristics of mechanical signals that lead to collective cell motion or matrix remodelling, and eventually to changes in tissue morphology. Third, we developed a platform that allows application of spatiotemporally defined matrix deformations using magnetically controlled microactuators. We recorded spatial and temporal changes in the distribution and shape of cultured cells along with the topological map of the fibre network using time-lapse confocal microscopy. We constructed an experimentally validated finite element model that can report stresses. Application of well-defined forces to the fibres facilitates the characterization of the constitutive properties of the structures. The combined experimental and computational approach has revealed important insights on the characteristics of mechanical stimuli along with fibre properties on the tissue architecture. Finally, we introduced a design and fabrication methodology for frequency addressable acoustic actuation of 3D nanoprinted hydrogel devices. We demonstrated remotely controlled operation of cell-sized devices with multiple pumps decorated with valves, filters, and chambers. We built analytical and numerical models that aid the design of the machines, calculate their resonance frequencies and modes, and provide quantitative information on their predicted performance. All the structures are printed as a monolithic piece from biocompatible hydrogels, enabling integration of the devices with living tissues.

The microrobotic toolkit that we have developed allows us to explore many aspects of mechanobiology that were previously inaccessible. The automated micromanipulation platforms will soon become instrumental to handle and operate on engineered tissues, organoids, and embryos. Performing protocols with robotic agents will democratise microsurgery and increase standardization of procedures across laboratories We used microfabrication, computational mechanics, light-sheet microscopy, and our robotic micromanipulation platforms to show that collagen gels covered with a contiguous epithelial sheet can be freely shaped using mechanical forces. This discovery opens up new avenues of research in tissue engineering with the hope that one day tissues developed in the lab will have the proper form and function to be implanted into a patient or used for testing therapies. The technology we developed also holds significant potential for translational medicine and targeted therapy. Our work introduced delivery methods that can not only release its contents at the target site in a dose-dependent manner, but also contain and protect the payload during transport. More specifically, we made two seminal contributions to the field. First, we have developed acoustically powered microscopic devices that can noninvasively collect biological samples from remote pathological sites and make on site measurements for diagnostic purposes or release biologics at target locations. Second, we discovered that ultraflexible slender instruments can harness power from biological flows such as cardiovascular and respiratory flows. This discovery is groundbreaking as we can now navigate microengineered electronic and microfluidic devices everywhere inside the vasculature of the brain and other organs.

An illustration of untethered soft robotic microdevices mechanically loading microtissues.

An artistic rendering of the microsurgery platform operating on arrays of microtissues.

Periodic Reporting for period 4 - ROBOCHIP (MicroRobotic toolkit to deliver spatiotemporally resolved physicochemical signals and control cell sociology)

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