Periodic Reporting for period 5 - PolarizeMe (Feeling Polarity: Integrating intracellular mechanics and forces for a biophysical understanding of epithelial polarity)
Reporting period: 2023-09-01 to 2024-08-31
Every living being is made up of a complicated and well organized assembly of cells, that give rise to tissue, organs and finally the macroscopic parts of the body, such as a head, arms and feet. The mechanical differences of these parts are known in everyday life. Bone is harder than even the strongest muscle, and in turn muscles are harder than fat tissue. Obviously, assemblies of cells, and their surrounding can give rise to mechanical differences on the large length-scale, which is obviously highly important for the relevant function. In turn, when opening a cell, we have only a very limited knowledge of the mechanical differences inside cells. To which extend such differences are important for the cellular organization, and might even be relevant for the correct cellular function remains unknown. PolarizeMe aims at illuminating the mechanical properties inside cells by using light, namely optical tweezers and microscopy. As model system we focus on classes of cell that are highly organized, name epithelial, or skin-, cells which show a structural differences between its basis and its top.
Such understanding of cellular mechanics is very important, as in many diseases the intracellular organization is lost, leading to severe problems. One example are kidney cells, for which it is of fundamental importance that the difference between the side facing the blood, and the side facing the urine is well established by the cell. Another important disease is represented by cancer, namely carcinomas which are of epithelial origin. Here epithelial cells loose many of their functions and start dividing and even migrating. The loss of polarity is one of the hallmarks on the transition from a healthy cell to a dangerous metastatic cancer cell. We try to understand if there is an intracellular mechanical structure that is supporting the polarity, and hence helping to maintain it even in a perturbed environment.
These studies are possible by using special properties of light, which are far beyond the everyday experience. When focused to a tiny spot, light starts applying a well defined force on all transparent objects, called a gradient force. This force finds its origin in the electromagnetic fields representing light and we use this to trap particles inside cells. We can thereby hold and manipulate such probe particles, and this is used to measure stiffness, viscosity and also forces inside cells. Another method is high resolution microscopy where we can precisely mark and track structures and small molecules inside the cell. Here we aim at understanding if there are systematic, and potentially circular material flows in the cell that would also lead to order by transporting organelles in a size dependent way. Finally, we want to understand how intracellular organelles and large structures are directionally transported inside the complicated environment of the cell.
Overall, we have the objective to give a detailed plan of the intracellular mechanical properties as well as of the intracellular forces. This will be first done in stable and polarized cells, and then in systematically perturbed cells.
In conclusion, we could significantly advance the field of intracellular mechanics, by defining the changes of mechanical properties during cell division, by providing a new mechanical fingerprint differentiation cells and by demonstrating a new statistical way to describe the active, non-equilibrium properties of cells with a new quantity, called the MBR (Mean Back Relaxation). While the assumption that epithelial cells are highly polarized was not confirmed, this project established a series of new and important findings on the search for the proposed intracellular polarization. This makes the project PolarizeMe a high success, far beyond what as originally thought we could achieve.
Such understanding of cellular mechanics is very important, as in many diseases the intracellular organization is lost, leading to severe problems. One example are kidney cells, for which it is of fundamental importance that the difference between the side facing the blood, and the side facing the urine is well established by the cell. Another important disease is represented by cancer, namely carcinomas which are of epithelial origin. Here epithelial cells loose many of their functions and start dividing and even migrating. The loss of polarity is one of the hallmarks on the transition from a healthy cell to a dangerous metastatic cancer cell. We try to understand if there is an intracellular mechanical structure that is supporting the polarity, and hence helping to maintain it even in a perturbed environment.
These studies are possible by using special properties of light, which are far beyond the everyday experience. When focused to a tiny spot, light starts applying a well defined force on all transparent objects, called a gradient force. This force finds its origin in the electromagnetic fields representing light and we use this to trap particles inside cells. We can thereby hold and manipulate such probe particles, and this is used to measure stiffness, viscosity and also forces inside cells. Another method is high resolution microscopy where we can precisely mark and track structures and small molecules inside the cell. Here we aim at understanding if there are systematic, and potentially circular material flows in the cell that would also lead to order by transporting organelles in a size dependent way. Finally, we want to understand how intracellular organelles and large structures are directionally transported inside the complicated environment of the cell.
Overall, we have the objective to give a detailed plan of the intracellular mechanical properties as well as of the intracellular forces. This will be first done in stable and polarized cells, and then in systematically perturbed cells.
In conclusion, we could significantly advance the field of intracellular mechanics, by defining the changes of mechanical properties during cell division, by providing a new mechanical fingerprint differentiation cells and by demonstrating a new statistical way to describe the active, non-equilibrium properties of cells with a new quantity, called the MBR (Mean Back Relaxation). While the assumption that epithelial cells are highly polarized was not confirmed, this project established a series of new and important findings on the search for the proposed intracellular polarization. This makes the project PolarizeMe a high success, far beyond what as originally thought we could achieve.
In the initial part of the project, we have constructed several new tools, such as automated optical tweezer systems, or a combination of optical tweezers with modern advanced microscopy techniques. Although we have performed many experiments, we cold not confirm any mechanical differences inside polarized cells. However, when investigating many cell types, we found out that during cell division, the intracellular properties undergo a drastic change. Furthermore, we found out that the intracellular mechanical properties, and force generation seems to be governed by a rather simple mathematical description, that suggests a fundamental relation between mechanical properties of the cells and their characteristic force generation. These findings led to a series of 26 publications, many in high impact factor journals, such as Science, Nature Materials, Nature Physics, Nature Communication, Advanced Science and eLife. Furthermore, we have explored new direction to generate polarity, which have resulted in several patents and the creation of a spinout company (ArtifiCell). This work is also supported by an ERC-Proof of Concept grant.
Especially the results that intracellular mechanics is on one side time dependent, and changes significantly during cell division, while on the other side it is possible to determine simple well defined relation for the intracellular mechanics have an important impact on our mechanical understanding of the cellular interior. Despite all the research done up to now, our picture of the mechanical situation inside cells is still blurred. This is due to the difficulty to ‘feel’ inside cells without destroying them. PolarizeMe solves this problem and the current results suggest that we will be able to generate general descriptions of the cellular interior in different situations and conditions of cells.
In summary, we have create a new systematic approach to describe both the active, non-equilibrium component of cell mechanics, but also the viscoelastic properties by providing a model for the viscoelastic shear modulus that is valid over more than 4 orders of magnitude. The surprising finding that the full activity and mechanics of such complicated materials as cells can be described by only 6 parameters remains surprising and opens the door for a complete mechanical description of intracellular mechanics that can be the basis for understanding intracellular mechanical differentiation and intracellular organization. Far beyond our current state of the art goes the developed quantity of mean back relaxation, which has the potential to revolutionize the field of cell mechanics, as it allows a simple and fast characterization of the active intracellular mechanics by a simple observation of spontaneous motion.
Thus the results and achievements obtained in the scope of this ERC project open up a new dimension in our understanding of cell biology and cell organization and provide a full new experimental access and quantitative description of active intracellular mechanics.
In summary, we have create a new systematic approach to describe both the active, non-equilibrium component of cell mechanics, but also the viscoelastic properties by providing a model for the viscoelastic shear modulus that is valid over more than 4 orders of magnitude. The surprising finding that the full activity and mechanics of such complicated materials as cells can be described by only 6 parameters remains surprising and opens the door for a complete mechanical description of intracellular mechanics that can be the basis for understanding intracellular mechanical differentiation and intracellular organization. Far beyond our current state of the art goes the developed quantity of mean back relaxation, which has the potential to revolutionize the field of cell mechanics, as it allows a simple and fast characterization of the active intracellular mechanics by a simple observation of spontaneous motion.
Thus the results and achievements obtained in the scope of this ERC project open up a new dimension in our understanding of cell biology and cell organization and provide a full new experimental access and quantitative description of active intracellular mechanics.