Prof. Leeya Engel
Faculty of Mechanical Engineering, Technion-Israel Institute of Technology, Israel
The ability of living cells to respond to mechanical cues from the microenvironment plays a vital role in physiological processes such as embryonic development and heart function. Over the past decade, the field of mechanobiology has seen major advances in identifying mechanosensitive molecules. However, we know comparatively little about the nanometer-scale organization of the cellular components that underly the cell’s ability to generate and sense mechanical force. This knowledge gap reflects a lack of tools that can visualize cellular organization at the nanoscale. In this talk, I will present two cell culture platforms developed to provide programmed mechanical cues to cells imaged with cryo-electron tomography (cryo-ET), a 3D transmission electron microscopy (TEM) modality that offers the highest resolution structural analysis within cells in a near-native state. The first is an extracellular matrix (ECM) micropatterning technology where we use maskless photolithography to functionalize TEM supports to shape cells and direct their positioning at high spatial accuracy, solving an important bottleneck in sample preparation. We demonstrate the utility of this technique for structural studies of lymphocytes, cardiac, and endothelial cells using cryo-ET.
We recently engineered a second cryo-ET platform to study cell sensitivity to ECM topography. We generated nanopatterned cell culture substrates from cryo-TEM by electrospinning ECM fibers directly onto TEM grids. By altering the electrospinning parameters (i.e., the rotating speed of the collector wheel), we programmed the orientation of the nanofibers to mimic healthy (aligned) or disrupted (randomly oriented) ECM organization. We successfully imaged nanoscale features within endothelial cells cultured on this new class of nanopatterned TEM grids, demonstrating that they are compatible with standard cryo-ET workflows (vitrification by plunge-freezing, clipping, etc.). Over the long term, these technologies will elucidate how spatial constraints from the cell microenvironment influence cellular ultrastructure, informing our understanding of the physical cues required for healthy tissue formation.