Figure Legends
Fig. 1. Molecular mechanisms of mechanotransduction in stem cells. Mechanotransduction describes the ability of cells to sense mechanical forces and translate them into biochemical reactions for signal transduction and biological responses. When external forces, such as fluid shear stress, are applied to cells, these forces are transferred from the ECM to the cell, propagating from the outside into the cell. Integrins are a family of transmembrane receptors connecting the cytoskeleton to the ECM. They are also known to initiate cytosolic signaling upon binding to extracellular ligands (outside-in signaling) as well have their binding affinity regulated intracellularly (inside-out signaling). Several mechanosensor proteins, including mechanosensing ion channels, cell-substrate and cell-cell junctional complexes, and cytoskeleton-associated complexes, have been identified in stem cells. Arrows and bars represent the net result of a signaling pathway. Abbreviations used: ECM, extracellular matrix; RhoA, Rho kinase; ROCK, Rho-associated protein kinase; LINC, linker of nucleoskeleton and cytoskeleton.
Fig. 2. Understanding the internal and external variability-generated intrinsic disorder in stem cell bioprocess based on mechanobiological considerations. The cell responses to internal and external forces are represented by the time-related phenomena in biological processes; time-dependency, time-delay, tardive, and uncertainty. These may lead to cellular bioprocess variability, the so-called intrinsic disorder that affects bioprocess efficiency.
Fig. 3. A simplified view of mechanosensing and mechanotransduction signaling pathways for regulating stem cell homeostasis. Cellular behavior is affected by mechanical forces through the process of mechanotransduction, in which mechanical stimuli from their extracellular environment or cell-generated forces are converted into a set of biochemical reactions and a cellular response. We categorized the “intrinsic cellular processes” as “trigger,” “effector,” and “memorizer” key factors involved in mechanotransduction. The actin-myosin cytoskeleton is defined as the “integrator” and “organizer” of mechanical and biochemical signaling inputs in these processes. Arrows and bars represent the net result of a signaling pathway. Arrows and bars represent the net result of a signaling pathway. Abbreviations used: RhoA, Rho kinase; MLC-P, phosphorylated myosin II regulatory light chain; MLC-P, myosin light chain phosphatase; MLCK, myosin light chain kinase.
Fig. 4. Development of criteria and indicators applicable to the study of stem cell bioprocess effectiveness based on mechanobiological mechanisms. We propose new indicators to assess the cell status associated with cell behavior-driven epigenetic memory formation in human induced pluripotent stem cell (hiPSC) culture, as a case study. The acquired memories are transformed from a short-term memory into a long-term memory in transition between exponential and long-term stationary phases. The relationship between the indicators in the same category was used to determine the cell state and differentiation potential. The combination and integration of qualitative and quantitative methods can be used to enhance stem cell bioprocess optimization without compromising the product quality.
Fig. 5. Potential strategies and practical considerations to stem cell bioprocess optimization with the conceptual framework based on mechanobiological mechanisms. We propose a new conceptual framework that systematically links three dimensions: inputs, outputs, and the associated system level outcomes of bioprocess effectiveness. Based on the dynamic relationship between the key factors (trigger, effector, and memorizer) affecting the formation of epigenetic memory (sensory, short-term, and long-term), the determination of the boundary conditions of applied mechanical forces is important to enhance cell quality and process robustness. The application of the conceptual frameworks can apply an important approach that facilitate movement to understand, predict, and control the intrinsic disorder in stem cell bioprocessing. Finally, this allows that the output of the design process, including a quality control plan, a bioprocessing plane, and a strategy for facilities, validation, and other requirements.