Title: Micropatterned substrates with physiological stiffness promote cell maturation in human induced pluripotent stem cell-derived skeletal myocytes
Legend: Human induced pluripotent stem cells (iPSCs) present exciting opportunities to study disease processes in vitro. Advances in bioengineering allow us to differentiate cells in a system more relevant to their native environment in order to observe naturally occurring phenomena. We used micropatterned substrates to see how topographical cues and substrate stiffness would affect skeletal myocyte differentiation. We found that differentiating skeletal myocytes from human iPSCs on micropatterned surfaces of physiological stiffness significantly improved the formation of well-aligned and multi-nucleated myotubes. These well-aligned myotubes looked similar to myofibers which spontaneously contracted along the long axis of the micropatterned lane. Next, we used patient iPSC-derived skeletal myocytes with Pompe disease (glycogen storage disease type II) and cultured in this system in order to observe specific cellular pathology. The micropatterned culture platforms enhanced disease phenotype from patient-oriented iPSCs with Pompe disease. Our culture system supports the hypothesis that geometric cues and surface stiffness influence cell morphology and behavior in culture and possibly offers more accurate analysis of disease processes that occur in vivo.
Citation: Jiwlawat N, Lynch E, Napiwocki BN, Stempien A, Ashton RS, Kamp TJ, Crone WC, Suzuki M (2019). Micropatterned substrates with physiological stiffness promote cell maturation and Pompe disease phenotype in human induced pluripotent stem cell‐derived skeletal myocytes. Biotechnology and Bioengineering, in press (Early View available). https://onlinelibrary.wiley.com/doi/10.1002/bit.27075
Abstract: Recent advances in bioengineering have enabled cell culture systems that more closely mimic the native cellular environment. Here, we demonstrated that human induced pluripotent stem cell (iPSC)‐derived myogenic progenitors formed highly‐aligned myotubes and contracted when seeded on two‐dimensional micropatterned platforms. The differentiated cells showed clear nuclear alignment and formed elongated myotubes dependent on the width of the micropatterned lanes. Topographical cues from micropatterning and physiological substrate stiffness improved the formation of well‐aligned and multinucleated myotubes similar to myofibers. These aligned myotubes exhibited spontaneous contractions specifically along the long axis of the pattern. Notably, the micropatterned platforms developed bundle‐like myotubes using patient‐derived iPSCs with a background of Pompe disease (glycogen storage disease type II) and even enhanced the disease phenotype as shown through the specific pathology of abnormal lysosome accumulations. A highly‐aligned formation of matured myotubes holds great potential in further understanding the process of human muscle development, as well as advancing in vitro pharmacological studies for skeletal muscle diseases.
About the Lab: The Suzuki group has demonstrated the therapeutic benefits of ex vivo gene therapy (stem cell-based growth/trophic factor delivery) targeting the skeletal muscle to prevent degeneration of motor neurons and associated neuromuscular junctions during ALS. Although most ALS research has focused on mechanisms of motor neuron cell death, degeneration is also observed in skeletal muscle, particularly at the neuromuscular connection. Glial cell line-derived neurotrophic factor and vascular endothelial growth factor (VEGF) promote survival of motor neurons and their neuromuscular junctions in neuromuscular disorders, such as ALS. Most recently, the lab delivered a combination of GDNF and/or VEGF to muscles using hMSCs; the hMSCs survive and synthesize and release growth factors, which slow disease progression in familial ALS model rats.