The potential of human induced pluripotent stem cell differentiated cardiomyocytes (hiPSC-CMs) is greatly limited by their functional immaturity. Strong relationships exist between CM structure and function, leading many in the field to seek ways to mature hiPSC-CMs by culturing on biomimetic substrates, specifically those that promote alignment. However, these in vitro models have so far failed to replicate the alignment that occurs during cardiac differentiation. We show that engineered alignment, incorporated before and during cardiac differentiation, affects hiPSC-CM electrochemical coupling and mitochondrial morphology. We successfully engineer alignment in differentiating hiPSCs as early as Day 4. We uniquely apply optical redox imaging to monitor the metabolic changes occurring during cardiac differentiation. We couple this modality with cardiac-specific markers, which allows us to assess cardiac metabolism in heterogeneous cell populations. The engineered alignment drives hiPSC-CM differentiation towards the ventricular compact CM subtype and improves electrochemical coupling in the short term. Moreover, we observe glycolysis to oxidative phosphorylation switch throughout differentiation and CM development. On the subcellular scale, we note changes in mitochondrial morphology in the long term. Our results demonstrate that cellular alignment accelerates hiPSC-CM maturity and emphasizes the interrelation of structure and function in cardiac development. We anticipate that combining engineered alignment with additional maturation strategies will result in improved development of mature CMs from hiPSC and strongly improve cardiac tissue engineering.
Older people have been disproportionately vulnerable to the current SARS-CoV-2 pandemic, with an increased risk of severe complications and death compared to other age groups. A mix of underlying factors has been speculated to give rise to this differential infection outcome, including changes in lung physiology, weakened immunity, and severe immune response. Our study focuses on the impact of biomechanical changes in lungs that occur as individuals age, i.e., the stiffening of the lung parenchyma and increased matrix fiber density. We used hydrogels with an elastic modulus of 0.2 and 50 kPa and conventional tissue culture surfaces to investigate how infection rate changes with parenchymal tissue stiffness in lung epithelial cells challenged with SARS-CoV-2 Spike (S) protein pseudotyped lentiviruses. Further, we employed electrospun fiber matrices to isolate the effect of matrix density. Given the recent data highlighting the importance of alternative virulent strains, we included both the native strain identified in early 2020 and an early S protein variant (D614G) that was shown to increase the viral infectivity markedly. Our results show that cells on softer and sparser scaffolds, closer resembling younger lungs, exhibit higher infection rates by the WT and D614G variant. This suggests that natural changes in lung biomechanics do not increase the propensity for SARS-CoV-2 infection and that other factors, such as a weaker immune system, may contribute to increased disease burden in the elderly.
The potential of human induced pluripotent stem cell differentiated cardiomyocytes (hiPSC-CMs) is greatly limited by their functional immaturity. Strong relationships exist between CM structure and function, leading many in the field to seek ways to mature hiPSC-CMs by culturing on biomimetic substrates, specifically those that promote alignment. However, these in vitro models have so far failed to replicate the alignment that occurs during cardiac differentiation. We show that engineered alignment, incorporated before and during cardiac differentiation, affects hiPSC-CM electrochemical coupling and mitochondrial morphology. We successfully engineer alignment in differentiating hiPSCs as early as Day 4. We uniquely apply optical redox imaging to monitor the metabolic changes occurring during cardiac differentiation. We couple this modality with cardiac-specific markers, which allows us to assess cardiac metabolism in heterogeneous cell populations. The engineered alignment drives hiPSC-CM differentiation towards the ventricular compact CM subtype and improves electrochemical coupling in the short term. Moreover, we observe glycolysis to oxidative phosphorylation switch throughout differentiation and CM development. On the subcellular scale, we note changes in mitochondrial morphology in the long term. Our results demonstrate that cellular alignment accelerates hiPSC-CM maturity and emphasizes the interrelation of structure and function in cardiac development. We anticipate that combining engineered alignment with additional maturation strategies will result in improved development of mature CMs from hiPSC and strongly improve cardiac tissue engineering.
Older people have been disproportionately vulnerable to the current SARS-CoV-2 pandemic, with an increased risk of severe complications and death compared to other age groups. A mix of underlying factors has been speculated to give rise to this differential infection outcome including changes in lung physiology, weakened immunity, and severe immune response. Our study focuses on the impact of biomechanical changes in lungs that occur as individuals age, that is, the stiffening of the lung parenchyma and increased matrix fiber density. We used hydrogels with an elastic modulus of 0.2 and 50 kPa and conventional tissue culture surfaces to investigate how infection rate changes with parenchymal tissue stiffness in lung epithelial cells challenged with SARS-CoV-2 Spike (S) protein pseudotyped lentiviruses. Further, we employed electrospun fiber matrices to isolate the effect of matrix density. Given the recent data highlighting the importance of alternative virulent strains, we included both the native strain identified in early 2020 and an early S protein variant (D614G) that was shown to increase the viral infectivity markedly. Our results show that cells on softer and sparser scaffolds, closer resembling younger lungs, exhibit higher infection rates by the WT and D614G variant. This suggests that natural changes in lung biomechanics do not increase the propensity for SARS-CoV-2 infection and that other factors, such as a weaker immune system, may contribute to increased disease burden in the elderly.
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