Assessing the impact of extracellular matrix fiber orientation on breast cancer cellular metabolism.

The extracellular matrix (ECM) is a dynamic and complex microenvironment that modulates cell behavior and cell fate. Changes in ECM composition and architecture have been correlated with development, differentiation, and disease progression in various pathologies, including breast cancer [1]. Studies have shown that aligned fibers drive a pro-metastatic microenvironment, promoting the transformation of mammary epithelial cells into invasive ductal carcinoma via the epithelial-to-mesenchymal transition (EMT) [2]. The impact of ECM orientation on breast cancer metabolism, however, is largely unknown. Here, we employ two non-invasive imaging techniques, fluorescence-lifetime imaging microscopy (FLIM) and intensity-based multiphoton microscopy, to assess the metabolic states of cancer cells cultured on ECM-mimicking nanofibers in a random and aligned orientation. By tracking the changes in the intrinsic fluorescence of nicotinamide adenine dinucleotide and flavin adenine dinucleotide, as well as expression levels of metastatic markers, we reveal how ECM fiber orientation alters cancer metabolism and EMT progression. Our study indicates that aligned cellular microenvironments play a key role in promoting metastatic phenotypes of breast cancer as evidenced by a more glycolytic metabolic signature on nanofiber scaffolds of aligned orientation compared to scaffolds of random orientation. This finding is particularly relevant for subsets of breast cancer marked by high levels of collagen remodeling (e.g. pregnancy associated breast cancer), and may serve as a platform for predicting clinical outcomes within these subsets [3-6].

Engineering Alignment Has Mixed Effects on Human Induced Pluripotent Stem Cell Differentiated Cardiomyocyte Maturation.

The potential of human induced pluripotent stem cell differentiated cardiomyocytes (hiPSC-CMs) is greatly limited by their functional immaturity. Strong relationships exist between cardiomyocyte (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 human induced pluripotent stem cells (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 toward the ventricular compact CM subtype and improves electrochemical coupling in the short term, at day 14 of differentiation. Moreover, we observe the glycolysis to oxidative phosphorylation switch throughout differentiation and CM development. On the subcellular scale, we note changes in mitochondrial morphology in the long term, at day 28 of differentiation. 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 hiPSCs and strongly improve cardiac tissue engineering.

Biomechanical Dependence of SARS-CoV-2 Infections.

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.