Chun, T.-H. | bioRxiv Science

bioRxiv2024

Tradeoffs in ATP metabolisms via hypoxic gradient migration assays

Migration and scratch assays are helpful tools to investigate wound healing and tissue regeneration processes, especially under disease conditions such as diabetes. However, traditional migration (injury-free) assays and scratch (with injury) assays are limited in their control over cellular environments and provide only simplified read-outs of their results. On the other hand, microfluidic-based cell assays offer a distinct advantage in their integration and scalability for multiple modalities and concentrations in a single device. Additionally, in situ stimulation and detection helps to avoid variabilities between individual bioassays. To realize an enhanced, smarter migration assay, we leveraged our multilayered oxygen gradient (1-16%) to study HaCaT migrations in diabetic conditions with spatial and metabolic read-outs. An analysis of the spatial migration over time observed a new dynamic between hypoxia (at 4.16-9.14% O2) and hyperglycemia. Furthermore, in situ adenosine triphosphate (ATP) and reactive oxygen species (ROS) responses suggest that this dynamic represents a switch between stationary versus motile modes of metabolism. Thus, elevated glucose and hypoxia are synergistic triggers of this switch under disease conditions. These findings illustrate the benefits of spatial microfluidics for modeling complex diseases such as hypoxia and diabetes, where multimodal measurements provide a more deterministic view of the underlying processes.

bioengineering↗

bioRxiv2023

Thrombospondin-1 promotes fibro-adipogenic stromal expansion and contractile dysfunction of the diaphragm in obesity

Pulmonary disorders impact 40-80% of individuals with obesity. Respiratory muscle dysfunction is linked to these conditions; however, its pathophysiology remains largely undefined. Mice subjected to diet-induced obesity (DIO) develop diaphragmatic weakness. Increased intra-diaphragmatic adiposity and extracellular matrix (ECM) content correlate with reductions in contractile force. Thrombospondin-1 (THBS1) is an obesity-associated matricellular protein linked with muscular damage in genetic myopathies. THBS1 induces proliferation of fibro-adipogenic progenitors (FAPs)--mesenchymal cells that differentiate into adipocytes and fibroblasts. We hypothesized that THBS1 drives FAP-mediated diaphragm remodeling and contractile dysfunction in DIO. We tested this by comparing effects of dietary challenge on diaphragms of wild-type (WT) and Thbs1 knockout (Thbs1-/-) mice. Bulk and single-cell transcriptomics demonstrated DIO-induced stromal expansion in WT diaphragms. Diaphragm FAPs displayed upregulation of ECM and TGF{beta}-related expression signatures, and augmentation of a Thy1-expressing sub-population previously linked to type 2 diabetes. Despite similar weight gain, Thbs1-/- mice were protected from these transcriptomic changes, and from obesity-induced increases in diaphragm adiposity and ECM deposition. Unlike WT controls, Thbs1-/-diaphragms maintained normal contractile force and motion after DIO challenge. These findings establish THBS1 as a necessary mediator of diaphragm stromal remodeling and contractile dysfunction in overnutrition, and potential therapeutic target in obesity-associated respiratory dysfunction.

cell biology↗