Abstract
Purpose :
Glucocorticoids, such as dexamethasone (Dex), are speculated to alter the contractile properties of human trabecular meshwork (HTM) cells. However, there are no definitive studies showing that dexamethasone treatment modulates the contractile forces exerted by HTM cells in vitro. Because cellular contraction is a dynamic process, it is important to quantify changes in contractile force generation over time. Additionally, observed heterogeneity in primary HTM cell cultures call for single cell contraction measurements. Here, we’ve adapted and implemented a contractility assay for HTM cells using a novel device to measure single cell contraction over time after dexamethasone treatment.
Methods :
Primary HTM cells were cultured for 5 days in the presence of DMSO vehicle, Dex (0.1µM), or medium alone. After 5 days, primary HTM cells from each condition were harvested, dissociated to single cells, and seeded into the wells of the fluorescently labelled elastomeric contractible surfaces (FLECS) assay for single cell contraction measurements. Data was collected at 1, 6, &16 hrs post-seeding for each condition and analyzed using computational algorithms.
Results :
Across all three conditions, a single large population of weakly contractile cells and smaller subpopulations of strongly contractile cells were identified. The strongly contractile subpopulations remained smaller compared to the single weakly contractile cell population throughout the course of 16 hrs across all conditions. Interestingly, the strongly contractile cell subpopulations became increasingly prominent in the DMSO vehicle and medium alone conditions. Dex treated cells also displayed small subpopulations of strongly contractile cells. However, these subpopulations were smaller compared to DMSO vehicle and medium conditions, and remained mostly unchanged throughout the course of 16 hrs.
Conclusions :
Here, we show a novel contractility assay that was able to resolve single-cell contractility within a population of human primary HTM cells.Contrary to our hypothesis, Dex treatment appeared to reduce the ability of a population of primary HTM cells to exert strong contractile forces. Future experiments will aim to further characterize the cell populations observed based on their physical properties.
This is a 2020 ARVO Annual Meeting abstract.