Dynamic regulation of single-cell contractile heterogeneity in the human trabecular meshwork

Glaucoma is characterized by increased outflow resistance of aqueous humor and higher intraocular pressure, with altered trabecular meshwork (TM) mechanics contributing to these symptoms. However, the regulation of contractility across individual TM cells is not well understood. Researchers employed a high-throughput single-cell contractility assay to quantify contractile behavior in primary human TM cells under various conditions, such as extracellular matrix (ECM) compositions, substrate stiffnesses, and pharmacological perturbations.

The data showed that TM contractility is not a uniform feature but a context-dependent state influenced by both external and internal factors.

The ECM composition set off distinct contractile programs. Collagen IV promoted a rapid convergence toward a highly contractile state, while collagen I supported a slower but similar response. Fibronectin, on the other hand, maintained a persistently heterogeneous low-contractility state. Intracellular perturbation with dexamethasone (DEX) did not uniformly increase contractility but selectively prolonged activity within the most mechanically active cells, sustaining highly contractile states.

However, this response was reduced as ECM stiffness increased and did not occur on pathologically stiff substrates, highlighting the role of extracellular mechanical context in glucocorticoid responsiveness.

In contrast, inhibition of ROCK (Rho-associated protein kinase) overrode both basal and DEX-induced contractile programs, causing the heterogeneous population to collapse into a uniformly low-contractility state. Intracellular protein abundance, however, poorly predicted functional mechanical behavior, with only weak pooled associations observed. These relationships varied among donors rather than reflecting consistent molecular-mechanical coupling.

In summary, the study reveals that TM contractility arises from the integration of intrinsic cellular states and the extracellular mechanical environment, rather than from either factor alone. Moreover, the research identifies single-cell contractile state as a functional aspect of TM cell identity, which cannot be determined from static molecular marker abundance or population-averaged mechanical measurements.