Flow-dependent traction in high- and low-flow normal trabecular meshwork cells.

Abstract

In the conventional aqueous outflow pathway, high-flow (HF) regions are compliant and low-flow (LF) regions are stiff and relatively shear-insensitive. We investigated whether HF- and LF-derived human trabecular meshwork/juxtacanalicular tissue (TM/JCT) cells differ in how shear flow due to aqueous humor outflow is transduced into cell-matrix biomechanics. Normal human HF and LF TM/JCT cells from one donor were cultured on type-I-collagen-coated polyacrylamide gels (∼4.7 kPa) embedded with FluoSpheres for 3D traction force microscopy. A recirculating system (computational fluid dynamics-calibrated) provided two conditions, No-Flow → Flow and Flow → No-Flow (6 h each). From time-lapse confocal Z-stacks we computed traction force and the derived divergence, curl, and strain fields. When we applied shear flow, HF traction increased over time (+343 Pa/h) and curl trended upward (+0.5 units/h), whereas corresponding LF traction and curl increased by +235 Pa/h and +0.3 units/h, respectively. By 12 h (last 6 h under flow), HF showed higher traction and curl than LF (medians 6.14 kPa compared to 1.68 kPa and 8.3 compared to 1.8, respectively); HF divergence remained negative (contractile) while LF divergence stayed near zero/positive. We identified a shear-associated traction-curl coupling in HF TM/JCT cells and a comparatively shear-insensitive state in LF cells. HF cells act as reversible shear transducers that tune contractility and rotational force with shear flow, whereas LF cells maintain sustained tension with blunted shear coupling. This regional data reveals a glaucoma-relevant mechanism as loss of shear sensing in LF regions sustains contractile forces and matrix compaction that may contribute to increasing outflow resistance. STATEMENT OF SIGNIFICANCE: Glaucoma increases eye pressure when the fluid-draining tissue becomes stiff. We directly quantify how physiological shear alters cell-matrix forces in human trabecular meshwork cells from naturally high- and low-flow regions. On collagen-coated hydrogels, 3D traction force microscopy tracked how cells pull, twist, and compact the matrix. High-flow cells sensed shear and ramped contractile and rotational forces (a "contractile-curl" signature); low-flow cells stayed relatively unresponsive and maintained tension. This region-specific loss of shear sensing explains how outflow resistance can rise as persistent tension compacts matrix. The assay results in a simple, time-resolved biomarker of shear responsiveness that can be used to screen drugs or devices aimed at restoring shear-coupled mechanics and lowering intraocular pressure.