Abstract
Purpose: :
To investigate the influence of the microtopography of the immediate extracellular environment to human trabecular meshwork (TM) cell morphology and proliferation, and to understand the impact of microtopography on regulation of hydraulic resistance of TM structures.
Methods: :
Microgrooves and micropillars with different geometries are fabricated on polydimethylsiloxane (PDMS) substrates using softlithography. After sterilizing the substrates, primary cultured TM cells from human donor rims are seeded and allowed to proliferate. Cell morphology is examined using FITC conjugated cell viability kits. Cell alignment is examined by measuring the orientation angle. The degree of cell elongation is also measured. Once a steady state condition of the cell volume and cell alignment is reached (after Day 6), the porosity of the TM cell monolayer is examined by interposing a contrast threshold on the microscopic images.
Results: :
The results showed that microgrooves are efficient in mimicking the in vivo like elongated TM cell morphology, while micropillars direct the cells into more isotropic shapes. In particular, the microgrooves effectively increase the porosity of the monolayer. Using the cells cultured on smooth surfaces as the baseline (100%), the cells cultured on microgrooves typically occupy less than 25%. Moreover, intergroup comparison demonstrates that in single spaced microgrooves, high porosity of the TM cell monolayer is associated with small line widths and small height-width ratios. According to Darcy’s law, the porosity directly relates to the hydraulic resistance of the monolayer. Therefore, it is expected that the microgrooves are able to regulate the hydraulic resistance of in vitro cultured TM cell monolayers. Furthermore, it is found that the cells seeded on the microgrooves with small height-to-width ratios proliferate across the neighboring structures and had a widespread angular distribution. This may affect pore geometries and therefore the Kozeny constant, which changes the permeability.
Conclusions: :
This work shows that engineered microstructures are effective in regulating the porosity of in vitro cultured TM cell monolayers. Since the porosity directly relates to the aqueous humor outflow, one may regulate the hydraulic resistance by designing appropriate microtopographies in the immediate extracellular matrix. This work provides a promising starting point for understanding microtopography-based intraocular pressure regulation, which may play a role in pharmacologic or laser interventions that produce outflow changes.
Keywords: trabecular meshwork • proliferation • topography