Abstract
Purpose :
The aim of this study is to create a bio-engineered 3D model of the trabecular meshwork (TM) and to evaluate its response to several external stresses. The morphology and composition of the TM and the mechanical properties were analyzed to evaluate similarities with the physiological features of the human TM and moreover, the effect of the stresses on its growth and functionality.
Methods :
The development of the 3D TM polycaprolactone (PCL) material is based on electrospinning techniques combined with fused deposition modeling or Melt electrospinning writing. Different fiber sizes, orientation and deposition density were applied to replicate the micro-anatomy of the human TM. Changes in fiber diameter facilitated the modification of the meshwork cell density and hence, scaffolds with different degrees of stiffness could be obtained. Human TM cells (HTMC) were received from human donors and cultured according to standard protocols. We evaluated the growth, distribution and functionality of HTMC in the 3D model. We used RNA sequencing to assess the HTMC response to increasing stiffness conditions as compared to HTMC cultivated in the classical 2D model.
Results :
HTMC demonstrated a fibroblasts-like morphology with slight differences depending on the donor. These cells were expanded according to the confluence observed in the traditional 2D models. HTMC remained functional and responded to dexamethasone stress in the 3D model. RNA sequencing analysis comparing HTMC growth in the 3D model with the 2D model showed changes in genes associated with gene ontology (GO) pathways. These changes included collagen binding, fibronectin binding, microtubule binding, extracellular matrix (ECM) structural constituent conferring compression resistance, among others.
Conclusions :
A bioengineered 3D HTM model using electrospinning techniques can reproduce human TM features. We achieved an emulation of the TM in varying degrees of stiffness. HTMCs were able to adhere and grow in the 3D structure. The induction of specific TM markers using dexamethasone demonstrated the functionality of the HTMC in our model. The HTMC responded to the changes in the stiffness by altering the expression of genes implicated in the ECM remodelling. These results facilitate the identification of new target genes associated with ECM remodelling, and other pathways.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.