Wednesday, January 15, 2025
1/15/2025
PROJECT SUMMARY/ABSTRACT Neuronal damage in glaucoma occurs following pathologic mechanical strains and fibrosis of the optic nerve head (ONH). Despite an abundance of evidence implicating mechanical strain and matrix fibrosis in glaucomatous damage, it is not known to what extent and how ONH cells signal to each other to produce matrix fibrosis in response to mechanical strains. This is due, in part, to a lack of adequate model systems to test mechanisms of matrix fibrosis following mechanical strain. In this exploratory R21, we propose to overcome limitations of existing models by establishing an in vitro 3D culture system that (a) accounts for native tissue stiffness, (b) permits dynamic matrix remodeling, (c) includes human ONH astrocytes and lamina cribrosa (LC) cells, and (d) allows for targeted manipulation of cell type-specific molecular pathways. We will then use this system to examine fibrotic signaling pathways between ONH astrocytes and LC cells in response to mechanical strain. Specifically, our preliminary data have identified the extra domain A isoform of fibronectin (FN+EDA) – Toll-Like Receptor 4 (TLR4) signaling axis as critical to matrix deposition by ONH cells. We will use our model system to test the role of the FN+EDA-TLR4 signaling axis in mechanical strain-induced matrix fibrosis. The experiments proposed in this R21 will advance our understanding of ONH mechanobiology by (i) investigating whether crosstalk between ONH astrocytes and LC cells is critical to mechanodysfunction, (ii) establishing quantitative models to test mechanisms of mechanical strain-induced matrix fibrosis, and (iii) investigating the potential of ONH matrix proteins to act as key signaling molecules to enhance ONH fibrosis. Our specific aims are: Aim 1, Generate quantitative models of the fibrotic response of human ONH astrocytes and LC cells after mechanical strain, in monocultures and co-cultures. Aim 2, Test whether astrocyte-derived FN+EDA stimulation of LC cell TLR4 promotes mechanical strain-induced matrix fibrosis.
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