Overexpression and Activation of αvβ3 Integrin Differentially Affects TGFβ2 Signaling in Human Trabecular Meshwork Cells

Abstract

Studies from our laboratory have suggested that activation of αvβ3 integrin-mediated signaling could contribute to the fibrotic-like changes observed in primary open angle glaucoma (POAG) and glucocorticoid-induced glaucoma. To determine how αvβ3 integrin signaling could be involved in this process, RNA-Seq analysis was used to analyze the transcriptomes of immortalized trabecular meshwork (TM) cell lines overexpressing either a control vector or a wild type (WT) or a constitutively active (CA) αvβ3 integrin. Compared to control cells, hierarchical clustering, PANTHER pathway and protein-protein interaction (PPI) analysis of cells overexpressing WT-αvβ3 integrin or CA-αvβ3 integrin resulted in a significant differential expression of genes encoding for transcription factors, adhesion and cytoskeleton proteins, extracellular matrix (ECM) proteins, cytokines and GTPases. Cells overexpressing a CA-αvβ3 integrin also demonstrated an enrichment for genes encoding proteins found in TGFβ2, Wnt and cadherin signaling pathways all of which have been implicated in POAG pathogenesis. These changes were not observed in cells overexpressing WT-αvβ3 integrin. Our results suggest that activation of αvβ3 integrin signaling in TM cells could have significant impacts on TM function and POAG pathogenesis.

A20 Attenuates the Fibrotic Response in the Trabecular Meshwork

“…Since NF-κB is necessary for the fibrotic response, we investigated how a suppressor of NF-κB, A20, may be able to rescue the fibrotic response within the TM. Importantly, the A20 gene has recently been shown to be downregulated in human TM cells that were made to have constitutively active αvβ3 integrin [22]. The activation of this integrin is suggested to contribute to the fibrotic-like changes observed in primary open-angle glaucoma, therefore A20's diminished presence may be contributing to the increased fibrotic response seen in glaucoma.…”

“…Future work will be needed to determine if A20 is altering the amount of NF-κB activation or its total levels in the TM. However, it is known that A20 gene expression can be downregulated in TM cells, and changes in the expression of A20 have been associated with glaucoma [12,22]. Specifically, immortalized trabecular meshwork cell lines with constitutively active αvβ3 integrin, activation of which could contribute to fibrosis in POAG, had downregulated expression of A20 compared to wild-type control cells [22].…”

“…However, it is known that A20 gene expression can be downregulated in TM cells, and changes in the expression of A20 have been associated with glaucoma [12,22]. Specifically, immortalized trabecular meshwork cell lines with constitutively active αvβ3 integrin, activation of which could contribute to fibrosis in POAG, had downregulated expression of A20 compared to wild-type control cells [22]. In glaucomatous human retinas, A20 expression has also been found to be downregulated [12].…”

Pathogenesis of glaucoma: Extracellular matrix dysfunction in the trabecular meshwork‐A review

“…177 This interaction produces CLANs observed in glaucomatous tissues and glucocorticoid-treated cells. 46,178,179 Activation of αvβ3 integrin also induces expression of TGFβ2, 180 inhibits phagocytosis, 51 and increases the deposition of fibronectin matrices. 180,181 Finally, deletion of this integrin in the mouse eye causes a decrease in IOP, while activating αvβ3 increases IOP, further suggesting a role of αvβ3 integrin in controlling IOP.…”

Gene Expression Data for Investigating Glaucoma Treatment Options and Pharmacology in the Anterior Segment, State-of-the-Art and Future Directions

“…The trabecular meshwork (TM) was the subject of a remarkable number of studies, by microarray and more recently by RNA-seq. In GSE180407 ( Filla et al, 2021 ), the effects of overexpressing either a wild type or a constitutively active avb3-integrin on the transcriptome of an immortalized line of human TM cells are described. GSE159793/GSE122652 ( Nettesheim et al, 2019 ; Shim et al, 2021 ) investigate human TM cells with and without silenced expression of the autophagy genes Atg5 and Atg7, some of which were subjected to cyclic mechanical stretch.…”