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Jingwen Cai, William M Johnson, Kristin Marie Perkumas, W Daniel Stamer, Yutao Liu; TGFβ Signaling in Human Schlemm’s Canal Endothelial Cells. Invest. Ophthalmol. Vis. Sci. 2018;59(9):5161.
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© ARVO (1962-2015); The Authors (2016-present)
TGFβ-related pathway plays a role in aqueous humor (AH) outflow by regulating extracellular matrix (ECM) turnover in the trabecular meshwork. However, its role in human Schlemm’s canal (HSC) biology remains unclear. HSC endothelial cells are constantly exposed to the mechanical strain generated by the fluctuating pressure gradient created by intraocular pressure versus episcleral venous pressure. Mechanical strain may induce ECM remodeling through TGFβ pathway. We aimed to determine how TGFβ-related target genes with differential expression in glaucoma HSC cells respond to mechanical strain in HSC cells.
We cultured 8 strains of HSC cells from 4 non-glaucoma and 3 glaucoma donors. Using droplet digital PCR (ddPCR) we examined the differential expression of 22 genes related with TGFβ signaling or ECM, including CYP1B1, ABCA1, GAS7, CAV1/CAV2, MGP, DCN, CLDN23, LOX, FMOD, LAMC2, LAMC3, EZR, CRIP2, HOXB2, CDH10, MYO10, DPT, EMILIN2, PODN, TGFBI, TGFBR, and GLDN. Primary HSC cells from 3 non-glaucoma donors were seeded on Bioflex membranes and exposed to 0% or 15% equibiaxial strain using a Flexcell FX-5000 Tension system for 24 hours. Using ddPCR, we examined the expression of the selected genes in response to the mechanical strain in HSC cells. Using the list of mechanical strain-responsive genes, we performed gene network analysis using Ingenuity Pathway Analysis (IPA).
We identified 18 genes with differential expression in glaucomatous HSC cells. In glaucomatous HSC cells, we found increased expression of TGFBR3 and TGFBI and corresponding expression changes of several TGFβ regulators (DPT, DCN) and target genes (LOX and MGP). Among them, 8 genes (CLDN23, DCN, EZR, TGFBR3, CAV2, GAS7, CYP1B1, and ABCA1) were found to have induced expression in the stretched primary HSC cells. IPA analysis illustrated that majority of these mechanical responsive genes were targets by TGFβ/Smad, Nf-kB, and ERK1/2 pathway. Compared to the non-stretched SC, stretched SC cells had higher expression of many TGFβ targeted genes such as CAV1/2, GAS7, CYP1B1, CLDN23 and DCN.
Our study indicates the potential role of TGFβ signaling in AH outflow through regulating SC cells. Further studies will need to validate this pilot finding.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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