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Colleen M McDowell, Tasneem Putliwala Sharma; TLR4 signaling in the human trabecular meshwork. Invest. Ophthalmol. Vis. Sci. 2019;60(9):5135.
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© ARVO (1962-2015); The Authors (2016-present)
The trabecular meshwork (TM) regulates aqueous humor outflow and intraocular pressure (IOP). Recently, we identified TGFβ2 and toll-like receptor 4 (TLR4) signaling crosstalk as a novel pathway involved in extracellular (ECM) regulation in the TM and ocular hypertension in mice. Here, we investigated the role of TLR4 in a human perfusion organ culture model to determine TGFβ2-TLR4 signaling crosstalk in human ocular hypertension and glaucoma.
Expression of TLR4 was determined by immunohistochemical analysis of TM in normal and glaucomatous human eyes (n=10/condition). To study human ocular hypertension, anterior segments of paired human eyes were perfused at a constant flow (2.5uL/minute) for 4 days to acquire stable baseline IOPs. Elevated IOP was induced by constant perfusion of anterior segments (n=4 pairs) with TGFβ2 (5 ng/mL) with vehicle control in the paired eye for 4 days. In a duplicate set of experiments (n=4 pairs), after stable baseline IOP we treated one eye with TGFβ2 (5 ng/mL) and the paired eye treated with TLR4 inhibitor TAK-242 (15 μM)+TGFβ2 (5 ng/mL) for 8 days. Perfusate and TM tissue from each eye were collected and analyzed for FN expression by western immunoblot analysis.
We observed increased expression of TLR4 in the glaucomatous human TM compared to normal by immunohistochemical staining. TLR4 expressin was ranked none, mild, moderate, or high, in a masked manner. TGFβ2 increased IOP in human perfusion organ cultures above baseline for 4 days post-treatment compared to vehicle control (p<0.05). TLR4 inhibitor TAK-24 blocked TGFβ2-induced ocular hypertension for 8 days post-treatment, while the paired eye treated with TGFβ2 alone induced significant IOP elevation above baseline (p<0.05). TLR4 inhibitor TAK-242 had no significant effect on IOP alone. TGFβ2 increased FN as evident by western blot analysis of TM and perfusate from perfusion experiments, and TLR4 inhibitor blocked TGFβ2- induced FN expression in the TGFβ2+TLR4 inhibitor treatment groups.
These studies identify TGFβ2 – TLR4 crosstalk as a novel pathway involved in ECM regulation in the TM and ocular hypertension. Future studies will determine the effect of TLR4 inhibition on TGFβ2 induced ocular hypertension in this model system. These data provide potential new targets to lower IOP and to further explore the molecular mechanisms involved in the development of glaucomatous TM damage.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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