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Jeffrey P. Mayer, Algis Kasniunas, Ryan D. McCarty, Stephen N. Schwartz, Beatrice Y.J.T. Yue, Paul A. Knepper; Toll-like Receptor 4, Low Molecular Weight Hyaluronic Acid and Cytokine Response in Trabecular Meshwork Cells. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4652.
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© ARVO (1962-2015); The Authors (2016-present)
Innate immunity is the first line of defense and is rapidly activated by cell trauma and infection leading to NF-Κβ activation responsive genes. Recent evidence suggests that low molecular weight hyaluronic acid (HA) signals through CD44-TLR-4-MD-2 receptor complex. The purpose of this study is to determine if low molecular weight HA elicits a cytokine response through Toll-like receptor 4 in trabecular meshwork (TM) cells.
TM primary cells were plated at 40,000 cells/well in 0.5 mL 10% FBS complete media for cell viability studies. Cells were treated for 24 hours with 40 µg/ml of low and high molecular weight HA, PBS control, and 25 µg LPS. Cells were harvested and counted using trypan blue. For cytokine studies, cells were treated with low and high molecular weight HA, and PBS control, and cell lysates were obtained after 6, 24, and 48 hours. The concentrations of the cytokines TNF-α, IFNr, GCSF, GMCSF, IL-1α, IL-8, IP-10 and Rantes were measured using a Human Inflammation ELISA strip (Signosis).
The data was corrected relative to the PBS control. The results indicated cell viabilities of 61.7% (+5.16%) for LPS, 80.2% (+3.61%) for 40 µg low molecular weight HA (P<0.02 compared to control), and 87.7% (+3.51%) for high molecular weight HA. Cytokine analysis profile of high molecular weight HA treated cells indicated decreased Rantes, GMCSF, IFNr, TNF-α and GCSF cytokine concentrations by 48 hours. Low molecular weight HA increased concentration of GCSF and IP-10 by 48 hours, and decreased the concentration of Rantes, GMCSF, a potent CNS neuroprotective cytokine, and TNF-α. Neither treatment of HA had an effect on IL-8 concentrations.
Our results indicate that administration of low molecular weight HA to TM cells decreases cell viability and induces cytokine responses in a time-dependent manner which may have pro-inflammatory as well as neuroprotective effects on TM cells.
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