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Jeffrey Dunmire, Rachida Bouhenni, Yang Yun, Deepak P. Edward; Internalization of L-Tyrosine Polyphosphate Nanoparticles by Human Trabecular Meshwork Cells. Invest. Ophthalmol. Vis. Sci. 2012;53(14):505.
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L-tyrosine polyphosphate (LTP) based nanoparticles can effectively bind to various chemical and biologic molecules, and have been considered as a means for targeted drug delivery. We investigated the in vitro biocompatibility and internalization of LTP nanoparticles in human trabecular meshwork (HTM) cells.
Primary HTM cells were seeded on 6-well culture plates 24 h prior to the addition of nanoparticles. Culture medium was replaced with rhodamine encapsulated LTP nanoparticles suspended in complete medium at 100, 10, 1, or 0.5 µg/ml. Cells were incubated at 37°C/5% CO2 in a humidified chamber for 48 h, 72 h, or 7 d. Negative controls included cells receiving unloaded LTP nanoparticles and cells receiving no nanoparticles. Following incubation, cells were rinsed, fixed, permeabilized, stained with both phalloidin-Alexa Fluor 488 and DAPI, and visualized by fluorescence microscopy.
Internalization of LTP nanoparticles by HTM cells was demonstrated, to varying degrees, at all time points for all concentrations of nanoparticle tested. Nanoparticle uptake efficiency increased to greater than 75% by day 7 using 10 µg/ml. Intracellular nanoparticles localized to the perinuclear region with few remaining free in the cytoplasm. Cell density and morphology were unchanged at all time points with 0.5, 1, and 10 µg/ml of LTP nanoparticle. However, 100 µg/ml of LTP nanoparticle had the apparent consequence of stress-induced morphological changes as evidenced by a more than 50% decrease in cell density and extension of numerous cytoplasmic processes.
This in vitro data suggests that HTM cells can internalize loaded LTP nanoparticles without affecting morphology or cell survival, except at high concentrations. LTP nanoparticles could potentially be used for delivery of chemical modulators of intra-ocular pressure, targeting cell systems that affect aqueous humor outflow.
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