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Aswani Dutt Vadlapudi, Ramya Krishna Vadlapatla, Dhananjay Pal, Ashim K Mitra; Functional Aspects And Molecular Evidence Of Organic Anion Transporting Polypeptide 2 (oatp2) In Human Corneal Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2012;53(14):4198.
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The objective of this study was to delineate the functional and molecular aspects of organic anion transporting polypeptide 2 (OATP2) in human corneal epithelial cells (HCEC) and to examine active involvement of this transporter in the uptake of various ophthalmic drugs.
HCEC were cultured using Dulbecco's Modified Eagle Medium: Nutrient Mixture F-12 (DMEM/F-12) medium. [3H] Estrone sulfate, a well known OATP2 substrate was employed to delineate the saturation kinetics of the transporter. The amount of radioactivity was quantified with a scintillation counter. Effects of temperature, extracellular pH, ions, metabolic inhibitors and several ocular drugs on the uptake of [3H] estrone sulfate were determined. Reverse transcription-polymerase chain reaction (RT-PCR) was performed to check the molecular evidence of OATP2. Further molecular evidence was also confirmed by immunoblot analysis.
Uptake of [3H] estrone sulfate was found to be saturable with a Km of 256.81 ± 26.32 µM and Vmax of 51.28 ± 2.56 pmol/mg protein/min. The uptake process was found to be temperature, pH, ion and energy dependent. Simvastatin, olmesartan and ritonavir significantly inhibited the uptake of [3H] estrone sulfate with IC50 values of 639.60 ± 1.24 nM, 135.90 ± 1.72 µM and 144 ± 1.12 nM, respectively. Molecular evidence was confirmed by both RT-PCR and immunoblot analysis.
This study demonstrates for the first time the functional aspects and molecular existence of OATP2 in human corneal epithelial cells. These findings suggest that OATP2 may be involved in the disposition of various ocular drugs.
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