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S. H. Boddu, K. G. Janoria, Z. Wang, A. K. Mitra; Functional Characterization of Sodium Dependent Multi- Vitamin Transporter (SMVT) on Human Retinal Pigmented Epithelial Cell Line (ARPE-19). Invest. Ophthalmol. Vis. Sci. 2007;48(13):707.
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© ARVO (1962-2015); The Authors (2016-present)
The objective of this study is to investigate the functional expression of sodium-dependent multiple vitamin transporter (SMVT) on Human Retinal Pigmented Epithelial cell line (ARPE-19) and also to utilize this transport system for delivery of ganciclovir prodrug (biotin-ganciclovir).
Human Retinal Pigmented Epithelial cell line (ARPE-19) was employed as the model. Uptake kinetics of [3H] biotin was determined at various concentrations and time points. Competitive inhibition studies were conducted in the presence of unlabeled biotin, its analog (desthiobiotin), pantothenic acid and lipoic acid. To delineate the mechanism of uptake; metabolic inhibitors (ouabain, 2,4- dinitrophenol (2,4-DNP) and sodium azide); amiloride, were employed. Sodium and chloride ion dependency, pH dependency on the uptake of biotin on ARPE-19 cells were examined. Uptake of biotin was also challenged with biotin ester conjugates such as biotin methyl ester, biocytin, biotin fluorescein and ganciclovir-biotin conjugate.
The uptake of [3H] biotin into ARPE-19 was linear over 7 minutes, and found to be saturable with a Km of 138.25 ÃÂ¼M and a Vmax of 38.84 pmol/min /mg protein. The process was inhibited by unlabeled biotin its analog (desthiobiotin), pantothenic acid and lipoic acid in a concentration dependent manner. Biotin uptake in ARPE-19 was found to be energy and Na+ dependent but H+ and Cl- independent. Uptake process was inhibited significantly in the presence of biotin conjugates (biotin methyl ester and biocytin). Further the uptake was also inhibited in the presence of biotin-ganciclovir conjugate.
Sodium-dependent multi-vitamin transporter responsible for the uptake of biotin was functionally identified and characterized on ARPE-19 cell line. Ganciclovir delivery into the retinal cells can be enhanced utilizing biotin-ganciclovir conjugate.Keywords: sodium-dependent multiple vitamin transporter (SMVT); Human Retinal Pigmented Epithelial cell line (ARPE-19), biotinSupported by: NIH grant R01 EY 09171, R01 EY 10659
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