Characterization of organic anion transporters in cultured MG-63 Human osteoblast cells

초록

The orgnaic anion excretion from the body, including numerous endogenous and exgenous compounds is mainly performed by plasmamembrane integral transport protein in the kidney and liver. these organic anions (such as a number of clinically used drugs, pesticides and herbicides) are chemically heterogeneous substances that posses a carbon backbone and a net gehative chage at physiological pH. Several integral bembrane protein families serve as organic anion transporters. There ate ATP binding cassette, organic anion transporting protein and drganic anion transporter (OAT). Among these OATS (including OAT1, 2 and 3) have extremely wide spectrum of substrate specificity. They mainly expressed in the brain, liver and kdney. The purpose of this study is to clarify the expression and characterization of OATs in cultured MG-63 human osteoblast cells. The expression of OAT1, 2 and 3 was determined using reverse transcription-polymerase chain reaction (PCR) and Western blotting analysis. Among these three isoforms of OATs, OAT3 messenger RNA was detected in cultured MG-63 osteoblast cells. No RT-PCR products of OAT1 and 2 were observed. By Western blot analysis, OAT3 protein was detected in plasma membrane fraction. The isotope labeled para-aminohippuric acid, estrone sulfate and dehydroepiandrosterone sulfate uptake at 37℃ were signifcantly increased comparing those of contron (at 0-4℃ ). The Km and Vmax values in MG-63 cells on estrone sulfate is 2.7 ±0.7 μM and 148.7 ± 19.7 pmol/.mg protein/min, respectively. The uptake was inhibited by some organic anion chemical/drug and several sulfate conjugates. From these results, obrained data would contribute to understanding of therapeutics and patho-physiology of bone disease. ℃