Functional Analysis of Organic Cation Transporter 3 Expressed in Human Placenta

TLDR

The study aims to compare uptake of organic cations into human placental basal membrane vesicles with that into OCT3‑expressing cells to elucidate the placental transport mechanism. OCT3 function was assessed by measuring sodium‑ and chloride‑independent, saturable uptake of 1‑methyl‑4‑phenylpyridinium (MPP⁺) in HEK293 cells expressing OCT3 and in basal membrane vesicles. OCT3 is the sole OCT isoform in human placenta, localized to basal membrane vesicles, and mediates sodium‑ and chloride‑independent, saturable uptake of MPP⁺ (Km ≈ 80 μM) that is inhibited by cationic drugs but not anionic or zwitterionic compounds, indicating a key role in placental cation transport.

Abstract

The aim of this study is to investigate the placental transport mechanism of cationic compounds by comparison of the uptake of an organic cation into human placental basal membrane vesicles (BLMVs) with that into organic cation transporter 3 (OCT3)-expressing cells. Reverse transcription-polymerase chain reaction analysis demonstrated that OCT3 is the only OCT isoform expressed in the human placenta. The function of OCT3 was investigated by measuring the uptake of 1-methyl-4-phenylpyridinium (MPP⁺) into human embryonic kidney (HEK)293 cells stably expressing OCT3 (HEK/OCT3 cells). The OCT3-mediated uptake of MPP⁺ was sodium- and chloride-independent and saturable, with a Michaelis constant (<i>K</i>_m) of 82.5 μM. The OCT3-mediated uptake was inhibited by various cationic drugs in a concentration-dependent manner but not by anionic compounds, such as <i>p</i>-aminohippuric acid and captopril, or a zwitterion, carnitine. Western blotting analysis of membrane vesicles prepared from human term placenta revealed that OCT3 is expressed only in BLMVs but not in microvillous membrane vesicles. The uptake of MPP⁺ into BLMVs was membrane potential-dependent and saturable, with a <i>K</i>_m value of 51.8 μM, which is similar to that in HEK293/OCT3 cells. The inhibitory spectrum of various compounds on MPP⁺ uptake by BLMVs was also similar to that in HEK293/OCT3 cells. These results suggest that OCT3 is expressed on the basal membrane of human trophoblast cells and plays an important role in the placental transport of cationic compounds.