OAT1/3 regulate the absorption of CP-25 in the rat synovium

Paeoniflorin-6’-O-benzene sulfonate (CP-25) is a new ester derivative formed by esterification of paeoniflorin. We found that CP-25 inhibits the abnormal proliferation and migration of inflammatory fibroblast-like synoviocytes (FLS). However, the mechanism by which CP-25 is absorbed by FLS remains unclear. Therefore, we established a UPLC–MS/MS methodology to study the mechanism of CP-25 uptake by FLS. Our research revealed that the uptake of CP-25 by rat FLS was time- and concentration-dependent and that the concentration of CP-25 reached a dynamic equilibrium after approximately 60 min. A glucose-free environment, a low-temperature environment or the inhibition of ATP synthesis significantly reduced the absorption of CP-25 by FLS. Furthermore, we confirmed through siRNA interference and overexpression through transient transfection that organic anion transporter (OAT)1/3 play a major role in the process of active uptake of CP-25 by FLS. Additionally, we validated the absorption of CP-25 mediated by OAT1/3 in the synovia with OAT1/3 substrates and inhibitors. Moreover, the absorption of CP-25 by FLS from adjuvant arthritis rats and rheumatoid arthritis patients was significantly lower than that by FLS from normal rats and normal individuals. Taken together, our data suggested that CP-25 is absorbed by FLS through an active transport process, which is mediated primarily by OAT1/3. This study provides an experimental and theoretical basis for further exploration of the molecular mechanism involved.