Kimura Osamu, Fujii Yukiko, Haraguchi Koichi, Kato Yoshihisa, Ohta Chiho, Koga Nobuyuki, Endo Tetsuya
School of Pharmaceutical Sciences, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido, 061-0293, Japan.
Daiichi University of Pharmacy, Tamagawa-cho, Minami-ku, Fukuoka, 815-8511, Japan.
Biochem Biophys Rep. 2020 Sep 7;24:100807. doi: 10.1016/j.bbrep.2020.100807. eCollection 2020 Dec.
We performed a detailed investigation of the uptake of sulfobromophthalein (BSP) from the apical membrane of Caco-2 cells, which is a substrate for organic anion transporting polypeptides (OATPs), and calculated the kinetic parameters of BSP uptake as follows: K = 13.9 ± 1.3 μM, V = 1.15 ± 0.07 nmol (mg protein) (5 min), and k = 38.2 ± 0.53 μL (mg protein) (5 min). Coincubation with medium-chain (C7-C11) perfluoroalkyl carboxylic acids (PFCAs), such as perfluoroheptanoic acid (PFHpA, C7), perfluorooctanoic acid (PFOA, C8), perfluorononanoic acid (PFNA, C9), perfluorodecanoic acid (PFDA, C10) and perfluoroundecanoic acid (PFUnDA, C11), significantly decreased BSP uptake by 27-55%, while coincubation with short- (C3-C6) and long-chain (C12-C14) PFCAs decreased the uptake only slightly. Dixon plotting suggested that PFOA, PFNA and PFDA competitively inhibited the BSP uptake with inhibition constant (K) values of 62.2 ± 1.3 μM, 35.3 ± 0.1 μM and 43.2 ± 0.3 μM, respectively. PFCAs with medium-chains could be substrates for OATPs, probably OATP2B1, which is the most abundantly expressed OATP isoform in Caco-2 cells.
我们对磺溴酞（BSP）从Caco-2细胞顶膜的摄取进行了详细研究，BSP是有机阴离子转运多肽（OATPs）的一种底物，并计算了BSP摄取的动力学参数如下：K = 13.9±1.3 μM，V = 1.15±0.07 nmol/（mg蛋白质）（5分钟），k = 38.2±0.53 μL/（mg蛋白质）（5分钟）。与中链（C7-C11）全氟烷基羧酸（PFCA）如全氟庚酸（PFHpA，C7）、全氟辛酸（PFOA，C8）、全氟壬酸（PFNA，C9）、全氟癸酸（PFDA，C10）和全氟十一酸（PFUnDA，C11）共同孵育，显著降低BSP摄取27%-55%，而与短链（C3-C6）和长链（C12-C14）PFCA共同孵育仅轻微降低摄取。Dixon作图表明，PFOA、PFNA和PFDA竞争性抑制BSP摄取，抑制常数（K）值分别为62.2±1.3 μM、35.3±0.1 μM和43.2±0.3 μM。中链PFCA可能是OATPs的底物，可能是OATP2B1，它是Caco-2细胞中表达最丰富的OATP亚型。
