Huang F-D, Kung F-L, Tseng Y-C, Chen M-R, Chan H-S, Lin C-J
School of Pharmacy, College of Medicine, National Taiwan University, 12F, 1 Jen-Ai Road, Section 1, Taipei 100, Taiwan.
Placenta. 2009 Feb;30(2):187-94. doi: 10.1016/j.placenta.2008.11.016. Epub 2008 Dec 16.
Placental OCTN2 is a high-affinity carnitine transporter that can interact with a number of therapeutic agents. The process of syncytialization is associated with the expression of a variety of genes. However, the association between syncytialization and OCTN2 expression is not yet clear. Given that forskolin induces BeWo cells to undergo biochemical and morphological differentiation, the purpose of the present study was to investigate whether the function and expression of OCTN2 are influenced by forskolin treatment during syncytialization. The forskolin-induced differentiation of BeWo cells was validated by secretion of beta-human chorionic gonadotropin (beta-hCG) and syncytin expression. Cellular localization of OCTN2 was analyzed by confocal microscopy. Expression of OCTN2 and the modular proteins PDZK1, PDZK2, NHERF1 and NHERF2 was analyzed by Western blotting and carnitine uptake by BeWo cells was estimated and the kinetic properties of uptake measured. The results showed that forskolin treatment increased beta-hCG secretion and syncytin expression, suggesting induction of syncytialization. Confocal images of BeWo cells showed the localization of OCTN2 in the brush-border membrane. OCTN2 protein expression was upregulated in isolated brush-border membranes by long-term forskolin treatment, but the V(m) for carnitine uptake was unchanged, although the K(m) increased. PDZK1, NHERF1 and NHERF2 protein expression in the brush-border membrane was downregulated by forskolin treatment, whereas PDZK2 levels remained unchanged. In conclusion, protein expression and function of OCTN2 in BeWo cells can be regulated by forskolin treatment. While the presence of forskolin results in an increase in OCTN2 protein expression, the increase in uptake capacity may be compensated by the decreased expression of PDZK1, NHERF1 or NHERF2.
胎盘有机阳离子转运体2（OCTN2）是一种高亲和力的肉碱转运体，可与多种治疗药物相互作用。合体化过程与多种基因的表达有关。然而，合体化与OCTN2表达之间的关联尚不清楚。鉴于福司可林可诱导BeWo细胞发生生化和形态学分化，本研究的目的是探讨在合体化过程中福司可林处理是否会影响OCTN2的功能和表达。通过β-人绒毛膜促性腺激素（β-hCG）分泌和合体素表达验证福司可林诱导的BeWo细胞分化。通过共聚焦显微镜分析OCTN2的细胞定位。通过蛋白质印迹法分析OCTN2以及模块化蛋白PDZK1、PDZK2、NHERF1和NHERF2的表达，并估计BeWo细胞的肉碱摄取量并测量摄取的动力学特性。结果表明，福司可林处理增加了β-hCG分泌和合体素表达，提示诱导了合体化。BeWo细胞的共聚焦图像显示OCTN2定位于刷状缘膜。长期福司可林处理使分离的刷状缘膜中OCTN2蛋白表达上调，但肉碱摄取的V(m)不变，尽管K(m)增加。福司可林处理使刷状缘膜中PDZK1、NHERF1和NHERF2蛋白表达下调，而PDZK2水平保持不变。总之，福司可林处理可调节BeWo细胞中OCTN2的蛋白表达和功能。虽然福司可林的存在导致OCTN2蛋白表达增加，但摄取能力的增加可能被PDZK1、NHERF1或NHERF2表达的降低所抵消。
