Sensken Sven-Christian, Bode Constantin, Gräler Markus H
Institute for Immunology, Hannover Medical School, 30625 Hanover, Germany.
J Pharmacol Exp Ther. 2009 Mar;328(3):963-9. doi: 10.1124/jpet.108.148163. Epub 2008 Dec 12.
The immunomodulator fingolimod (FTY720) induces lymphopenia by inhibiting lymphocyte egress from thymus and secondary lymphoid organs (SLOs). It is phosphorylated mainly by sphingosine kinase (SK) 2 in vivo. FTY720-phosphate (FTY-P) activated and rapidly internalized S1P(1), which is the major sphingosine 1-phosphate (S1P) receptor for mediating lymphocyte egress. Although FTY-P is thought to be the active metabolite for triggering the onset of lymphopenia, nonphosphorylated FTY720 was much more potent in inhibiting cellular calcium flux and splenocyte chemotaxis via S1P(1) than FTY-P after preincubation. Determination of both compounds by liquid chromatography coupled to mass spectrometry revealed efficient uptake and accumulation of FTY720 but not FTY-P by splenocytes. Coculture experiments of B and T cells with and without FTY720 pretreatment led to rapid cellular transfer and phosphorylation by mouse lymphocytes. The presence of FTY720 in lymphoid tissues of FTY720-treated SK2-deficient mice without onset of lymphopenia excluded a potential role of the nonphosphorylated compound for lymphocyte egress. Local concentrations of both phosphorylated and nonphosphorylated FTY720 were much higher in lymphoid tissues than in blood. Therefore, we conclude that cellular accumulation of FTY720 generates a reservoir in thymus and SLOs, leading to sustained FTY-P production and activation of S1P(1) within tissues.
免疫调节剂芬戈莫德（FTY720）通过抑制淋巴细胞从胸腺和二级淋巴器官（SLOs）逸出而诱导淋巴细胞减少。它在体内主要由鞘氨醇激酶（SK）2磷酸化。磷酸化芬戈莫德（FTY-P）激活并迅速内化S1P(1)，S1P(1)是介导淋巴细胞逸出的主要1-磷酸鞘氨醇（S1P）受体。尽管FTY-P被认为是引发淋巴细胞减少症发作的活性代谢物，但预孵育后，非磷酸化的FTY720在通过S1P(1)抑制细胞钙通量和脾细胞趋化性方面比FTY-P更有效。通过液相色谱-质谱联用测定这两种化合物，结果显示脾细胞对FTY720有高效摄取和积累，而对FTY-P则没有。对B细胞和T细胞进行有无FTY720预处理的共培养实验，结果显示小鼠淋巴细胞能使细胞快速转移并磷酸化。在未经淋巴细胞减少症发作的经FTY720处理的SK2缺陷小鼠的淋巴组织中存在FTY720，这排除了非磷酸化化合物在淋巴细胞逸出中的潜在作用。磷酸化和非磷酸化的FTY720在淋巴组织中的局部浓度均远高于血液中的浓度。因此，我们得出结论，FTY720在细胞内的积累在胸腺和SLOs中形成了一个储备库，导致组织内持续产生FTY-P并激活S1P(1)。
