Budde Holger, Berntsch Ulrike, Riggert Joachim, Legler Tobias J
University Medical Center Göttingen, Göttingen, Germany.
Cent Eur J Immunol. 2017;42(1):1-9. doi: 10.5114/ceji.2017.67312. Epub 2017 May 8.
Extracorporeal photopheresis (ECP) is an important second-line therapy for graft-versus-host disease. A central therapeutic mechanism is the induction of immune tolerance through apoptosis in patient's leukocytes, caused by ex vivo incubation with 8-methoxypsoralen (8-MOP) and subsequent UVA irradiation. We hypothesized that different 8-MOP incubation times and an additional 8-MOP removal step could influence the apoptosis kinetics of leukocytes in general and in particular could lead to different apoptotic levels in the leukocyte subpopulations. After 8-MOP/UVA treatment of human leukocytes, cells were cultured and the percentage of annexin V positive cells from several leukocyte subpopulations was determined. Only regulatory T cells (Tregs) were relatively resistant to 8-MOP/UVA induced apoptosis. When cells were incubated for 30 minutes with 8-MOP prior to UVA exposure, higher percentages of annexin V positive cells were detected on day 1 and day 2 after treatment. Removal of 8-MOP after UVA exposure caused no significant changes in the apoptosis kinetics during the 72 h culture period compared with unwashed cells. The results of our in vitro study indicate that it could be possible to adjust the apoptosis kinetics via modulation of the 8-MOP incubation time. In further in vivo studies it should be elucidated to which extent different apoptosis kinetics influence the therapeutic effect of ECP since steady-state apoptosis levels are probably important for establishing a long lasting immune tolerance. Furthermore we found that Tregs, according to their well-known tolerogenic function, are more resistant to apoptosis after 8-MOP/UVA treatment compared to GvHD inducing T cell populations.
体外光化学疗法（ECP）是治疗移植物抗宿主病的一种重要二线疗法。其核心治疗机制是通过8-甲氧基补骨脂素（8-MOP）体外孵育及随后的紫外线A（UVA）照射，诱导患者白细胞凋亡，从而产生免疫耐受。我们推测，不同的8-MOP孵育时间以及额外的8-MOP去除步骤，可能会影响白细胞的凋亡动力学，尤其是可能导致白细胞亚群出现不同的凋亡水平。对人白细胞进行8-MOP/UVA处理后，将细胞进行培养，并测定几个白细胞亚群中膜联蛋白V阳性细胞的百分比。只有调节性T细胞（Tregs）对8-MOP/UVA诱导的凋亡相对具有抗性。在UVA照射前用8-MOP将细胞孵育30分钟，在处理后的第1天和第2天检测到更高百分比的膜联蛋白V阳性细胞。与未洗涤的细胞相比，UVA照射后去除8-MOP在72小时培养期内对凋亡动力学没有显著影响。我们的体外研究结果表明，有可能通过调节8-MOP孵育时间来调整凋亡动力学。在进一步的体内研究中，应该阐明不同的凋亡动力学在多大程度上影响ECP的治疗效果，因为稳态凋亡水平可能对建立持久的免疫耐受很重要。此外，我们发现，根据其众所周知的致耐受功能，与诱导移植物抗宿主病的T细胞群体相比，Tregs在8-MOP/UVA处理后对凋亡更具抗性。
