In vitro effects of different 8-methoxypsoralen treatment protocols for extracorporeal photopheresis on mononuclear cells.
In vitro effects of different 8-methoxypsoralen treatment protocols for extracorporeal photopheresis on mononuclear cells.
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DOI：
10.5114/ceji.2017.67312
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发表时间：
2017
期刊：
影响因子：
--
通讯作者：
Legler TJ
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文献类型：
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作者：
Budde H;Berntsch U;Riggert J;Legler TJ
体外光分离术(ECP)是治疗移植物抗宿主病的重要二线疗法。一个主要的治疗机制是通过患者白细胞的凋亡诱导免疫耐受，这是由8-甲氧补骨脂素(8-MOP)体外孵育和随后的UVA照射引起的。我们假设，不同的8-MOP孵育时间和额外的8-MOP去除步骤可以影响一般白细胞的凋亡动力学，特别是可能导致不同的白细胞亚群的凋亡水平。用8-MOP/UVA处理人白细胞后，进行细胞培养，并测定不同白细胞亚群的Annexin V阳性细胞百分率。只有调节性T细胞(Tregs)对8-MOP/UVA诱导的细胞凋亡具有相对抗性。在UVA照射前用8-MOP孵育细胞30min，在处理后第1天和第2天，Annexin V阳性细胞百分率较高。UVA照射后去除8-MOP后，细胞在72 h培养期间的凋亡动力学与未清洗细胞相比无明显变化。我们的体外研究结果表明，通过调节8-MOP孵育时间来调节细胞凋亡动力学是可能的。在进一步的体内研究中，应该阐明不同的细胞凋亡动力学对ECP治疗效果的影响程度，因为稳定的细胞凋亡水平对于建立持久的免疫耐受可能是重要的。此外，我们还发现，与GvHD诱导的T细胞群体相比，Tregs在8-MOP/UVA治疗后具有更强的抗凋亡能力。
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.