Division of Haematology and Children's Research Center, University Children's Hospital Zurich, Zurich, Switzerland.
Division of Transfusion Medicine, University Hospital Greifswald, Greifswald, Germany.
PLoS One. 2021 Jan 11;16(1):e0244848. doi: 10.1371/journal.pone.0244848. eCollection 2021.
Apoptotic pathways in platelets are important for their survival and function. Platelet apoptosis may be involved in the pathogenesis of immune thrombocytopenia (ITP), an autoimmune-mediated disease. In contrast to the intrinsic apoptosis pathway, not much is known about the extrinsic pathway mechanisms in platelets.
To investigate the expression of proteins involved in the extrinsic apoptosis pathway, including the death receptors, adaptor and regulator proteins in human platelets. To determine a possible trigger of the extrinsic apoptosis pathway in platelets.
To investigate the expression of key markers of the extrinsic pathway we used targeted immunofluorescence and flow cytometry assays. To study their expression and interaction we performed Western blotting and co-immunoprecipitation. Treated platelets with different apoptosis triggers were subjected to flow cytometry.
We could identify the protein expression of the pro-apoptotic proteins TRADD (Tumor Necrosis Factor Receptor type 1- Associated DEATH Domain protein), TRAF2/5, (TNF Associated Factor) and DEDAF (Death Effector Domain- Associated Factor), FADD (Fas-Associated protein with death domain) as well as the anti-apoptotic proteins DJ-1 (Deglycase 1) and c-FLIP in human platelets. ABT-737 treatment induced a disruption in the co-localization of DJ-1 with FADD. Platelets treated with ABT-737 showed an activation in caspase-3 and -8. The exposure to TNF (Tumor Necrosis Factor), FasL (Fas ligand), and TWEAK or to plasma derived from ITP patients, did not lead to changes in caspase-3 and -8 activation in platelets.
Human platelets express some proteins of the extrinsic apoptosis pathway which can be modulated only by ABT-737 treatment. However so far, no other apoptosis trigger or interaction with an external receptor have been yet identified.
血小板中的凋亡途径对于其存活和功能很重要。血小板凋亡可能与免疫性血小板减少症（ITP）的发病机制有关，ITP 是一种自身免疫介导的疾病。与内在凋亡途径相比，人们对血小板外在凋亡途径的机制知之甚少。
研究参与外在凋亡途径的蛋白质表达，包括人血小板中的死亡受体、衔接和调节蛋白。确定血小板外在凋亡途径的可能触发因素。
为了研究外在途径的关键标志物的表达，我们使用了靶向免疫荧光和流式细胞术检测。为了研究它们的表达和相互作用，我们进行了 Western blot 和共免疫沉淀。用不同凋亡触发剂处理血小板，然后进行流式细胞术检测。
我们能够鉴定出促凋亡蛋白 TRADD（肿瘤坏死因子受体 1 相关死亡结构域蛋白）、TRAF2/5（TNF 相关因子）和 DEDAF（死亡效应结构域相关因子）、FADD（带有死亡结构域的 Fas 相关蛋白）以及抗凋亡蛋白 DJ-1（去糖基酶 1）和 c-FLIP 在人血小板中的蛋白表达。ABT-737 处理诱导 DJ-1 与 FADD 的共定位中断。用 ABT-737 处理的血小板显示 caspase-3 和 -8 的激活。暴露于 TNF（肿瘤坏死因子）、FasL（Fas 配体）、TWEAK 或 ITP 患者的血浆不会导致血小板 caspase-3 和 -8 激活发生变化。
人血小板表达外在凋亡途径的一些蛋白质，这些蛋白质只能通过 ABT-737 处理来调节。然而，到目前为止，尚未发现其他凋亡触发因素或与外部受体的相互作用。
