Functional expression of the ATP-gated P2X7 receptor in human iPSC-derived astrocytes.

IF 3 4区 医学 Q2 NEUROSCIENCES
Purinergic Signalling Pub Date : 2024-06-01 Epub Date: 2023-07-15 DOI:10.1007/s11302-023-09957-8
Jaideep Kesavan, Orla Watters, Laura de Diego-Garcia, Aida Menéndez Méndez, Mariana Alves, Klaus Dinkel, Michael Hamacher, Jochen H M Prehn, David C Henshall, Tobias Engel
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引用次数: 0

Abstract

Activation of the ATP-gated P2X7 receptor (P2X7R), implicated in numerous diseases of the brain, can trigger diverse responses such as the release of pro-inflammatory cytokines, modulation of neurotransmission, cell proliferation or cell death. However, despite the known species-specific differences in its pharmacological properties, to date, most functional studies on P2X7R responses have been analyzed in cells from rodents or immortalised cell lines. To assess the endogenous and functional expression of P2X7Rs in human astrocytes, we differentiated human-induced pluripotent stem cells (hiPSCs) into GFAP and S100 β-expressing astrocytes. Immunostaining revealed prominent punctate P2X7R staining. P2X7R protein expression was also confirmed by Western blot. Importantly, stimulation with the potent non-selective P2X7R agonist 2',3'-O-(benzoyl-4-benzoyl)-adenosine 5'- triphosphate (BzATP) or endogenous agonist ATP induced robust calcium rises in hiPSC-derived astrocytes which were blocked by the selective P2X7R antagonists AFC-5128 or JNJ-47965567. Our findings provide evidence for the functional expression of P2X7Rs in hiPSC-derived astrocytes and support their in vitro utility in investigating the role of the P2X7R and drug screening in disorders of the central nervous system (CNS).

Abstract Image

ATP 门控 P2X7 受体在人 iPSC 衍生星形胶质细胞中的功能表达。
ATP 门控 P2X7 受体(P2X7R)与多种脑部疾病有关,激活 P2X7R 可引发多种反应,如释放促炎细胞因子、调节神经传递、细胞增殖或细胞死亡。然而,尽管其药理特性存在已知的物种特异性差异,但迄今为止,有关 P2X7R 反应的大多数功能研究都是在啮齿动物细胞或永生细胞系中进行分析的。为了评估 P2X7R 在人类星形胶质细胞中的内源性和功能性表达,我们将人类诱导多能干细胞(hiPSCs)分化为表达 GFAP 和 S100 β 的星形胶质细胞。免疫染色显示了明显的点状P2X7R染色。P2X7R 蛋白表达也得到了 Western 印迹的证实。重要的是,强效非选择性 P2X7R 激动剂 2',3'-O-(苯甲酰基-4-苯甲酰基)-腺苷 5'- 三磷酸(BzATP)或内源性激动剂 ATP 可诱导 hiPSC 衍生星形胶质细胞中的钙离子强劲上升,而选择性 P2X7R 拮抗剂 AFC-5128 或 JNJ-47965567 可阻断这种上升。我们的研究结果为 P2X7R 在 hiPSC 衍生的星形胶质细胞中的功能表达提供了证据,并支持其在体外研究 P2X7R 的作用和中枢神经系统(CNS)疾病的药物筛选中的实用性。
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来源期刊
Purinergic Signalling
Purinergic Signalling 医学-神经科学
CiteScore
6.60
自引率
17.10%
发文量
75
审稿时长
6-12 weeks
期刊介绍: Nucleotides and nucleosides are primitive biological molecules that were utilized early in evolution both as intracellular energy sources and as extracellular signalling molecules. ATP was first identified as a neurotransmitter and later as a co-transmitter with all the established neurotransmitters in both peripheral and central nervous systems. Four subtypes of P1 (adenosine) receptors, 7 subtypes of P2X ion channel receptors and 8 subtypes of P2Y G protein-coupled receptors have currently been identified. Since P2 receptors were first cloned in the early 1990’s, there is clear evidence for the widespread distribution of both P1 and P2 receptor subtypes in neuronal and non-neuronal cells, including glial, immune, bone, muscle, endothelial, epithelial and endocrine cells.
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