Trubiani Oriana, Horenstein Alberto L, Caciagli Francesco, Caputi Sergio, Malavasi Fabio, Ballerini Patrizia
J Cell Biochem. 2014 Jun;115(6):1138-46. doi: 10.1002/jcb.24756.
ATP is released by human periodontal ligament cells (hPDLCs) and has been shown to regulate PDL regeneration and responses to mechanical stress through activation of P2Y receptors. This nucleotide, however, has also been reported to trigger the pro-inflammatory cascade by inducing the maturation and/or release of chemokines/cytokines from various cell types mainly via P2X7 receptors. Much less is known on the possible role of ATP in stem cells deriving from PDL (hPDLSCs) which are considered to be a promising tool for cell-based therapy to restore lesions. Given the role played by P2X7 in pathophysiological conditions, in this study we investigated the expression of P2X7 ATP receptors in hPDLSCs. The results obtained showed that hPDLSCs express P2X7 receptors evaluated by means of cytofluorimetric, immunohistochemistry, reverse transcriptase-PCR, and Western blot analyses. P2X7 ligation by 2',3'-(benzoyl-4-benzoyl)-ATP (BzATP), a specific receptor agonist, was followed by an increase in intracellular Ca2+ and in the uptake of ethidium bromide. These effects were dramatically reduced by oxidized ATP (oATP), the P2X7 irreversible inhibitor, suggesting that the P2X7 is the functional receptor involved. At 24 h treatment of hPDLSCs with BzATP it enhanced the release of the pro-inflammatory agents IL8 and CCL20, without influencing cell viability. These effects were counteracted by pre-treating the cells with oATP or with A-740003, a selective and potent P2X7 competitive antagonist. Collectively, these results indicated that extracellular ATP mediate a pro-inflammatory response via P2X7 receptors in hPDLSCs opening a further approach to control hPDLSCs behavior in their possible application as therapeutic tool.
三磷酸腺苷（ATP）由人牙周膜细胞（hPDLCs）释放，并且已显示其通过激活P2Y受体来调节牙周膜再生以及对机械应力的反应。然而，据报道，这种核苷酸还主要通过P2X7受体诱导各种细胞类型趋化因子/细胞因子的成熟和/或释放，从而触发促炎级联反应。关于ATP在源自牙周膜的干细胞（hPDLSCs）中的可能作用知之甚少，而hPDLSCs被认为是基于细胞的治疗修复损伤的一种有前景的工具。鉴于P2X7在病理生理条件下所起的作用，在本研究中，我们调查了hPDLSCs中P2X7 ATP受体的表达。获得的结果表明，通过细胞荧光分析、免疫组织化学、逆转录聚合酶链反应和蛋白质免疫印迹分析评估，hPDLSCs表达P2X7受体。用特异性受体激动剂2'，3'-（苯甲酰-4-苯甲酰）-ATP（BzATP）连接P2X7后，细胞内钙离子（Ca2+）增加，溴化乙锭摄取增加。这些效应被P2X7不可逆抑制剂氧化ATP（oATP）显著降低，表明P2X7是相关的功能性受体。用BzATP处理hPDLSCs 24小时后，它增强了促炎因子白细胞介素8（IL8）和CC趋化因子配体20（CCL20）的释放，而不影响细胞活力。用oATP或选择性强效P2X7竞争性拮抗剂A-740003预处理细胞可抵消这些效应。总体而言，这些结果表明，细胞外ATP通过hPDLSCs中的P2X7受体介导促炎反应，为在hPDLSCs作为治疗工具的可能应用中控制其行为开辟了另一种途径。
