P2RX7是一种嘌呤受体，具有多个活性，在炎症组织中发现高水平的胞外ATP会激活该受体。P2RX7具有免疫调节和抗肿瘤特性，因此是治疗各种疾病的治疗靶点。已经开发了几种化合物，旨在抑制或增强其活性。但是，对P2RX7活性影响的研究仅限于需要使用非生理介质的体外和体外研究。目前，评估P2RX7调节剂在体内对受体的活性的唯一方法是间接的。我们成功开发了一种协议，利用其独特的大孔形成能力，在小鼠肺中检测P2RX7活化，从而解决了这个问题。该协议基于TO-PRO™-3的鼻内给药和流式细胞术测量荧光。我们表明，ATP主要以P2RX7依赖方式增强了小鼠肺免疫细胞中的TO-PRO™-3荧光。所描述的方法已成功分析了WT和转基因C57BL6小鼠肺中的P2RX7活性。提供的详细指导和建议将支持使用该协议研究针对P2RX7的药理或生物化合物的效力。©2023.作者。

P2RX7 is a purinergic receptor with pleiotropic activities that is activated by high levels of extracellular ATP that are found in inflamed tissues. P2RX7 has immunomodulatory and anti-tumor proprieties and is therefore a therapeutic target for various diseases. Several compounds are developed to either inhibit or enhance its activation. However, studying their effect on P2RX7's activities is limited to in vitro and ex vivo studies that require the use of unphysiological media that could affect its activation. Up to now, the only way to assess the activity of P2RX7 modulators on the receptor in vivo was in an indirect manner.We successfully developed a protocol allowing the detection of P2RX7 activation in vivo in lungs of mice, by taking advantage of its unique macropore formation ability. The protocol is based on intranasal delivery of TO-PRO™-3, a non-permeant DNA intercalating dye, and fluorescence measurement by flow cytometry. We show that ATP enhances TO-PRO™-3 fluorescence mainly in lung immune cells of mice in a P2RX7-dependant manner.The described approach has allowed the successful analysis of P2RX7 activity directly in the lungs of WT and transgenic C57BL6 mice. The provided detailed guidelines and recommendations will support the use of this protocol to study the potency of pharmacologic or biologic compounds targeting P2RX7.© 2023. The Author(s).