Heat stress-activated P2X7 receptor induces astrocyte activation and regulates glioma tumor microenvironment via calcium signaling pathway

Background and aims: The effects of adjuvant hyperthermia on glioblastoma-associated astrocytes remain poorly characterized. This study aimed to investigate the role of the purinergic P2X7 receptor, an ATP-gated ion channel, in mediating heat-induced astrocyte activation and its impact on tumor progression. Methods: Primary mouse astrocytes were subjected to heat stress (mild hyperthermia at 42°C). P2X7 signaling was examined using a specific antagonist (A-740003), siRNA-mediated knockdown, and live-cell calcium imaging. Astrocyte activation was evaluated by assessing Glial Fibrillary Acidic Protein (GFAP) expression and pro-inflammatory markers. The pro-tumorigenic potential of astrocyte-conditioned medium was tested on U87 glioblastoma cells. An orthotopic mouse model was used to validate the effects of local hyperthermia, with or without P2X7 inhibition. Results: Mild hyperthermia upregulated P2X7 expression in astrocytes and induced a substantial calcium influx, leading to activation of the Calcineurin-NFAT pathway. This process promoted a reactive astrocyte phenotype and a pro-tumorigenic secretory profile, enhancing U87 cell proliferation, migration, and invasion. In vivo, mild hyperthermia was associated with increased tumor progression, which was attenuated by pharmacological inhibition of P2X7. Conclusion: Heat stress facilitates glioblastoma progression by activating astrocytes through the P2X7-mediated calcium–calcineurin–NFAT signaling pathway. These findings highlight P2X7 as a potential therapeutic target for optimizing hyperthermia-based strategies in glioblastoma treatment.