Traumatic brain injury (TBI) represents a major cause of long-term disability worldwide. Primary damage to brain tissue leads to complex secondary injury mechanisms involving inflammation, oxidative stress and cellular activation/reactivity. The molecular pathways that exacerbate brain cell dysfunction after injury are not well understood and provide challenges to developing TBI therapeutics. This study aimed to delineate mechanisms of astrocyte activation induced by mechano-stimulation, specifically involving extracellular adhesion and cationic transduction. An in vitro model was employed to investigate 2D and 3D cultures of primary astrocytes, in which cells were exposed to a single high-rate overpressure known to cause upregulation of structural and proliferative markers within 72 h of exposure. An inhibitor of focal adhesion kinase (FAK) phosphorylation, TAE226, was used to demonstrate a relationship between extracellular adhesion perturbations and structural reactivity in the novel 3D model. TAE226 mitigated upregulation of glial fibrillary acidic protein in 3D cultures by 72 h post-exposure. Alternatively, incubation with gadolinium (a cationic channel blocker) during overpressure, demonstrated a role for cationic transduction in reducing the increased levels of proliferating cell nuclear antigen that occur at 24 h post-stimulation. Furthermore, early changes in mitochondrial polarization at 15 min and in endogenous ATP levels at 4−6 h occur post-overpressure and may be linked to later changes in cell phenotype. By 24 h, there was evidence of increased amine metabolism and increased nicotinamide adenine dinucleotide phosphate oxidase (NOX4) production. The overproduction of NOX4 was counteracted by gadolinium during overpressure exposure. Altogether, the results of this study indicated that both extracellular adhesion (via FAK activation) and cationic conductance (via ion channels) contribute to early patterns of astrocyte activation following overpressure stimulation. Mechano-stimulation pathways are linked to bioenergetic and metabolic disruptions in astrocytes that influence downstream oxidative stress, aberrant proliferative capacity and structural reactivity.

颅脑外伤（TBI）代表了全球范围内长期残疾的主要原因。对脑组织的原发性损伤导致复杂的继发性损伤机制，涉及炎症，氧化应激和细胞活化/反应性。损伤后加重脑细胞功能障碍的分子途径尚未得到很好的理解，并为开发TBI治疗剂提出了挑战。这项研究旨在描述机械刺激诱导星形胶质细胞活化的机制，特别是涉及细胞外粘附和阳离子转导。一种在体外该模型被用于研究原代星形胶质细胞的2D和3D培养，其中细胞暴露于单一的高速超压下，该超高压已知会在暴露后72小时内引起结构和增殖标志物的上调。粘着斑激酶（FAK）磷酸化的抑制剂，TAE226，被用于证明新型3D模型中细胞外粘附扰动和结构反应性之间的关系。TAE226可以在暴露后72小时内减轻3D培养物中神经胶质纤维酸性蛋白的上调。或者，在超压过程中与（（阳离子通道阻滞剂）一起孵育，表明阳离子转导在减少刺激后24小时发生的增殖细胞核抗原水平升高方面发挥了作用。此外，线粒体极化的早期变化在15分钟后发生，内源性ATP水平在4-6 h发生在超压后，并且可能与后来的细胞表型变化有关。到24小时，有证据表明胺代谢增加，烟酰胺腺嘌呤二核苷酸磷酸氧化酶（NOX4）产生增加。在过压暴露期间，g可抵消NOX4的过量生产。总而言之，这项研究的结果表明，细胞外粘附（通过FAK激活）和阳离子电导（通过离子通道）都有助于超压刺激后星形胶质细胞激活的早期模式。机械刺激途径与星形胶质细胞的生物能和代谢破坏有关，影响下游的氧化应激，异常的增殖能力和结构反应性。