Department of Neurosurgery, Wuxi Taihu Hospital, Wuxi Clinical College of Anhui Medical University, Wuxi, 214044, Jiangsu, China.
Mol Biol Rep. 2023 Sep;50(9):7237-7244. doi: 10.1007/s11033-023-08619-7. Epub 2023 Jul 7.
Necroptosis, a newly defined regulatable necrosis with membrane disruption, has been demonstrated to participate in trauma brain injury (TBI) related neuronal cell death. Heat shock protein 70 (HSP70) is a stress protein with neuroprotective activity, but the potential protective mechanisms are not fully understood.
Here, we investigated the effects of HSP70 regulators in a cellular TBI model induced by traumatic neuronal injury (TNI) and glutamate treatment. We found that necroptosis occurred in cortical neurons after TNI and glutamate treatment. Neuronal trauma markedly upregulated HSP70 protein expression within 24 h. The results of immunostaining and lactate dehydrogenase release assay showed that necroptosis following neuronal trauma was inhibited by HSP70 activator TRC051384 (TRC), but promoted by the HSP70 inhibitor 2-phenylethyenesulfonamide (PES). In congruent, the expression and phosphorylation of receptor interacting protein kinase 3 (RIPK3) and mixed lineage kinase domain-like protein (MLKL) were differently regulated by HSP70. Furthermore, the expression of HSP90α induced by neuronal trauma was further promoted by PES but decreased by TRC. The data obtained from western blot showed that the phosphorylation of RIPK3 and MLKL induced by HSP70 inhibition were reduced by RIPK3 inhibitor GSK-872 and HSP90α inhibitor geldanamycin (GA). Similarly, inhibition of HSP90α with GA could partially prevented the increased necroptosis induced by PES.
Taken together, HSP70 activation exerted protective effects against neuronal trauma via inhibition of necroptosis. Mechanistically, the HSP90α-mediated activation of RIPK3 and MLKL is involved in these effects.
坏死性凋亡是一种新定义的具有膜破裂的可调控性坏死，已被证明参与创伤性脑损伤（TBI）相关的神经元细胞死亡。热休克蛋白 70（HSP70）是一种具有神经保护活性的应激蛋白，但潜在的保护机制尚不完全清楚。
在这里，我们研究了 HSP70 调节剂在创伤性神经元损伤（TNI）和谷氨酸处理诱导的细胞性 TBI 模型中的作用。我们发现 TNI 和谷氨酸处理后皮质神经元发生坏死性凋亡。创伤性神经元明显上调 HSP70 蛋白表达在 24 小时内。免疫染色和乳酸脱氢酶释放试验的结果表明，HSP70 激活剂 TRC051384（TRC）抑制神经元创伤后的坏死性凋亡，但 HSP70 抑制剂 2-苯乙磺酰胺（PES）促进坏死性凋亡。相应地，受体相互作用蛋白激酶 3（RIPK3）和混合谱系激酶结构域样蛋白（MLKL）的表达和磷酸化受 HSP70 调节。此外，神经元创伤诱导的 HSP90α 的表达进一步被 PES 促进，而被 TRC 降低。Western blot 获得的数据表明，HSP70 抑制诱导的 RIPK3 和 MLKL 的磷酸化被 RIPK3 抑制剂 GSK-872 和 HSP90α 抑制剂格尔德霉素（GA）减少。同样，GA 抑制 HSP90α 可部分阻止 PES 诱导的坏死性凋亡增加。
综上所述，HSP70 的激活通过抑制坏死性凋亡对神经元创伤发挥保护作用。在机制上，HSP90α 介导的 RIPK3 和 MLKL 的激活参与了这些作用。
