Zhou Qinxiang, Tang Hao, Bai Dingqun, Kong Yuhan
Department of Rehabilitation Medicine, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China.
Chongqing Key Laboratory of Neurology, The First Affiliated Hospital of Chongqing Medical University, Chongqing, China.
PeerJ. 2022 Feb 15;10:e12886. doi: 10.7717/peerj.12886. eCollection 2022.
7,8-dihydroxyflavone (7,8-DHF), a selective agonist of tropomyosin related kinase receptor B (TrkB), is known to exert protective effects in neurodegenerative diseases. However, the role of 7,8-DHF in TrkB signaling after ischemic stroke has remained elusive.
In the vitro model of ischemic stroke, we investigated the neuroprotective effect of 7,8-DHF through activation of TrkB signaling. Neurons subjected to oxygen and glucose deprivation/reperfusion were treated with the protein kinase inhibitor K252a and a knockdown of TrkB. Cell counting kit-8 (CCK-8) assay, Flow Cytometric Analysis (FACS), TdT-mediated dUTP nick end labeling (TUNEL) assay were conducted for measuring cell viability and numbers of apoptotic cells. And apoptosis-associated proteins were analyzed by Western blotting.
Compared with the Control group, OGD/R group revealed lower cell viability by CCK-8 assay FACS and TUNEL assay showed increased rates of neuronal apoptosis. However, 7,8-DHF treatment increased cell viability and reduced neuronal apoptosis. Western blotting indicated upregulated Bax and cleaved caspase-3 and but downregulated Bcl-2 following OGD/R. Whereas 7,8-DHF treatment downregulated Bax and cleaved caspase-3 but upregulated Bcl-2. These changes were accompanied by a significant increase in the phosphorylation of TrkB and Akt following 7,8-DHF administration. However, the administration of K252a and knockdown of TrkB could alleviate those effects.
Our study demonstrates that activation of TrkB signaling by 7,8-DHF protects neurons against OGD/R injury via the TrkB/Akt pathway, which provides the evidence for the role of TrkB signaling in OGD-induced neuronal damage and may become a potential therapeutic target for ischemic stroke.
7,8-二羟基黄酮（7,8-DHF）是原肌球蛋白相关激酶受体B（TrkB）的选择性激动剂，已知其在神经退行性疾病中发挥保护作用。然而，7,8-DHF在缺血性中风后TrkB信号传导中的作用仍不清楚。
在缺血性中风的体外模型中，我们通过激活TrkB信号传导研究了7,8-DHF的神经保护作用。用蛋白激酶抑制剂K252a和TrkB敲低处理经历氧和葡萄糖剥夺/再灌注的神经元。进行细胞计数试剂盒-8（CCK-8）测定、流式细胞术分析（FACS）、TdT介导的dUTP缺口末端标记（TUNEL）测定以测量细胞活力和凋亡细胞数量。并通过蛋白质印迹分析凋亡相关蛋白。
与对照组相比，氧糖剥夺/再灌注（OGD/R）组通过CCK-8测定显示细胞活力较低，FACS和TUNEL测定显示神经元凋亡率增加。然而，7,8-DHF处理提高了细胞活力并减少了神经元凋亡。蛋白质印迹表明，OGD/R后促凋亡蛋白Bax和裂解的半胱天冬酶-3上调，但抗凋亡蛋白Bcl-2下调。而7,8-DHF处理使Bax和裂解的半胱天冬酶-3下调，但使Bcl-2上调。这些变化伴随着7,8-DHF给药后TrkB和Akt磷酸化的显著增加。然而，K252a的给药和TrkB的敲低可以减轻这些作用。
我们的研究表明，7,8-DHF激活TrkB信号传导可通过TrkB/Akt途径保护神经元免受OGD/R损伤，这为TrkB信号传导在OGD诱导的神经元损伤中的作用提供了证据，并可能成为缺血性中风的潜在治疗靶点。
