Department of Neurosurgery, Jinling Hospital, School of Medicine, Nanjing University, Nanjing, Jiangsu 210002, P.R. China.
Department of Neurosurgery, Zhongdu Hospital, Bengbu, Anhui 233004, P.R. China.
Int J Mol Med. 2018 Jun;41(6):3203-3210. doi: 10.3892/ijmm.2018.3567. Epub 2018 Mar 14.
The transcription factor nuclear factor-κB (NF-κB) has been shown to function as a key regulator of cell death or survival in neuronal cells. Previous studies indicate that the biphasic activation of NF-κB occurs following experimental neonatal hypoxia-ischemia and subarachnoid hemorrhage. However, the comprehensive understanding of NF-κB activity following traumatic brain injury (TBI) is incomplete. In the current study, an in vitro model of TBI was designed to investigate the NF-κB activity and expression of p65 and c-Rel subunits following traumatic neuronal injury. Primary cultured neurons were assigned to control and transected groups. NF-κB activity was detected by electrophoretic mobility shift assay. Western blotting and immunofluorescence were used to investigate the expression and distribution of p65 and c-Rel. Reverse transcription-quantitative polymerase chain reaction was performed to assess the downstream genes of NF-κB. Lactate dehydrogenase (LDH) quantification and trypan blue staining were used to estimate the neuronal injury. Double peaks of elevated NF-κB activity were observed at 1 and 24 h following transection. The expression levels of downstream genes exhibited similar changes. The protein levels of p65 also presented double peaks while c-Rel was elevated significantly in the late stage. The results of the trypan blue staining and LDH leakage assays indicated there was no sustained neuronal injury during the late peak of NF-κB activity. In conclusion, biphasic activation of NF-κB is induced following experimental traumatic neuronal injury. The elevation of p65 and c-Rel levels at different time periods suggests that within a single neuron, NF-κB may participate in different pathophysiological processes.
转录因子核因子-κB（NF-κB）已被证明在神经元细胞中作为细胞死亡或存活的关键调节剂发挥作用。先前的研究表明，NF-κB 的双相激活发生在实验性新生儿缺氧缺血和蛛网膜下腔出血后。然而，对于创伤性脑损伤（TBI）后 NF-κB 活性的全面理解并不完整。在本研究中，设计了一种体外 TBI 模型，以研究创伤性神经元损伤后 NF-κB 活性和 p65 和 c-Rel 亚基的表达。原代培养神经元分为对照组和横切组。通过电泳迁移率变动分析检测 NF-κB 活性。Western blot 和免疫荧光用于研究 p65 和 c-Rel 的表达和分布。逆转录定量聚合酶链反应用于评估 NF-κB 的下游基因。通过乳酸脱氢酶（LDH）定量和台盼蓝染色来评估神经元损伤。横切后 1 和 24 小时观察到 NF-κB 活性的双峰升高。下游基因的表达水平也发生了相似的变化。p65 的蛋白水平也呈现双峰，而 c-Rel 在晚期显著升高。台盼蓝染色和 LDH 漏出测定的结果表明，在 NF-κB 活性的晚期高峰期间没有持续的神经元损伤。总之，实验性创伤性神经元损伤后诱导 NF-κB 的双相激活。在不同时间点 p65 和 c-Rel 水平的升高表明，在单个神经元内，NF-κB 可能参与不同的病理生理过程。
