Shi Wei, Sun Yanqing, Wang Juncheng, Tang Yifan, Zhou Shengyuan, Xu Zheng, Yuan Bo, Geng Xiangwu, Chen Xiongsheng
Spine Center, Department of Orthopaedics, Shanghai Changzheng Hospital, Second Military Medical University Shanghai, China.
Department of Orthopedics, Naval Special Medical Center, Second Military Medical University Shanghai, China.
Am J Transl Res. 2021 Jun 15;13(6):6117-6125. eCollection 2021.
This research aimed to study the impact and regulatory mechanism of Trem1 in spinal cord ischemia-reperfusion injury (SCIRI).
Temporary aortic cross clamp followed by reperfusion was used to establish SCIRI mice model. Mice motion function was estimated by Basso, Beattie, Bresnahan (BBB) score. Spinal cord infract zone was analyzed by HE and TUNEL staining. High throughput sequencing was performed to explore potential target for SCIRI. N2a cells were used to simulate the pathophysiological process of SCIRI in vitro with oxygen-glucose-serum deprivation/restoration (OGSD/R). RT-PCR and Western blot were token to determine mRNA and protein expression levels. Knockdown of Trem1 was performed with siRNA transfection and shRNA adenovirus injection . The relationship between Trem1 and SYK was analyzed by immunoprecipitation and immunofluorescence.
We observed that neuronal apoptosis of spinal cord was aggravated after SCIRI. Trem1 expression was dramatically upregulated as shown by high throughput sequencing, RT-PCR and Western blot results. Furthermore, Trem1 triggered apoptosis of N2a cells induced by OGSD/R, and knockdown of Trem1 by siRNAs blocked apoptosis via PI3K/AKT and NF-κB signaling pathway by interacting with SYK. In addition, we found that intrathecal injection of adenovirus with Trem1 shRNA could downregulate SYK and inhibit neuron apoptosis caused by SCIRI .
Trem1 interacts with SYK and mediates neuronal apoptosis via the PI3K/AKT and NF-κB signaling pathway. Trem1 may be a therapeutic candidate for patients with SCIRI.
本研究旨在探讨触发受体表达分子1（Trem1）在脊髓缺血再灌注损伤（SCIRI）中的作用及调控机制。
采用临时主动脉交叉夹闭并再灌注的方法建立SCIRI小鼠模型。通过Basso、Beattie、Bresnahan（BBB）评分评估小鼠运动功能。采用苏木精-伊红（HE）染色和末端脱氧核苷酸转移酶介导的缺口末端标记（TUNEL）染色分析脊髓梗死区域。进行高通量测序以探索SCIRI的潜在靶点。在体外采用氧-葡萄糖-血清剥夺/恢复（OGSD/R）处理神经母细胞瘤细胞（N2a细胞）以模拟SCIRI的病理生理过程。采用逆转录聚合酶链反应（RT-PCR）和蛋白质免疫印迹法检测mRNA和蛋白表达水平。通过小干扰RNA（siRNA）转染和短发夹RNA（shRNA）腺病毒注射敲低Trem1。通过免疫沉淀和免疫荧光分析Trem1与脾酪氨酸激酶（SYK）之间的关系。
我们观察到SCIRI后脊髓神经元凋亡加剧。高通量测序、RT-PCR和蛋白质免疫印迹结果显示Trem1表达显著上调。此外，Trem1可触发OGSD/R诱导的N2a细胞凋亡，siRNA敲低Trem1可通过与SYK相互作用，经磷脂酰肌醇-3激酶（PI3K）/蛋白激酶B（AKT）和核因子κB（NF-κB）信号通路阻断凋亡。此外，我们发现鞘内注射携带Trem1 shRNA的腺病毒可下调SYK并抑制SCIRI引起的神经元凋亡。
Trem1与SYK相互作用，并通过PI3K/AKT和NF-κB信号通路介导神经元凋亡。Trem1可能是SCIRI患者的一个潜在治疗靶点。
