Department of Paediatrics, Linyi People's Hospital, Linyi, China.
Department of Paediatrics, Linyi People's Hospital, Linyi, China.
J Biol Chem. 2021 Aug;297(2):100537. doi: 10.1016/j.jbc.2021.100537. Epub 2021 Mar 12.
The present study aimed to explore specific mechanisms involved in mediating the neuroprotective effects of Smad ubiquitination regulatory factor 2 (Smurf2) in cerebral ischemic injury. A middle cerebral artery occlusion (MCAO) mouse model and an oxygen-glucose deprivation (OGD)-treated neuron model were developed. The expression of Smurf2, Yin Yang 1 (YY1), hypoxia-inducible factor-1 alpha (HIF1α), and DNA damage-inducible transcript 4 gene (DDIT4) was analyzed. Thereafter, the expression of Smurf2, YY1, HIF1α, and DDIT4 was altered in the MCAO mice and OGD-treated neurons. Apoptosis in tissues and cerebral infarction were assessed. In neurons, the expression of apoptosis-related proteins, viability, and apoptosis were assessed, followed by evaluation of lactate dehydrogenase leakage rate. The interaction between Smurf2 and YY1 was analyzed by coimmunoprecipitation assay and that between YY1 ubiquitination by in vivo ubiquitination experiment. The results showed downregulation of Smurf2 and upregulation of YY1, HIF1α, and DDIT4 in both MCAO mice and OGD-treated neurons. Smurf2 elevated YY1 ubiquitination and degradation, and YY1 increased HIF1α expression to promote DDIT4 in neurons. Overexpressed Smurf2 or downregulated YY1, HIF1α, or DDIT4 reduced the volume of cerebral infarction and apoptosis in MCAO mice, while enhancing cell viability and reducing apoptosis and lactate dehydrogenase leakage in OGD-treated neurons. In summary, our findings elucidated a neuroprotective role of Smurf2 in cerebral ischemic injury via inactivation of the YY1/HIF1α/DDIT4 axis.
本研究旨在探讨介导 Smad 泛素化调节因子 2 (Smurf2) 在脑缺血损伤中神经保护作用的具体机制。建立了大脑中动脉闭塞 (MCAO) 小鼠模型和氧葡萄糖剥夺 (OGD) 处理神经元模型。分析了 Smurf2、Yang Yang 1 (YY1)、缺氧诱导因子-1α (HIF1α) 和 DNA 损伤诱导转录物 4 基因 (DDIT4) 的表达。此后，改变了 MCAO 小鼠和 OGD 处理神经元中的 Smurf2、YY1、HIF1α 和 DDIT4 的表达。评估了组织和脑梗死中的细胞凋亡。在神经元中，评估了凋亡相关蛋白的表达、活力和凋亡情况，然后评估乳酸脱氢酶漏出率。通过共免疫沉淀测定分析 Smurf2 和 YY1 之间的相互作用，并通过体内泛素化实验分析 YY1 泛素化。结果显示，在 MCAO 小鼠和 OGD 处理神经元中 Smurf2 下调和 YY1、HIF1α 和 DDIT4 上调。Smurf2 升高了 YY1 的泛素化和降解，YY1 增加了 HIF1α 的表达，促进了神经元中的 DDIT4。过表达 Smurf2 或下调 YY1、HIF1α 或 DDIT4 减少了 MCAO 小鼠的脑梗死体积和凋亡，同时增强了细胞活力并减少了 OGD 处理神经元中的凋亡和乳酸脱氢酶漏出。总之，我们的研究结果阐明了 Smurf2 通过失活 YY1/HIF1α/DDIT4 轴在脑缺血损伤中的神经保护作用。
