电针通过KAT3B介导的ACSL4琥珀酰化抑制铁蛋白沉积以缓解脑缺血再灌注损伤
摘要
电针（EA）是中国传统针灸与现代电疗相结合的一种治疗脑缺血再灌注损伤（CIRI）的有效方法。然而，EA在CIRI过程中的下游分子机制仍是一个未知数。本研究旨在揭示EA对CIRI大鼠的治疗作用及其调控机制。首先，我们构建了大脑中动脉闭塞（MCAO）大鼠模型，然后用EA治疗，观察其病理变化。结果表明，EA能减少MCAO模型大鼠的梗死体积（43.81 ± 3.34 vs 15.96 ± 2.22）和神经系统评分（3.33 ± 0.52 vs 1.67 ± 0.52），并抑制细胞凋亡。在铁变态反应分析中，EA降低了MCAO模型大鼠的Fe2 +（0.08±0.01 vs 0.06±0.01）、MDA（36.61±4.29 vs 21.72±2.79）和LPS（5.25±0.69 vs 2.89±0.42）含量，增加了GSH（4.94±1.04 vs 11.69±1.88）含量。我们接下来检测了琥珀酰化是否介导了 EA 处理的 I/R 损伤。根据免疫沉淀和 Western 印迹分析，EA 治疗可降低 MCAO 大鼠体内琥珀酰化和 KAT3B 的水平。此外，机制实验还发现，KAT3B能促进铁突变相关蛋白ACSL4在K661位点的琥珀酰化，从而稳定ACSL4。最后，给经EA处理的MCAO大鼠进一步注射KAT3B表达载体。结果显示，KAT3B的过表达增加了EA处理的MCAO模型大鼠的梗死体积（31.44 ± 3.92 vs 7.94 ± 2.84）和神经评分（2.67 ± 0.51 vs 1.33 ± 0.51），并促进了细胞凋亡。在铁变态反应分析中，KAT3B的过表达增加了EA处理的MCAO模型大鼠的Fe2 + (0.08 ± 0.01 vs 0.05 ± 0.01)、MDA (29.24 ± 4.30 vs 22.06 ± 1.89)和LPO (5.07 ± 0.45 vs 2.88 ± 0.49)含量，降低了GSH (7.86 ± 1.09 vs 11.06 ± 1.76)含量。总之，我们的研究表明，EA通过抑制KAT3B诱导的ACSL4稳定化来抑制铁氧化，从而在CIRI中发挥治疗作用。这些发现有助于我们了解 EA 神经保护作用的分子机制，并为开发 CIRI 的创新治疗策略开辟了新途径。Electro-acupuncture (EA) is identified as an effective therapeutic method for cerebral ischemia/reperfusion injury (CIRI), which is a combination of Chinese traditional acupuncture and modern electro-therapy. However, the downstream molecular mechanisms of EA in CIRI process remains largely unknown. The purpose of the present study is to unveil the therapeutic effect of EA on CIRI rat and its regulatory mechanisms. At first, we constructed middle cerebral artery occlusion (MCAO) rat models and then treated them with EA to observe the pathological changes. The results indicated that EA decreased the infarct volume (43.81 ± 3.34 vs 15.96 ± 2.22) and the neurological scores (3.33 ± 0.52 vs 1.67 ± 0.52) and suppressed the apoptosis in MCAO model rats. For ferroptosis analysis, EA decreased the Fe2 + (0.08 ± 0.01 vs 0.06 ± 0.01), MDA (36.61 ± 4.29 vs 21.72 ± 2.79), and LPS (5.25 ± 0.69 vs 2.89 ± 0.42) contents and increased the GSH (4.94 ± 1.04 vs 11.69 ± 1.88) content in MCAO model rats. We next detected whether succinylation mediated EA-treated I/R injury. According to immunoprecipitation and western blot analysis, EA treatment could lower both levels of succinylation and KAT3B in MCAO rats. Moreover, mechanism experiments unveiled that KAT3B promoted the succinylation of the ferroptosis-related protein ACSL4 at K661 site and thus stabilizing ACSL4. Finally, EA-treated MCAO rats were further injected with KAT3B expression vector. The results showed that KAT3B overexpression increased the infarct volume (31.44 ± 3.92 vs 7.94 ± 2.84) and the neurological scores (2.67 ± 0.51 vs 1.33 ± 0.51) and promoted the apoptosis in EA treated MCAO model rats. For ferroptosis analysis, KAT3B overexpression increased the Fe2 + (0.08 ± 0.01 vs 0.05 ± 0.01), MDA (29.24 ± 4.30 vs 22.06 ± 1.89), and LPO (5.07 ± 0.45 vs 2.88 ± 0.49) contents and decreased the GSH (7.86 ± 1.09 vs 11.06 ± 1.76) content in EA treated MCAO model rats. Collectively, our study demonstrates that EA plays a therapeutic role in CIRI through suppressing KAT3B-induced stabilization of ACSL4 to inhibit ferroptosis. These findings contribute to our understanding of the molecular mechanisms underlying the neuroprotective effects of EA and open new avenues for the development of innovative therapeutic strategies for CIRI.
求助内容：
应助结果提醒方式：
京公网安备 11010802042870号
