硫化氢在大鼠弥漫性轴索损伤中的作用

Role of hydrogen sulfide in diffuse axonal injury in rats

目的探讨弥漫性轴索损伤(diffuse axonal injury,DAI)后脑组织内源性硫化氢(hydrogen sulfide,H2S)含量的变化及外源性H2S对DAI的作用,阐明H2S作为一种新的气体信号分子在脑组织中发挥的作用,为DAI后神经功能障碍的防治提供理论基础。方法采用大鼠头颅瞬间旋转装置制备大鼠DAI模型,给予外源性H2S干预,以造模后6h、24h、7d为时间点,分别用亚甲蓝分光光度法检测脑组织H2S含量的变化,流式细胞仪检测海马CA3区神经元的凋亡及Western blot检测脑组织β淀粉样前体蛋白(β-amyloid precursor protein,β-APP)的变化。结果 DAI模型组皮层、海马、脑干内源性H2S的含量较对照组均显著增加,且DAI+H2S干预组脑组织H2S含量增加更明显;DAI模型组较对照组海马神经元早期凋亡百分比明显增加,致伤时间越长,凋亡百分比越高,且DAI(7d)+H2S干预组均较DAI(7d)模型组的海马神经元凋亡百分比明显增高;DAI模型组皮层、海马、脑干β-APP的表达较对照组明显增高,且DAI+H2S干预组均较DAI模型组β-APP的表达更明显。结论 DAI后脑组织神经元凋亡及β-APP表达的增加可能与脑组织自身内源性H2S含量的增加有关,其可能作为一种新的神经递质参与DAI的继发性神经元损伤过程。

Objective To detect the content changes of endogenous hydrogen sulfide （HzS） in the cerebral tissues after diffuse axonal injury （DAI） and observe the role of exogenous H2S in DAI so as to clarify the role of H2 S as a new gas signaling molecule involved in the process of DAI and provide rationale supports for the prevention and treatment of neurological dysfunction after DAI. Methods The DAI model was established by a coronal rotation device and exogenous HzS was administered for intervention. Time points of 6 h, 24 h and 7 d after DAI were chosen for observation. Methylene blue spectrophotometric method was used for detecting H2 S content change in the brain tissues after DAI, flow cytometry for measuring the apoptosis of hippocampal CA3 neurons, and Western blot for determining the change of β-amyloid precursor protein （β-APP） expression in the cortex, hippocampus and brain stem. Results Compared with that in the control group, endogenous H2S content in the cortex, hippocampus and brainstem significantly increased in DAI groups, with DAI ＋ H2S intervention groups having significantly higher H2 S content than DAI model groups. The apoptosis of hippocampal CA3 neurons showed an increased early apoptosis percentage in the DAI model groups and more apoptotic neuron ceils were observed inthe longer time post-injury in DAI model groups. Furthermore, DAI （7 d） ~ HzS group had a significantly higher early apoptosis percentage than DAI （7 d） group. More importantly, Western blot showed that 13-APP in the cortex, hippocampus and brain stem in DAI model groups was expressed higher than in the control group, with DA! ~ HzS intervention groups having a significantly higher I^-APP expression than DAI model groups. Gonclusion The increase of neuronal apoptosis and ^-APP expression may be related to the increase of endogenous Hz S after DAI. It suggests that H2S may be involved in the secondary neuronal injury of DAI as a new neurotransmitter.