百事乐加味方对卒中后抑郁大鼠海马神经元自噬蛋白的影响
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摘要
目的观察百事乐加味方对卒中后抑郁(PSD)大鼠神经功能缺损评分、形态学改变及海马自噬蛋白m TOR、Beclin-1、LC-3Ⅱ表达的影响,探讨PSD神经元细胞自噬机制及中药防治作用。方法采用大脑中动脉栓塞+慢性不可预见温和应激+孤养法制作PSD模型,分组前进行行为学评分,随机分为假手术组、抑郁组、卒中组、PSD组、阳性药组、百事乐加味方组,各给药组给予相应药物,连续28 d。HE、Nissl染色观察大鼠海马形态;免疫组化检测PSD大鼠海马m TOR、Beclin-1、LC-3Ⅱ的表达。结果与假手术组比较,PSD组大鼠神经功能缺损评分明显升高,海马m TOR表达明显降低,Beclin-1、LC-3Ⅱ表达明显升高(P<0.01);与PSD组比较,百事乐加味方组大鼠功能缺损评分显著降低,海马组织病理形态明显改善,海马m TOR蛋白表达明显升高,Beclin-1、LC-3Ⅱ蛋白表达明显降低(P<0.05,P<0.01)。结论百事乐加味方可有效改善PSD大鼠行为学、病理学表现,抑制神经细胞异常自噬可能是其抗PSD的重要机制之一。
Objective To observe the effects of modified Baishile Formula on the neurological deficit score and the expressions of mTOR, Beclin-1 and LC-3Ⅱ in hippocampus of post-stroke depression(PSD) rats; To explore the preventive and therapeutic mechanism of TCM for post-stroke depression from neuronal autophagy. Methods The rat model of focal cerebral ischemia was established by the method of middle cerebral artery occlusion(MCAO), and the PSD model was established by combining with chronic unpredictable mild stress(CUMS) and solitary nourishment. The behavioral scores were evaluated by Longa grading method before dividing the groups. The rats were randomly divided into sham-operation group, depression group, post-stroke group, PSD group, positive medicine group and modified Baishile Formula group. Each administration group was given relevant medicine for 28 d. The morphological changes of hippocampus were observed by HE staining and Nissl staining. Immunohistochemical method was used to detect the expressions of mTOR, Beclin-1 and LC-3Ⅱ in hippocampus of PSD rats. Results Compared with the sham-operation group, the neurological deficit score of the PSD group significantly increased, the expression of mTOR in the hippocampus significantly decreased, and the expressions of Beclin-1 and LC-3Ⅱ significantly increased(P<0.01). Compared with the PSD group, the neurological deficit score of the rats in modified Baishile Formula groupsignificantly decreased, the pathological morphology of the hippocampus was significantly improved, the expression of m TOR protein in the hippocampus significantly increased, and the expression of Beclin-1 and LC-3Ⅱ proteins significantly decreased(P<0.05, P<0.01). Conclusion Modified BaishileFormula can effectively improve the behavioral and pathological symptoms of PSD rats and inhibit autonomic phagocytosis. It may be one of the important mechanisms of treating PSD.
Objective To observe the effects of modified Baishile Formula on the neurological deficit score and the expressions of mTOR, Beclin-1 and LC-3Ⅱ in hippocampus of post-stroke depression(PSD) rats; To explore the preventive and therapeutic mechanism of TCM for post-stroke depression from neuronal autophagy. Methods The rat model of focal cerebral ischemia was established by the method of middle cerebral artery occlusion(MCAO), and the PSD model was established by combining with chronic unpredictable mild stress(CUMS) and solitary nourishment. The behavioral scores were evaluated by Longa grading method before dividing the groups. The rats were randomly divided into sham-operation group, depression group, post-stroke group, PSD group, positive medicine group and modified Baishile Formula group. Each administration group was given relevant medicine for 28 d. The morphological changes of hippocampus were observed by HE staining and Nissl staining. Immunohistochemical method was used to detect the expressions of mTOR, Beclin-1 and LC-3Ⅱ in hippocampus of PSD rats. Results Compared with the sham-operation group, the neurological deficit score of the PSD group significantly increased, the expression of mTOR in the hippocampus significantly decreased, and the expressions of Beclin-1 and LC-3Ⅱ significantly increased(P<0.01). Compared with the PSD group, the neurological deficit score of the rats in modified Baishile Formula groupsignificantly decreased, the pathological morphology of the hippocampus was significantly improved, the expression of m TOR protein in the hippocampus significantly increased, and the expression of Beclin-1 and LC-3Ⅱ proteins significantly decreased(P<0.05, P<0.01). Conclusion Modified BaishileFormula can effectively improve the behavioral and pathological symptoms of PSD rats and inhibit autonomic phagocytosis. It may be one of the important mechanisms of treating PSD.
引文
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[12] NAZIM U M, MOON J H, LEE Y J, et al. Park SYG lipizide sensitizes lungcancercellstoTRAIL-inducedapoptosisviaAkt/mTOR/autophagy pathways oncotarget[J]. Oncotarget,2017,8(59):100021-100033.
[13] LI W D, LI N P, SONG D D, et al. Metformin inhibits endothelial progenitor cell migration by decreasing matrix metalloproteinases,MMP-2 and MMP-9, via the AMPK/mTOR/autophagy pathway[J]. Int J Mol Med,2017,39(5):1262-1268.
[14] ZHU Y, WANG J. Wogonin increasesβ-amyloid clearance and inhibits tau phosphorylation via inhibition of mammalian target of rapamycin:potential drug to treat Alzheimer's disease[J].Neurol Sci,2015,36(7):1181-1188.
[15]郭向飞,赵雅宁,李建民,等.mTOR/自噬通路在间歇性低氧全脑缺血再灌注大鼠海马区的表达及意义[J].中华耳鼻咽喉头颈杂志,2016,51(10):761-767.
[16] WANG Y, WANG W, LI D, et al. IGF-1 alleviates NMDA-induced excitotoxicity in cultured hippocampal neurons against autophagy viatheNR2B/PI3K-AKT-mTORpathway[J].JCellPhysiol,2014,229(11):1618-1129.
[17]SHEHATAM,MATUMURAH,OKUBO-SUZUKI,etal.Neuronal stimulation induces autophagy in hippocampal neurons that is involved in AMPA receptor degradation after chemical long-term depression[J]. J Neurosci,2012,32(30):10413-10422.
[18] LI R, JIN Y, LI Q, et al. MiR-93-5p targeting PTEN regulates the NMDA-induced autophagy of retinal ganglion cells via AKT/mTOR pathway in glaucoma[J]. Biomed Pharmacother,2018,100:1-7.
[19]范世珍,陈旭娜,于波海,等.姜黄素通过抑制PI3K/Akt/mTOR促进SiHa细胞自噬[J].中国实验诊断学,2017,21(5):897-900.
[20]吴仲敏,程正文,倪桂莲,等.人参皂苷Rg1对创伤后应激障碍大鼠行为学变化和海马神经元自噬的影响[J].中国病理生理杂志,2017,33(5):896-901.
[2]刘林,杨蕙,刘羽,等.百事乐加味方对卒中后抑郁大鼠海马-前额叶皮层环路形态及BDNF影响的实验研究[J].中华中医药杂志,2016,31(4):1398-1401.
[3]刘林,刘羽,廖亮英,等.百事乐加味方抗抑郁作用的实验研究[J].中华中医药学刊,2016,34(2):317-319.
[4] LONGA E Z, WEISTEIN P R, CARLSON S. Reversible middle cerebral artery occlusion without craniectomy in rats[J]. Stroke,1989,20:84-89.
[5]李丹丹,肖学进,秦书俭,等.脑缺血再灌注损伤程度与自噬相关基因LC3、mTOR表达的相关性研究[J].中国临床解剖学杂志,2018,36(2):182-186.
[6]徐爱军,刘昊,田艳霞,等.柴胡疏肝散对抑郁症大鼠行为学和海马神经元凋亡及自噬的影响[J].吉林大学学报,2014,40(4):801-804.
[7]BOCKAERTJ,MARINP.mTORinbrainphysiologyand pathologies[J]. Physiol Rev,2015,95(4):1157-1187.
[8] SINGH A K, KASHYAP M P, TRIPATHI V K. Neuroprotection through rapamycin-induced activation-of autophagy and PI3K/Akt1/mTOR/CREBsignalingagainstAmyloid-β-inducedoxidativestress,synaptic/neurotransmission dysfunction, and neurodegeneration in adult rats[J]. Mol Neurobiol,2017,54(8):5815-5828.
[9] PAZINI F L, CUMHA M P, ROSA J M. Creatine, similar to ketamine,counteracts depressive-like behavior induced by corticosterone via PI3K/Akt/mTOR pathway[J]. Mol Neurobiol,2016,53(10):6818-6834.
[10]沈忠飞,王志坚,潘巍巍,等.氟西汀调控+CUMS+抑郁大鼠海马突触重塑[J].中国病理生理杂志,2016,32(9):1642-1647.
[11] JIA J, LE W. Molecular network of neuronal autophagy in the pathophysiology and treatment of depression[J]. Neurosci Bull,2015,31(4):427-434.
[12] NAZIM U M, MOON J H, LEE Y J, et al. Park SYG lipizide sensitizes lungcancercellstoTRAIL-inducedapoptosisviaAkt/mTOR/autophagy pathways oncotarget[J]. Oncotarget,2017,8(59):100021-100033.
[13] LI W D, LI N P, SONG D D, et al. Metformin inhibits endothelial progenitor cell migration by decreasing matrix metalloproteinases,MMP-2 and MMP-9, via the AMPK/mTOR/autophagy pathway[J]. Int J Mol Med,2017,39(5):1262-1268.
[14] ZHU Y, WANG J. Wogonin increasesβ-amyloid clearance and inhibits tau phosphorylation via inhibition of mammalian target of rapamycin:potential drug to treat Alzheimer's disease[J].Neurol Sci,2015,36(7):1181-1188.
[15]郭向飞,赵雅宁,李建民,等.mTOR/自噬通路在间歇性低氧全脑缺血再灌注大鼠海马区的表达及意义[J].中华耳鼻咽喉头颈杂志,2016,51(10):761-767.
[16] WANG Y, WANG W, LI D, et al. IGF-1 alleviates NMDA-induced excitotoxicity in cultured hippocampal neurons against autophagy viatheNR2B/PI3K-AKT-mTORpathway[J].JCellPhysiol,2014,229(11):1618-1129.
[17]SHEHATAM,MATUMURAH,OKUBO-SUZUKI,etal.Neuronal stimulation induces autophagy in hippocampal neurons that is involved in AMPA receptor degradation after chemical long-term depression[J]. J Neurosci,2012,32(30):10413-10422.
[18] LI R, JIN Y, LI Q, et al. MiR-93-5p targeting PTEN regulates the NMDA-induced autophagy of retinal ganglion cells via AKT/mTOR pathway in glaucoma[J]. Biomed Pharmacother,2018,100:1-7.
[19]范世珍,陈旭娜,于波海,等.姜黄素通过抑制PI3K/Akt/mTOR促进SiHa细胞自噬[J].中国实验诊断学,2017,21(5):897-900.
[20]吴仲敏,程正文,倪桂莲,等.人参皂苷Rg1对创伤后应激障碍大鼠行为学变化和海马神经元自噬的影响[J].中国病理生理杂志,2017,33(5):896-901.