水溶性灵芝多糖对大鼠脑缺血损伤的神经保护作用。

Neuroprotective effects of water-soluble Ganoderma lucidum polysaccharides on cerebral ischemic injury in rats.

机构信息

Laboratory of Neurology, Institute of Integrative Medicine, Zhongshan Hospital, Fudan University, Shanghai 200032, China.

出版信息

J Ethnopharmacol. 2010 Aug 19;131(1):154-64. doi: 10.1016/j.jep.2010.06.023. Epub 2010 Jun 18.

DOI:10.1016/j.jep.2010.06.023

PMID:20600765

Abstract

AIM OF THE STUDY

To investigate the neuroprotective effects of water-soluble Ganoderma lucidum polysaccharides (GLPS) on cerebral ischemic injury in rats, and to explore the involved mechanisms.

MATERIALS AND METHODS

Two models [middle cerebral artery occlusion (MCAO) in Sprague-Dawley (SD) rats and oxygen and glucose deprivation (OGD) in primary cultured rat cortical neurons] were employed to mimic ischemia-reperfusion (I/R) damage, in vivo and in vitro, respectively. Cerebral infarct area was measured by tetrazolium staining, and neurological functional deficits were assessed at 24h after I/R. Neuronal apoptosis was studied by Nissl staining and DNA fragmentation assay. Neuronal injury was assessed by morphological examination using phase-contrast microscopy and quantified by measuring the amount of lactate dehydrogenase (LDH) leakage, cell viability was measured by sodium 3'-1- (phenylaminocarbonyl)-3, 4-tetrazolium-bis (4-methoxy-6-nitro) benzene sulfonic acid (XTT) reduction. Neuronal apoptosis was determined by flow cytometry, and electron microscopy was used to study morphological changes of neurons. Caspase-3, -8 and -9 activation and Bcl-2, Bax protein expression were determined by western blot analysis.

RESULTS

Oral administration of GLPS (100, 200 and 400mg/kg) significantly reduced cerebral infarct area, attenuated neurological functional deficits, and reduced neuronal apoptosis in ischemic cortex. In OGD model, GLSP (0.1, 1 and 10 microg/ml) effectively reduced neuronal cell death and relieved cell injury. Moreover, GLPS decreased the percentage of apoptotic neurons, relieved neuronal morphological damage, suppressed overexpression of active caspases-3, -8 and -9 and Bax, and inhibited the reduction of Bcl-2 expression.

CONCLUSIONS

Our findings indicate that GLPS protects against cerebral ischemic injury by inhibiting apoptosis by downregulating caspase-3 activation and modulating the Bcl-2/Bax ratio.

摘要

目的

研究水溶性灵芝多糖(GLPS)对大鼠脑缺血损伤的神经保护作用，并探讨其作用机制。

材料与方法

采用大脑中动脉阻塞（MCAO）模型和原代培养大鼠皮质神经元氧糖剥夺（OGD）模型，分别在体内和体外模拟缺血再灌注（I/R）损伤。通过四唑盐染色测量脑梗死面积，在 I/R 后 24 小时评估神经功能缺损。通过尼氏染色和 DNA 片段化测定研究神经元凋亡。通过相差显微镜观察形态学改变评估神经元损伤，并通过测量乳酸脱氢酶（LDH）漏出量来量化细胞活力，通过 XTT 还原法评估细胞活力。通过流式细胞术测定神经元凋亡，电子显微镜观察神经元形态变化。通过 Western blot 分析测定 caspase-3、-8 和 -9 的激活以及 Bcl-2、Bax 蛋白的表达。

结果

GLPS（100、200 和 400mg/kg）口服可显著减少脑梗死面积，减轻缺血皮质区的神经功能缺损，减少神经元凋亡。在 OGD 模型中，GLPS（0.1、1 和 10μg/ml）可有效减少神经元细胞死亡，减轻细胞损伤。此外，GLPS 降低了凋亡神经元的比例，减轻了神经元形态损伤，抑制了活性 caspase-3、-8 和 -9 以及 Bax 的过表达，并抑制了 Bcl-2 表达的减少。