Wang Qiao-Yun, Liu Feng, Wu Feng-Jie, Li Jin-Lian
College of Basic Medicine, Binzhou Medical College, Shandong (264003), China.
Zhongguo Zhong Xi Yi Jie He Za Zhi. 2013 Feb;33(2):229-34.
To study the possible anti-apoptotic mechanism of ginsenoside Rg1 on the apoptosis of hippocampal neuron after cerebral ischemia/reperfusion (I/R) injury rats.
Totally 120 healthy male adult SD rats were randomly divided into the cerebral I/R model group (the model group), the low dose ginsenoside Rg1 group (10 mg/kg), the middle dose ginsenoside Rg1 group (20 mg/kg), the high dose ginsenoside Rg1 group (40 mg/kg), and the sham-operation group, 18 in each group. Rats received medication by peritoneal injection. Equal volume of normal saline was peritoneally injected to rats in the sham-operation group and the model group, once daily, for 7 successive days. The cerebral I/R injury model was prepared by 2-h middle cerebral artery occlusion (MCAO) followed by 24-h reperfusion. Rats in the sham-operation group received the same surgical procedure without the carotid arteries occluded. The neurofunction was assessed using Longa EZ method. The injury of hippocampal pyramidal cells was observed by Nissel staining and TUNEL assay. The nerve cell apoptosis rate was calculated. The protein expression levels of extracellular signal-regulated kinase 1/2 (ERK1/2), phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2), c-Jun N-terminal kinases (JNK), and phosphorylated c-Jun N-terminal kinase (p-JNK) were detected using Western blot.
Compared with the sham-operation group, the score of neurofunction, the apoptosis rate, the expression levels of p-JNK and p-ERK1/2 increased, the survived number of pyramidal cells decreased in the model group (P < 0.05, P < 0.01). Compared with the model group, the score of neurofunction and the apoptosis rate decreased in each ginsenoside Rg1 group (P < 0.05, P < 0.01). The survived number of pyramidal cells increased in the high and middle dose ginsenoside Rg1 groups, the expression of p-JNK in the hippocampal CA1 region decreased, and the expression level of p-ERK1/2 increased (P < 0.05, P < 0.01). Compared with the low dose ginsenoside Rg1 group, the score of neurofunction, the apoptosis rate, the p-JNK protein expression decreased, the survived number of pyramidal cells increased, the expression of p-ERK1/2 increased in the high and middle dose ginsenoside Rg1 groups (P < 0.05, P < 0.01). Three to four layers of pyramidal cells were arranged tightly and compactly in the hippocampal CA1 region of the sham - operation group. The nucleus was big and round under high power lens, with 1 -2 kernel. After cerebral I/R injury, the hippocampal nerve cells were severely injured. Normal structure was lost in the CA1 region, with disarranged cell line and reduced cell amount. Partial neurons were shrunken, and the kernel was condensed and darkenedly stained. They were in triangular, long strip, fusiform, or irregular shape. The staining of nucleus was clustered and the kernel was not clear. Ginsenoside Rg1 (20 and 40 mg/kg) could improve the morphology of ischemic nerve cells, reduce their loss. Of them, stronger effects were shown in the high dose ginsenoside Rg1 group than in the middle dose ginsenoside Rg1 group. The JNK protein band was divided into two subzones, JNK1 (46 kD) and JNK2 (54 kD). ERK protein band was also divided into two subzones, ERK1 (44 kD) and ERK2 (42 kD).
The protective effect of ginsenoside Rg1 on cerebral I/R injury was correlated with inhibiting the apoptosis of hippocampal neurons, regulating the expression levels of p-ERK1/2 and p-JNK.
研究人参皂苷Rg1对脑缺血/再灌注（I/R）损伤大鼠海马神经元凋亡的可能抗凋亡机制。
将120只健康成年雄性SD大鼠随机分为脑I/R模型组（模型组）、低剂量人参皂苷Rg1组（10 mg/kg）、中剂量人参皂苷Rg1组（20 mg/kg）、高剂量人参皂苷Rg1组（40 mg/kg）和假手术组，每组18只。大鼠通过腹腔注射给药。假手术组和模型组大鼠腹腔注射等体积的生理盐水，每日1次，连续7天。采用大脑中动脉闭塞2小时后再灌注24小时的方法制备脑I/R损伤模型。假手术组大鼠接受相同手术操作，但不结扎颈动脉。采用Longa EZ法评估神经功能。通过尼氏染色和TUNEL法观察海马锥体细胞损伤情况。计算神经细胞凋亡率。采用蛋白质印迹法检测细胞外信号调节激酶1/2（ERK1/2）、磷酸化细胞外信号调节激酶1/2（p-ERK1/2）、c-Jun氨基末端激酶（JNK）和磷酸化c-Jun氨基末端激酶（p-JNK）的蛋白表达水平。
与假手术组相比，模型组神经功能评分、凋亡率、p-JNK和p-ERK1/2表达水平升高，锥体细胞存活数量减少（P<0.05，P<0.01）。与模型组相比，各人参皂苷Rg1组神经功能评分和凋亡率降低（P<0.05，P<0.01）。高、中剂量人参皂苷Rg1组锥体细胞存活数量增加，海马CA1区p-JNK表达降低，p-ERK1/2表达水平升高（P<0.05，P<0.01）。与低剂量人参皂苷Rg1组相比，高、中剂量人参皂苷Rg1组神经功能评分、凋亡率、p-JNK蛋白表达降低，锥体细胞存活数量增加，p-ERK1/2表达升高（P<0.05，P<0.01）。假手术组海马CA1区锥体细胞排列紧密，3～4层。高倍镜下细胞核大而圆，有1～2个核仁。脑I/R损伤后，海马神经细胞损伤严重，CA1区正常结构消失，细胞排列紊乱，细胞数量减少。部分神经元皱缩，核浓缩，染色加深，呈三角形、长条形、梭形或不规则形，核染色聚集，核仁不清。人参皂苷Rg1（20和40 mg/kg）可改善缺血神经细胞形态，减少其丢失。其中，高剂量人参皂苷Rg1组效果强于中剂量人参皂苷Rg1组。JNK蛋白条带分为两个亚区，JNK1（46 kD）和JNK2（54 kD）。ERK蛋白条带也分为两个亚区，ERK1（44 kD）和ERK2（42 kD）。
人参皂苷Rg1对脑I/R损伤的保护作用与抑制海马神经元凋亡、调节p-ERK1/2和p-JNK表达水平有关。
