芍药苷对血虚肝郁证候模型大鼠海马CA3区组织形态及一氧化氮通路的影响

Effects of paeoniflorin on histomorphology and NO pathway in hippocampus CA3 Zone in a rat model of blood deficiency and liver depression pattern

目的探讨血虚肝郁证模型大鼠海马CA3区组织形态、谷氨酸(Glu)、一氧化氮(NO)含量、诱导型一氧化氮合酶(i NOS)蛋白表达的变化,以及芍药苷的干预作用。方法 56只雄性SD大鼠采用辐照结合束缚制作血虚肝郁证大鼠模型,随机分为模型组,芍药苷高、中、低剂量组,另设正常SD大鼠为空白组,每组14只。造模第1天开始芍药苷高、中、低剂量组每天分别灌胃40、20、10mg/kg的药物,模型组和空白组灌胃等量的纯净水,共21 d。采用大脑切片HE染色,观察病理形态学改变;邻苯二甲醛(OPA)柱前衍生HPLC荧光检测法测定海马中Glu含量;格里斯法(Griess法)测定NO含量;免疫组化法测定海马CA3区i NOS蛋白的表达。结果与空白组比较,模型组海马CA3区神经元胞体收缩、染色质加深、细胞核深染、核收缩呈三角形、核仁不明显;海马组织中Glu、NO含量显著升高(P<0.05),i NOS蛋白表达显著增多(P<0.05)。与模型组比较,芍药苷组海马CA3区神经元细胞核未见明显异常,神经元形态基本完整;芍药苷高剂量组海马组织中Glu含量显著降低(P<0.05),芍药苷高、中剂量组海马组织中NO含量显著降低(P<0.05),i NOS蛋白表达显著降低。结论血虚肝郁证模型大鼠海马CA3区神经元出现萎缩等病理状态,Glu、NO含量升高,i NOS蛋白表达增多,而芍药苷对海马组织的病理变化具有改善作用,其发挥神经保护的功能可能是通过降低Glu和NO含量,降低i NOS蛋白的表达等多途径实现的。

Objective To investigate the effects of paeoniflorin on histomorphology,Glu and NO levels in hippocampus,and on protein expressions of iNOS in the hippocampus CA3 Zone in a rat model of blood deficiency and liver depression pattern. Methods The rat model of blood deficiency and liver depression pattern was established with chronic restraint stress in conjunction with radiation. Fifty-six male SD rats were randomly divided into model group, high-dose, medium-dose, and low-dose paeoniflorin groups,as well as the blank control group of healthy SD rats（ n = 14 in each group）. SD rats were treated with intragastric administration of paeoniflorin in the high-dose,medium-dose,and low-dose paeoniflorin groups while pure water was intragastrically administered in the model and blank groups for21 days. Histopathological changes of hippocampus CA3 Zone were observed by using HE staining methods,and Glu and NO levels in hippocampus tissues was measured by using HPLC and the Griess methods,respectively,and protein expression of iNOS in hippocampus CA3 zone was detected with immunohistochemistry. Results Compared with the blank group,significant changes were observed in the neuron cells in the model group,including cell body shrinkage,deeply stained chromatin nuclei,triangular nuclear shrinkage and no remarkable nucleoli. There was no significant changes in paeoniflorin group. The levels of Glu and NO increased significantly in model group compared with normal group（ P〈0. 05）,and decreased significantly in paeoniflorin group compared with model group（ P〈0. 05）. The protein expression of iNOS in hippocampus CA3 zone increased significantly in the model group compared with the blank control group（ P〈0. 05）,and decreased significantly in paeoniflorin group compared with the model group（ P〈0. 05）. Conclusion Pathological changes including neuron shrinkage were present in hippocampus CA3 Zone in the model of blood deficiency and liver depression pattern.Paeoniflorin could improve hippocampus functional structural damage in the rat model of blood deficiency and liver depression pattern,which may be related to decreased levels of Glu and NO and protein expression of iNOS in hippocampus CA3 Zone.