"poison Damage Brain Network Disease Hypothesis Of The Cell Biology Of Basic Research

Ischemic cerebral vascular disease is a major lethal disease, has the characteristic of high incidence rate, high crippling rate, high recurring rate. In recent years, through the repeatedly clinical observation and the practice, it is thought that this disease belong to the category of'collateral disease'in Chinese medicine,'toxin hurts brain collaterals'is the key point in its pathogenesis, and remarkable clinical curative effect has obtained with the treatment targeting on the functional characteristic of collateral. Also, it is believed that the the injury of brain vascular endothelial cell is the early pathological change and the basic agent of the attack of cerebral ischemia, and has close relationship with the development of cerebral vascular diseases. Therefore, under the instruction of'collateral theory'of Chinese medicine and focus the target on the Brain Microvascular Endothelial Cells (MVEC), we further studied the regulatory influence of rat brain microvascular endothelial cells on the activity of cortical neurons.Objective :We collected the conditioned media from 4 different cultured cerebral endothelial cell groups, which were normal endothelial cells, the normal ones and treated with Tong Luo Jiu Nao Injection, the injured ones damaged by simulated cerebral ischemia, as well as the injured ones and treated with Tong Luo Jiu Nao Injection, respectively(N-CM, NT-CM, I-CM, IT-CM). Then, the conditioned medium was added into the cultures of the normal neurons and the damage ones which are injured by simulated cerebral ischemia as well, respectively. The effects of each type of conditioned medium on the activities of neurons and nomadic calcium ion concentration and the NMDA receptor as well as the mitochondrial function were determined, and carried on the analysis of difference protein in conditioned media from different cultured endothelial cells using proteomics techniques to discuss the material basis of regulatory function of vascular endothelial cells on neurons. Thus presented the modern biology basis of'toxin hurts brain collaterals'pathogenesis theory, and provided significant ideas for the pathway and target of treatment of central nervous system disease with Chinese Materia Medica.Methods:1. Primary culture and identification of rat brain MVEC: Get the brain MVEC using the methods of homogenating and twice filtering as well as collagenase digesting, then cultivate , purify , passage and identify it with immunofluorescence labelling of vWF.2.Establishment of model of the brain MVEC damaged by simulated cerebral ischemia: Using method of cell oxygen deficit combined with culture fluid absent of sugar, observed the activity change of MVEC in different time after injury, thus established a reliable experimental model of MVEC damaged by simulated cerebral ischemia in vitro.3. Screening the application density of Tong Luo Jiu Nao Injection: Observe the influence of...