| ObjectiveCarbon monoxide(Carbon monoxide, CO) is a kind of colourless, odourless and no excitant gas. In daily life, people can be exposed to CO harmful gases,such as iron making, blacksmiths, coking, heat treatment and casting of industrial production and household gas water heaters, stoves and automobile exhaust. Acute CO poisoning is the largest number of acute poisoning in China,and its morbidity and mortality is also biggest. The most serious neurological sequelae in patients with acute CO poisoning is delayed encephalopathy, its pathogenesis has not been completely elucidated.We establish the acute CO poisoning model of mice to observe the performance of poisoning process and detect the different groups of mice brain tissue of MDA,the activities of SOD, GSH-PX and the number of caspase-3in the Id,3d,7d,14d and21d after poisoning.We also observe the pathological changes of poisoning of mice brain tissue and protective effect of EDA in order to further explore the mechanism of delayed encephalopathy after acute CO poisoning, and to provide new ideas for clinical treatment of patients.Methods1. Healthy male Kunming mice were poisonous by CO in the airtight container. After it, they were randomly divided into CO poisoning group and the EDA group (n= 40). Control group was not poisonous mice selected randomly(n=40).EDA group mice were intraperitoneal injection of EDA (0.1ml/10g) respectively in2h and4h after they were awaked, and the remaining two groups of mice received normal saline by intraperitoneal injection.2. Set up the acute CO poisoning model:Observe the behavior of mice which were in the cabinet in the whole poisoning process.3. Mice growth index determination:Weighed all mice after CO poisoning, and weighed again at different time points, and take the whole brain tissue and weigh it. Observe the changes of the different groups of mice at the same time point, calculate weight gain and brain tissue factor.4. Take mice brain tissue and prepare organization homogenate. Using the ultraviolet spectrophotometer to measure the content of MDA, SOD and GSH-PX separately at different time points.5. Morphologic detection:the paraffin section HE dyeing of mice brain tissue was observed under light microscopy and we intuitively determine whether there is the morphological change.6. We use the immunohistochemistry (SP) to measure the changes of the number of caspase-3of different groups of mice brain tissue.Results1. The poisoning behavior of experimental animals:the whole poisonous process of poisoning animals’symptoms are obvious:5-10min, mice move more frequently, and hit the cage;15-20min, the poisonous mice appear less dynamic, spiritual malaise, and become convulsive;30-40min, mice are shortness of breath, almost without moving or even become incontinent, some individual mice appear the coma; About55min entire mice are in the coma.2. The changes of growth index in mice:compared with the control group, the weight gain of mice in CO poisoning group and EDA group was shorten in the21day, there is a significant difference (P<0.05); three groups of mice brain tissue factor was no statistically differences. 3. Changes in the MDA:the MDA content of CO poisoning group at five time points were all significantly increased (P<0.05) compared with the control group; the EDA group was P<0.05at3,7,14days compared with control group.4. Changes in the SOD:compared with the control group, the SOD content of CO poisoning group were marked differences (P<0.01) in addition to the21day; the EDA group was at1,3and7days (P<0.05).5. Changes in the GSH-PX:compared with the control group, the GSH-PX levels of CO poisoning group at five time points were statistically different (P<0.05); there were the obvious differences in the first3,7,14days, P<0.01in EDA group.6. HE staining:there are a large number of neurons cell swelling in the mice brain tissue of CO poisoning group, the cells surrounding appear a large number of gaps, we can see a large number of nerve cells become ischemic degeneration, necrosis, the most significant section are the3d and7d; EDA group is consistent with CO poisoning group, but the damage is a lesser extent.7. The number of caspase-3:the number of caspase-3in acute CO poisoning group mice brain tissue is a significant increase from the1day, compared with the control group at each time point, the number of caspase-3was statistically significant (P<0.05). EDA group, P<0.05, in the1,3,7,14days compared with the control group.Conclusion1. If the mice is about50mins in the airtight container where CO is1550PPM-1650PPM, we can successfully establish the acute poisoning model.2. Acute CO poisoning can cause the changes of free radical system.3. Acute CO poisoning can cause the damage of brain tissue, there are a significant increase in both the number of caspase-3and neuronal apoptosis.4. The mechanism of apoptosis and the mechanism of free radical may be the important pathogenesis of DEACMP.5. The EDA may alleviate the symptoms of CO poisoning and can be used for clinical prevention and treatment of DEACMP. |
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