| Object: To evaluate the pathologic injuries and oxygen free radical contents in lung, liver and kidney after paraquat poisoning in order to determine the injuries action of oxygen free radical induced by paraquat. Methods: The rat pulmonary fibrosis model has been founded by our groups. Male wistar rats were be used in the model, whose body weight were 200±10g, toxic dose 25mg/kg,administration by peritoneal injection, and the incidence rate of pulmonary fibrosis was more than 90%. Simultaneously, two lower and two higher poisoning groups were applied. There were five poisoning groups plus one normal control group for observation. The detailed process : 88 Wistar rats were randomly divided into six groups, including A, B, C, D, E, F, and A was normal control group, eight rats, non-poisoning, and the other five groups were sixteen rats in each, poisoning dosis were divided by 15mg/kg, 20 mg/kg, 25 mg/kg, 30 mg/kg, 35 mg/kg, and eight experimental rats were sacrificed by exsanguination via the abdominal aorta at four hour and twelve hour after poisoning, the groups sacrificed at four hour were B1,C1,D1,E1,F1, and at twelve hour were B2,C2,D2,E2,F2. After sacrificed the tissue of lung, liver and kidney of every rat were taken quickly. The big lobe of left lungs were torrefied at 80℃in the oven and got dry weight, then calculated lung coefficients (including lung-to-body weight ratio, dry lung-to-body weight ratio, water content in the lung and lung wet-to-dry weight ratio) in order to comparing lung coefficients among interclass using t test in SAS statistics software. About 0.5 cm width of superior lobe of right lung was incubated fifteen minutes with 2',7'- dichloro-dihydrofluorescein diacetate (DCFH-DA) and cut frozen sections, then examined fluorescence intensity using fluorescence microscope to determine oxygen free radical quantity. Volume 1mm×1mm×1mm at edge of the fourth right lung was fixed with 3 percentage glutaric dialdehyde solution to check with transmission electron microscope. The second and third lobe of right lung was fixed with 10 percentage neutral formalin solution for HE sections after two days. About 0.5 cm of liver superior lobe was incubated fifteen minutes with DCFH-DA and cut frozen section, then examined fluorescence intensity using fluorescence microscope to determine oxygen free radical quantity. Volume 1mm×1mm×1mm at edge of liver was fixed with 3 percentage glutaric dialdehyde solution to check with transmission electron microscope. Another lobe of liver was fixed with 10 percentage neutral formalin solution for HE sections after two days. Left kidney was intersected middle, half was incubated fifteen minutes with DCFH-DA and cut frozen sections, then examined fluorescence intensity using fluorescence microscope to determine oxygen free radical quantity, the other half was taken 1mm×1mm×1mm at edge and fixed with 3 percentage glutaric dialdehyde solution to check with transmission electron microscope. Another kidney was fixed with 10 percentage neutral formalin solution for HE sections after two days. Results: The pulmonary tissue structure in A group was almost normal. Compared with control group, the lung coefficients of B1, C1, D1, E1, F1, B2, C2 and D2 groups changed nothing, but E2 and F2 group increased obviously, P level less than 0.05. The higher dose of PQ poisoning, the stronger of fluorescence intensity in the tissue(lung, kidney and liver) which were incubated with DCFH-DA, that is higher contents of oxygen free radical. In the alveolar space of B1, C1, D1, E1, F1 groups, there was protein hydropsia effusion and asphyxial membrane under light microscope in HE sections, but leucocytes infiltrated little, the alveolar septum still show monolayer cells, just a little thicker, content of effusion in different poisoning groups present quiet. In B2, C2, D2, E2 and F2 groups, leucocytes infiltrated obviously, and can also view pulmonary hemorrhage,capillary broadening and engorging,endothelial cell swelling, the more poisoning, the more serious of pulmonary alveoli inflammatory infiltration, alveolar spaces in E2 and F2 groups almost disappeared and pulmonary hemorrhage was obvious. In transmission electron microscope, the higher poisoning, the more serious injuries of pulmonary subcellular structure. The hepatic tissue structure in A group was almost normal, and organelle were no abnormalities. After poisoning, pathological changes were based on dropsy, the degree was related to poisoning quantity, the higher poisoning, the stronger fluorescence intensity in liver, that is more content of oxygen free radical, and hepatocytes injury were no difference between 4h and 12h. The nephric tissue structure in A group was nearly normal, and nephric organelle ultra-microstructure were normalities. After poisoning, pathological changes emerged vacuolar degeneration, and were based on dropsy. Frozen sections of kidney which were incubated with DCFH-DA emerged stronger fluorescence intensity at higher poisoning, suggested more content of oxygen free radical, and there were no different nephric injury between 4h and 12h. Conclusion: All of lung, liver and kidney tissues were injuried after PQ poisoning, and the degree of injuries were correlated with quantity of poisoning, the higher poisoning the more serious of tissue injury and more content of oxygen free radical. Before 12 hours, lung injury had obvious time dependence, exsudation was obvious 4 hours after poisoning and inflammatory infiltration was more, however liver and kidney hadn't.
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