| Acute pulmonary embolism is a group of the disease that various embolus obstructed pulmonary artery system as a cause. It has been concerned because of its morbidity and mortality increased year by year. At present, thrombolytic therapy is the method recommended by Clinic doctors, but the reduction of mortality and sudden death rate wasn’t as pronounced after thrombolytic therapy. The reasons of this phenomenon may closely connected to lung ischemia-reperfusion injury appeared at Earlier thrombolytic therapy. The reasons of forming lung ischemia-reperfusion injury are manifold, such as ischemia basis caused by APE and restores Blood flow might accelerate lung injury. Endothelin is the materials which can contraction of the smooth muscle of vascular and air passages, especially effect the small vessel (resistance renal blood vessels) of lungs. So endothelin may be promote lung ischemia-reperfusion injury appeared. In recent years, Great progress has been made in the research of antagonist-sing the effect of endothelin against lung ischemia-reperfusion injury.BQ-123is the ET-1receptor antagonists, and could competitive inhibitor of ET-1receptor.We established the rabbit model of acute pulmonary embolism, when thrombolytic we used endothelin-1receptor antagonist BQ-123intervention. We measured the mean pulmonary arterial pressure before remobilization and0.5h,2h,4h after embolization and the levels of ET-1in the venous plasma concentrations. We also measured the ratio of lung tissue wet weight/dry weight, the SOD activity and MAD content at the end of the experiment. The research was divided into three parts as the followed.Objective:To establish the model of acute pulmonary embolism in rabbit and observe the effects of embolism.Methods:1. The animal groups:12Big-eared Japanese rabbits(SPF,no limit of male or female)are divided into2groups randomly (six in each group), that is SHAM group (sham-operated group), PE group (pulmonary embolism group).SHAM groups: Required tracheotomy and ventilation, the mean pulmonary artery pressure was measured before0.3ml/kg0.9%NS injected the pulmonary slowly through trocar. PE groups:The mean pulmonary artery pressure was measured before autologous blood clots0.3ml/kg injected the pulmonary artery slowly through trocar. The other operation is the same as the SHAM group.2. Animal model:The10%chloralic hydras4ml/kg intraperitoneal injection, we tracheotomy and ventilation with DH-150type small animal breathing machine when succeed at anesthesia, breathing rate50/min, tidal volume set up to10ml/kg, the vital signs was monitor by American HP M3046A electro card-scope monitor;20G trocar was connected with bar-receptor after inserting the main pulmonary artery root through thoracotomy; The trocar were also inserted in femoral vein and femoral artery;2ml arterial blood put in the disposable sterilized syringe, stand for natural solidification, mothball; The arteria cruralis of right used as ambulatory blood pressure monitoring and femoral venous of left used as intravenous drug.3. The marker for the detection:(1) Mean pulmonary artery pressure(MPAP): Record the different time points of MPAP such as before embolization and0.5h,2h,4h after embolization.(2) The ratio of lung tissue wet weight/dry weight:Executed rabbits after embolization for4h to observe the thromboembolism, and take about1g lung tissue, measured the wet weight with electronic analytical balance and dry weight after baking at80℃in the electric thermostatic drier over48h, calculated the wet weight/dry weight (W/D).(3) The ET-1of plasma concentration:Draw2ml blood from femoral venous into EDTA tube before embolization and at0.5h2h,4h after embolization,3000r/min centrifugal15min at the place of4℃, detected ET-1of plasma concentration by radioimmunoassay.(4) Determination the lung tissue Superoxide Dismutase (SOD) activity and Malondialdehyde content:Mixed the lung tissue and0.9%NS then homogenate the lung,3000r/min centrifugal15min at the place of4℃。 Measured the concentration changes of MDA and SOD level by colorimetric method, take on the operation strictly according to the operation instructions.(5) looked at the lung tissues under an electron microscope.The statistical description through Mean±SD(x±s), and comparisons between groups were analyze with one-way ANOVA test. The comparisons between pair-wise with the LSD test. P<0.05is considered significant.Results1. Mean pulmonary artery pressure:Analyze with one-way ANOVA test we found there were no difference between groups before embolization(F=1.125, P=0.314). But there were significant difference between different groups at different time point after embolization(F0.5h=109.641、F2h=262.091、F4h=271.936, P=0.000). Analyze with LSD test we found when compared with the SHAM group, the MPAP of PE group increased more significantly when embolization after0.5h (P<0.05). Researches indicate that MPAP rise obviously caused by pulmonary vessel were obstructed in the APE.2. The ratio of lung tissue wet weight/dry weight:Analyze with one-way ANOVA test we found there were difference between PE and SHAM groups(F=61.866, P=0.000). Analyze with LSD test we found when compared with SHAM group, W/D increased obviously in PE group (P<0.01). Researches indicate that pulmonary edema occurred in APE.5. There was no thrombosis in SHAM group when we anatomized lung, but the thrombosis can be found in the pulmonary artery system in PE group. We can found that the structure of mitochondria and epithelial cells are normal in SHAM group. Mitochondria dissolved into vacuole in PE group under an electron microscope.Conclusion1. Control the dosage of blood clot within0.3ml/kg injected to the pulmonary artery slowly can establish the model of acute pulmonary embolism in rabbit.2. The change of MPAP of PE group, ET-1of plasma concentration, The ratio of lung tissue wet weight/dry weight, SOD concentration and the MDA content in lung tissue indicate that acute injury of lung.Objective:To observe lung ischemia-reperfusion injury after thrombolytic in the APE.Methods:1. The animal groups:18Big-eared Japanese rabbits(SPF,no limit of male or female)are divided into3groups randomly (six in each group), that is SHAM group (sham-operated group), PE group (pulmonary embolism group), UK group (thrombolytic group).2. The methods of Establish Animal model:SHAM groups:Required tracheotomy and ventilation, the mean pulmonary artery pressure was measured before0.3ml/kg0.9%NS injected the pulmonary slowly through trocar. PE groups: The mean pulmonary artery pressure was measured before autologous blood clots0.3ml/kg injected the pulmonary artery slowly through trocar. UK group:Inject heparin and the urokinase20000U/kg through trocar in the pulmonary artery slowly with micro perfusion pump at0.5h after embolism, all the urokinase should be injected within2h. The other operation is the same as the part1.3. The marker for the detection:(1) Mean pulmonary artery pressure(MPAP): Record the different time points of MPAP such as before embolization and0.5h,2h,4h after embolization.(2) The ratio of lung tissue wet weight/dry weight:Executed rabbits after embolization for4h to observe the thromboembolism, and take about1g lung tissue, measured the wet weight with electronic analytical balance and dry weight after baking at80℃in the electric thermostatic drier over48h, calculated the wet weight/dry weight (W/D).(3) The ET-1of plasma concentration:Draw2ml blood from femoral venous into EDTA tube before embolization and at0.5h2h,4h after embolization,3000r/min centrifugal15min at the place of4℃, detected ET-1of plasma concentration by radioimmunoassay.(4) Determination the lung tissue Superoxide Dismutase (SOD) activity and Malondialdehyde content:Mixed the lung tissue and0.9%NS then homogenate the lung,3000r/min centrifugal15min at the place of4℃Measured the concentration changes of MDA and SOD level by colorimetric method, take on the operation strictly according to the operation instructions.(5) Looked at the lung tissues under an electron microscope. The statistical description through Mean±SD(x±s), and comparisons between groups were analyze with one-way ANOVA test. The comparisons between pair-wise with the LSD test. P<0.05is considered significant.Results1. Mean pulmonary artery pressure (MPAP):Analyze with one-way ANOVA test we found there were no difference between groups before embolization(F=1.400, P=0.277); There were no difference between PE group and UK group. The MPAP in UK group is lower than PE group at2h(F=126.206, P=0.000),4h(F=141.075, P=0.000) after embolization. But analyze with LSD test we found when compared with the SHAM group, the MPAP increased more significantly in PE group and UK group at0.5h after embolization (P<0.01),2. The ET-1of plasma concentration:Analyze with one-way ANOVA test we found the ET-1of plasma concentration have no difference between the three groups before embolization(F=0.084,P=0.920). But there were some difference in the ET-1of plasma concentration in SHAM group(Fo.5h=30.611、F2h=197.901、F4h=213.732, P均=0.000), and analyze with LSD test we found UK group are higher than the PE group at different time points after embolization(P=0.003); The ET-1of plasma concentration have no difference between PE group and UK group at0.5h after embolization (P=0.272).3. The SOD concentration and the MDA content in lung tissue:Analyze with one-way ANOVA test we found the SOD concentration(F=31.152, P=0.000)and the MDA content(F=37.897, P=0.000) in lung tissue have some difference between the three groups. Analyze with LSD test we found when compared with SHAM group, the MDA content increased significantly and the SOD concentration decreased in PE group and UK group(P<0.01). Compared with PE group, the MDA content increased significantly and the SOD concentration decreased in UK group(P<0.01).4. The wet/dry (W/D) of lung tissue:Analyze with one-way ANOVA test we found the W/D of lung tissue have some difference between the three groups(F=105.246, P=0.000). Analyze with LSD test we found when compared with SHAM group, W/D increased obviously in PE group and UK group (P<0.01). The W/D increased in UK group than PE group.5. A lot of lamellar body formatted in lung cell in the UK group under an electron microscope.Conclusion We found lung injury obviously in acute pulmonary embolism from our research; and the injury aggravated with thrombolytic, so called ischemia-reperfusion lung injury.Objective:To explore the protective effects of endothelin-1receptor antagonist BQ-123against ischemia-reperfusion injury after thrombolytic in acute pulmonary embolism.Method:1.24Big-eared Japanese rabbits(SPF,no limit of male or female)are divided into4groups randomly (six in each group), that is SHAM group (sham-operated group), PE group (pulmonary embolism group), UK group (thrombolytic group), BQ group (UK+BQ-123group)。 The SHAM group, PE group, UK group were operatied as the part1and part2, the UK+BQ group:Inject heparin and the urokinase20000U/kg through trocar in the pulmonary artery slowly with micro perfusion pump at0.5h after embolism, all the urokinase should be injected within2h. and at the same time inject the BQ-123(50μg/kg), The other operation is the same as the part2.2. The marker for the detection:(1) Mean pulmonary artery pressure(MPAP): Record the different time points of MPAP such as before embolization and0.5h,2h,4h after embolization.(2) The ratio of lung tissue wet weight/dry weight:Executed rabbits after embolization for4h to observe the thromboembolism, and take about1g lung tissue, measured the wet weight with electronic analytical balance and dry weight after baking at80℃in the electric thermostatic drier over48h, calculated the wet weight/dry weight (W/D).(3) The ET-1of plasma concentration:Draw2ml blood from femoral venous into EDTA tube before embolization and at0.5h2h,4h after embolization,3000r/min centrifugal15min at the place of4℃, detected ET-1of plasma concentration by radioimmunoassay.(4) Determination the lung tissue Superoxide Dismutase (SOD) activity and Malondialdehyde content:Mixed the lung tissue and0.9%NS then homogenate the lung,3000r/min centrifugal15min at the place of4℃Measured the concentration changes of MDA and SOD level by colorimetric method, take on the operation strictly according to the operation instructions.(5) Looked at the lung tissues under an electron microscope. The statistical description through Mean±SD(x±s), and comparisons between groups were analyze with one-way ANOVA test. The comparisons between pair-wise with the LSD test. P<0.05is considered significant.Results:1.Analyze with one-way ANOVA test we found the W/D of lung tissue have some difference in the four groups before embolization(F=1.382, P=0.277). Analyze with LSD test we found when compared with the SHAM group, the PE group, UK group, and BQ group increased significantly at0.5h after embolization (P<0.01);2. Analyze with one-way ANOVA test we found the levels of ET-1have difference in different groups at different time points(F0.5h=25.553、F2h=148.401、 F4h=161.716, P=0.000). Analyze with LSD test we found the levels of ET-1in the plasma of BQ+UK group and UK group have no difference at2h after embolization(P=0.346).3. Analyze with one-way ANOVA test we found the MAD content(F=36.409, P=0.000), SOD activity(F=19.489,P=0.000), the W/D (F=86.343, P=0.000)have difference in different groups. Analyze with LSD test we found when compared with the SHAM group, the W/D and MAD content increased but SOD activity reduced in PE group and UK group at the end of the experiment(P<0.01), UK group compared with PE is have the same result (P<0.01). the W/D(P=0.029)and MAD content is lower in BQ+UK group than UK group, but the SOD activity is more higher(P<0.01). A few of mitochondria dissolved into vacuole in the UK+BQ under an electron microscope.Conclusions:Lung ischemia-reperfusion injury appeared in thrombolytic when we curried the APE, and BQ-123maybe antagonizes that reaction. |
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