| Experiment 1: Effect of FK506 Pretreatment on Endotoxinshock MiceObjective: To study the effect of FK506 pretreatment on endotoxin shock mice induced by LPS. Methods: (1)Established animal model and divided group: Endotoxin shock model was developed by intraperitoneal injection of LPS(25 mg/kg). 155 Kunming mice were random divided into two groups between LPS(n=80) and LPS+FK(n=75). 2 days before experiment, the mice in LPS+FK group were given FK.506 in a dose of 0.5 mg/kg and maintained the same dosage each 12 hours; the LPS group mice were given saline as control. 20 mice of LPS group and 15 of LPS+FK group were chosen to observe the dynamic survival rate change. At 0, 0.5, 2, 4, 6, 8 hours after LPS injection, 10 mice of each group were killed seperately, the blood samples were get and lung and liver were harvested. (2)Monitor index: a. Observe on pathologic respond of mice in two groups in 72 hours after LPS injection,the first death time and death number; draw Kaplan-Meier survival curve and calculate the mid-survival time. b. take the left lung, absorb the purface bloodand liquid quickly and weigh with electronic balance; put into oven(80C) for 48 hours, weigh again and compute the lung wet/dry ratio. Take the right lung and right robe liver, fix with formalin, stain with hematoxylin and eosin, and observe under microscope, c. Place the collected blood under room temperature for a hour, centrifugate at a speed of 2500 r/min for 15 minutes after completely coagulation, and suck out the serum. Estimate sermu ALT, AST levels at 0 h, 8 h after LPS injection with TRACE CB171 semi-automatic biochemistry analyzer; Assay serum TNF-ou IL-1B levels with radio-immunity method. Results: (1)With intervention of FK.506, the survival quality of endotoxin shock mice improved. The mid-survival time lengthened from 16 hours to 54.5 hours, and the survival rate at 72 hours after LPS administration raised from 10% to 46.67%. (2)With intervention of FK506, the lung W/D ratio of endotoxin shock mice decreasd. Histopathologic analysis revealed that FK506 pretreatment reduced the extent of lung tissue lesions, such as intravascular neutrophil agglomerating, perivascular/ peribronchial damage, parenchyma edema and thickening of alveolar wall, ヾecreased with intervention of FK506. Histologically FK506 pretreatment alleviated the acute liver injury induced, including cell swelling, eosinophilic degeneration and PMN infiltration. (4)The serum levels of TNF-a and IL-1B of LPS+FK group mice were lower than that of LPS group mice at each time point. Both TNF-a and IL-1|3 levels of the two groups had a tendency of first increasing and then decreasing. 2 hours after LPS administration, TNF-a level reached its peak, while IL-1B did it two hours later. Conclusion: FK506 pretreatment could decrease the multi-organ lesion induced by LPS and down-regulate the inflammatory reaction. Its mechanism may be ralated to the inhibition of production of TNF-a and IL-1B.Experiment 2: Effect of FK506 Pretreatment on Severe AcutePancreatitis ratsObjective: To study the effect of FK506 pretreatment on severe acute pancreatitis (SAP) rats. Methods: (1)Established animal model and divided group: Severe acute pancreatitis model was developed by retrograde injection of sodium taurocholate solution into the biliopancreatic duct (40 mg/kg). 54 healthy female SD rats were random divided into three groups between SAP(n=18), SAP+FK(n=18) and SO(n=18). 2 days before experiment, the rats in LPS+FK group were given FK506 in a dose of 0.5 mg/kg and maintained the same dosage each 12 hours; the SAP group and SO group rats were given saline as control. At 3, 6, 12 hours after operation, 6 rats of each group were sacrificed seperately, the blood samples were get and lung and pancreas tissues were harvested. (2)Monitor index: a. Observe on survival rate, ascites volume and pathologic changes of lung and pancreas, b. Place the collected blood samples under room temperature for a hour, centrifugate at a speed of 2500 r/min for 15 min... |
