| Endotoxin produced by Intestinal Obstruction often cause damages of distal organs in neonatal period, and acute lung injury (ALI) is the most common injury Among them. The clinical manifestations is hypoxia, moist rales, cough pink frothy sputum and progressive dyspnea, some serious paeients can develop into acute respiratory distress syndrome, and then Trigger multiple organ dysfunction syndrome, The mortality rate of it is more than 44%, so prevention ALI has a very important clinical significance.ObjectiveThis study intends to adopt neonatal rats acute lung injury model by intraperitoneal infection of lipopolysaccharide, through the preoperative give penehyclindine hydrochloride intervention, to observed the lung pathological changes by light-microscopy, and detective the change of Cyt-c^ Caspase-3,MDA,SOD, discusses the protective mechanism of penehyclindine hydrochloride on acute lung injury in neonatal rats. Materials and Methods1 Study ObjectForty 7 days old Wistar rats weighing between 12 to 18g provided By the laboratory animal center provides of henan province.2 Methods2.1 animals modelsA neonate rat model of ALI was induced by intraperitoneal infection of lipopolysaccharide(LPS 5mg·kg). The animal were fixed on the operating table, and given LPS injection intraperitoneally (5mg·kg) With 1 ml syringe.Then put The rats in cage, move freely,not banning diet and water.established A success neonate rat model of acute lung injury Based on optical microscope morphologic changes in the structure of lung tissue2.2 Animal groupForty 7-day-old Wistar rats weighing between 12 to 18g were randomly divided into 4 groups (n=10 each):control group(group NS), which was given saline injection intraperitoneally; ALI group(group LPS), which was given LPS injection intraperitoneally (5mg·kg) to reproduce the ALI neonatal rats models; low dose group (group PL), which was given intraperitoneal injection of PHC 0.1mg/kg, After 30 minutes, they were induced by intraperitoneal infection of LPS (5mg·kg); high dosegroup (group PH), which was given intraperitoneal injection of PHC 1.0 mg/kg, After 30 minutes, they were induced by intraperitoneal infection of LPS (5mg·kg). After 6 hours,the pups were anesthesia with ether, take off the lung quickly, then fixed the lung with 4% formaldehyde solution.2.3 Collection and disposal of samples2.3.1 Collection and disposal of blood samplesAfter 6 hours of operation, all rats were anaesthetized by aether and Supine fixed on the operating table. Blood were extracted from Right ventricular quickly after Conventional disinfected with iodine,alcohol. Specimen of blood rest for 20 minutes, centrifuge at room temperature (3000 r·min, 10min),serum was stored at -20℃.2.3.2 Collection and disposal of Lung samplesAfter Blood were extracted from Right ventricular, physiological saline Flush, Remove lung quickly, fixed left lung in 4% formaldehyde 24 hours, Paraffin embedded the lung and cut into slices in 4μm thickness 5 times,then HE dyed, the lung pathologic structure were observed under the optical microscope electron microscope. Remove right lung quickly with same methode,and fixed, dehydration, encasement, cut sheet wait for Cyt-c,Caspase-3 test.2.4 Detection Indexes2.4.1 The histology of lungParaffin embedded the lung and cut into slices in 4μm thickness 5 times,then HE dyed, the lung pathologic structure were observed under the optical microscope electron microscope.2.4.2 Serum IndexesThe expression of SOD were detected by xanthine oxidase method; The expression of MDA in the were detected by sulfuretted barbitone method.2.4.3 Apoptosis proteins in lungLevels of Cyt-c and Caspase-3 in lung tissues were determined with immunohistochemical method (IHC).3 Statistical treatmentSPSS 12.0 was used for studies.All data were calculated using mean±SD. comparing data by one-way ANOVA within the group, Mean differences between binary comparison using q-test. There was a statistical significance when P<0.05.Results1. The general status observation:The neonate rats of LPS group had series of symptoms such as scrunch, Surface wet,rarely exercise, tachypnea and respiratory embarrassment,some rats had cyanosis after 4 hours. The PL group and PH group both had symptoms such as rarely exercise, tachypnea. The PL group also had symptoms such as respiratory embarrassment, cyanosis,but less than the LPS group. The NS do not had such symptoms.2.By microscopy,each group had classic change presenting acute lung injury which included, the infiltration of inflammatory cells, alveolar septum widen,the leakage of erythrocyte except group NS,which structure of alveoli and interalveolar septum was normal,the alveolar space was clear.Group LPS had most severe pathologic changes.In the group PH, the according pathological changes were slighter than group PL.3. There was significant difference on expression of Cyt-c and Caspase-3 in lung of neonatal rats among four groups.Compared with group NS,the expression of Cyt-c and Caspase-3 of lung tissues in the group LPS increased obviously(P<0.05). Compared with LPS group, the level of Cyt-c and Caspase-3 of lung tissues in the group PL and PH decreased(P<0.05). Furthermore,group PH was lower than that of group PL.4. Compared with group NS, the Content of MDA were higher in group LPS,PL and PH (P<0.05), While the Content of SOD were lower (P<0.05) Compared with group LPS, the Content of MDA were lower in group PL and PH(P<0.05), While the Content of SOD were higher (P<0.05),groups PH more significant changes.ConclusionsPHC has the protective effect on ALI.The mechanisms of the effect are due to its ability of inhibiting the expression of Cyt-c protein,Caspase-3 protein in lung, and lighting the expression of MDA and improving the expression of SOD.thereby inhibit oxidatie stress and mitochondrial mediated apoptosis at the development of ALI... |
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