壳寡糖硬脂酸嫁接物纳米材料对小鼠肺部毒性研究
发布时间：2018-08-27 07:15
【摘要】：目的 壳寡糖硬脂酸嫁接物纳米材料(chitosan oligosaccharide-stearic acid, CSO-SA)是由浙江大学药学院合成的纳米靶向载药物质。我们在前期研究中发现小鼠尾静脉注射CSO-SA后,会发生呼吸窘迫,表现为呼吸局促,频次变多,呼吸深度变浅。死亡小鼠解剖时发现肺脏颜色偏红,部分肺叶有出血点、水肿等现象。本研究的目的是探索CSO-SA导致小鼠急性肺损伤的可能机制。 方法 开展3个体内实验：血气分析实验、单次给药急性毒性实验、重复给药肺部毒性实验。血气分析实验设置阴性对照组(NaCl),低、中、高剂量组(25、50、125mg/kg),阳性对照组(脂多糖)。给药1h后颈动脉采血,分析血液中pH值、动脉氧分压、二氧化碳分压和血氧饱和度等指标。单次给药急性毒性实验设置阴性对照组,低、中、高剂量组(25、50、75mg/kg),阳性对照组。给药后24小时进行肺泡灌洗。灌洗液测定白细胞数量、肺蛋白含量、超氧化物歧化酶(SOD)、乳酸脱氢酶(LDH)、碱性磷酸酶(AKP)活性变化,肺组织匀浆测定丙二醛(MDA)含量变化。重复给药肺部毒性实验设置阴性对照组,4个实验组(5、10、25、50mg/kg),阳性对照组。连续5天给药,最后一次给药后24小时股动脉放血处死小鼠。检测灌洗液中白细胞数量、蛋白含量,肺组织匀浆中SOD、LDH、AKP活性、MDA含量变化；计算肺脏体系数,组织匀浆检测炎症因子白介素1p(IL-1β)、肿瘤坏死因子(TNF-a)的变化和肺表面活性相关蛋白(SP-A)表达量变化,HE染色和电镜染色观察肺组织损伤情况。SPSS18.0对数据进行统计分析,P0.05认为有统计学差异。 结果 给药后125mg/kg组血液呈现酸性,pH值与阴性对照相比有显著性差异(P0.05)。重复给药后25、50mg/kg组体重增长减少,肺脏体系数升高,与阴性对照组存在显著性差异(P0.05)。单次及连续5天给药后,各剂量组都能引起灌洗液中总细胞计数上升,差异有显著性(p0.05)。75mg/kg组作用24小时和50mg/kg组连续5天给药肺泡灌洗液蛋白含量增加,与阴性对照相比差异有显著性(p0.05)。重复给药25、50mg/kg引起肺组织匀浆LDH升高,10、25、50mg/kg引起AKP升高,与阴性对照相比差异有显著性(p0.05)。重复给药各剂量组中IL-1β水平与阴性对照相比升高,差异有显著性(p0.05),而5、25、50mg/kg均引起了TNF-α水平升高,差异有显著性(p0.05)。各组SP-A表达水平与阴性对照相比减少,其中10、25、50mg/kg组存在显著性差异(P0.05)。HE染色显示50mg/kg的肺泡结构受到破坏。电镜显示50mg/kg组的肺泡上皮细胞凋亡比正常组增多,Ⅱ型上皮细胞特有的板层体结构空泡化增多。 结论 1.小鼠单次或重复尾静脉注射CSO-SA后,主要靶器官为肺,能引起肺部炎症和出血,这可能是小鼠急性死亡的主要原因。 2.反复尾静脉注射CSO-SA,能诱导小鼠肺部细胞凋亡,破坏肺泡Ⅱ型上皮细胞结构,减少表面活性物质,可能影响肺泡表面张力和通气功能。
[Abstract]:Objective chitosan oligosaccharide stearic acid graft nanomaterials (chitosan oligosaccharide-stearic acid, CSO-SA) were synthesized by Zhejiang University School of Pharmacology. In our previous study, we found that respiratory distress occurred after CSO-SA was injected into tail vein of mice, which was characterized by shortness of breath, more frequency and shallower depth of respiration. Dead mice dissection found that the color of the lungs red, some lobes bleeding, edema and other phenomena. The aim of this study was to explore the possible mechanism of acute lung injury induced by CSO-SA in mice. Methods three in vivo experiments were carried out: blood gas analysis test, single dose acute toxicity test and repeated lung toxicity test. Blood gas analysis showed that the negative control group had low, medium and high (NaCl), (2550125 mg / kg) and the positive control group (lipopolysaccharide). Blood samples were collected from carotid artery for 1 hour after administration. PH, partial pressure of oxygen, partial pressure of carbon dioxide and saturation of oxygen in blood were analyzed. Single dose acute toxicity test was performed in negative control group, low, medium and high dose group (2550mg / kg) and positive control group. Alveolar lavage was performed 24 hours after administration. The white blood cell count, lung protein content, superoxide dismutase (SOD),) lactate dehydrogenase (LDH),) alkaline phosphatase (AKP) activity and malondialdehyde (MDA) content in lung homogenate were measured. The pulmonary toxicity test of repeated administration was carried out with negative control group, 4 experimental groups (5 ~ 10 ~ 10 ~ 2550mg / kg), and a positive control group. The mice were sacrificed by femoral artery bloodletting 24 hours after the last administration for 5 days. The changes of white blood cell count, protein content and SOD,LDH,AKP activity in lung homogenate were detected, and the number of lung system was calculated. The changes of inflammatory factor interleukin 1 (IL-1 尾), tumor necrosis factor (TNF-a) and the expression of pulmonary surfactant associated protein (SP-A) were detected in tissue homogenate. Results there was significant difference in pH value between 125mg/kg group and negative control group (P0.05). After repeated administration, the weight increased and the number of lung system increased in 25g / kg group, which was significantly different from that in negative control group (P0.05). The total cell count in the lavage fluid was increased in each dose group after a single and continuous administration for 5 days. The difference was significant (p0.05) .75mg / kg group treated for 24 hours and the 50mg/kg group increased the protein content of alveolar lavage fluid for 5 days. There was a significant difference compared with the negative control (p0.05). Repeated administration of 25g / kg of 25 渭 g / kg increased LDH in lung homogenate (10 ~ 2550mg / kg), which was significantly higher than that of negative control (p0.05). The level of IL-1 尾 in the repeated administration group was significantly higher than that in the negative control group (p0.05), while the level of TNF- 伪 was significantly increased (p0.05). The expression of SP-A in each group was significantly lower than that in the negative control group (P0.05). The alveolar structure of 50mg/kg was damaged by HE staining. Electron microscope showed that the apoptosis of alveolar epithelial cells in 50mg/kg group was higher than that in normal group, and the vacuolation of lamellar structure of type 鈪，
本文编号：2206520
[Abstract]:Objective chitosan oligosaccharide stearic acid graft nanomaterials (chitosan oligosaccharide-stearic acid, CSO-SA) were synthesized by Zhejiang University School of Pharmacology. In our previous study, we found that respiratory distress occurred after CSO-SA was injected into tail vein of mice, which was characterized by shortness of breath, more frequency and shallower depth of respiration. Dead mice dissection found that the color of the lungs red, some lobes bleeding, edema and other phenomena. The aim of this study was to explore the possible mechanism of acute lung injury induced by CSO-SA in mice. Methods three in vivo experiments were carried out: blood gas analysis test, single dose acute toxicity test and repeated lung toxicity test. Blood gas analysis showed that the negative control group had low, medium and high (NaCl), (2550125 mg / kg) and the positive control group (lipopolysaccharide). Blood samples were collected from carotid artery for 1 hour after administration. PH, partial pressure of oxygen, partial pressure of carbon dioxide and saturation of oxygen in blood were analyzed. Single dose acute toxicity test was performed in negative control group, low, medium and high dose group (2550mg / kg) and positive control group. Alveolar lavage was performed 24 hours after administration. The white blood cell count, lung protein content, superoxide dismutase (SOD),) lactate dehydrogenase (LDH),) alkaline phosphatase (AKP) activity and malondialdehyde (MDA) content in lung homogenate were measured. The pulmonary toxicity test of repeated administration was carried out with negative control group, 4 experimental groups (5 ~ 10 ~ 10 ~ 2550mg / kg), and a positive control group. The mice were sacrificed by femoral artery bloodletting 24 hours after the last administration for 5 days. The changes of white blood cell count, protein content and SOD,LDH,AKP activity in lung homogenate were detected, and the number of lung system was calculated. The changes of inflammatory factor interleukin 1 (IL-1 尾), tumor necrosis factor (TNF-a) and the expression of pulmonary surfactant associated protein (SP-A) were detected in tissue homogenate. Results there was significant difference in pH value between 125mg/kg group and negative control group (P0.05). After repeated administration, the weight increased and the number of lung system increased in 25g / kg group, which was significantly different from that in negative control group (P0.05). The total cell count in the lavage fluid was increased in each dose group after a single and continuous administration for 5 days. The difference was significant (p0.05) .75mg / kg group treated for 24 hours and the 50mg/kg group increased the protein content of alveolar lavage fluid for 5 days. There was a significant difference compared with the negative control (p0.05). Repeated administration of 25g / kg of 25 渭 g / kg increased LDH in lung homogenate (10 ~ 2550mg / kg), which was significantly higher than that of negative control (p0.05). The level of IL-1 尾 in the repeated administration group was significantly higher than that in the negative control group (p0.05), while the level of TNF- 伪 was significantly increased (p0.05). The expression of SP-A in each group was significantly lower than that in the negative control group (P0.05). The alveolar structure of 50mg/kg was damaged by HE staining. Electron microscope showed that the apoptosis of alveolar epithelial cells in 50mg/kg group was higher than that in normal group, and the vacuolation of lamellar structure of type 鈪，
本文编号：2206520
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