【摘要】：目的 殼寡糖硬脂酸嫁接物納米材料(chitosan oligosaccharide-stearic acid, CSO-SA)是由浙江大學(xué)藥學(xué)院合成的納米靶向載藥物質(zhì)。我們?cè)谇捌谘芯恐邪l(fā)現(xiàn)小鼠尾靜脈注射CSO-SA后,會(huì)發(fā)生呼吸窘迫,表現(xiàn)為呼吸局促,頻次變多,呼吸深度變淺。死亡小鼠解剖時(shí)發(fā)現(xiàn)肺臟顏色偏紅,部分肺葉有出血點(diǎn)、水腫等現(xiàn)象。本研究的目的是探索CSO-SA導(dǎo)致小鼠急性肺損傷的可能機(jī)制。 方法 開(kāi)展3個(gè)體內(nèi)實(shí)驗(yàn)：血?dú)夥治鰧?shí)驗(yàn)、單次給藥急性毒性實(shí)驗(yàn)、重復(fù)給藥肺部毒性實(shí)驗(yàn)。血?dú)夥治鰧?shí)驗(yàn)設(shè)置陰性對(duì)照組(NaCl),低、中、高劑量組(25、50、125mg/kg),陽(yáng)性對(duì)照組(脂多糖)。給藥1h后頸動(dòng)脈采血,分析血液中pH值、動(dòng)脈氧分壓、二氧化碳分壓和血氧飽和度等指標(biāo)。單次給藥急性毒性實(shí)驗(yàn)設(shè)置陰性對(duì)照組,低、中、高劑量組(25、50、75mg/kg),陽(yáng)性對(duì)照組。給藥后24小時(shí)進(jìn)行肺泡灌洗。灌洗液測(cè)定白細(xì)胞數(shù)量、肺蛋白含量、超氧化物歧化酶(SOD)、乳酸脫氫酶(LDH)、堿性磷酸酶(AKP)活性變化,肺組織勻漿測(cè)定丙二醛(MDA)含量變化。重復(fù)給藥肺部毒性實(shí)驗(yàn)設(shè)置陰性對(duì)照組,4個(gè)實(shí)驗(yàn)組(5、10、25、50mg/kg),陽(yáng)性對(duì)照組。連續(xù)5天給藥,最后一次給藥后24小時(shí)股動(dòng)脈放血處死小鼠。檢測(cè)灌洗液中白細(xì)胞數(shù)量、蛋白含量,肺組織勻漿中SOD、LDH、AKP活性、MDA含量變化；計(jì)算肺臟體系數(shù),組織勻漿檢測(cè)炎癥因子白介素1p(IL-1β)、腫瘤壞死因子(TNF-a)的變化和肺表面活性相關(guān)蛋白(SP-A)表達(dá)量變化,HE染色和電鏡染色觀察肺組織損傷情況。SPSS18.0對(duì)數(shù)據(jù)進(jìn)行統(tǒng)計(jì)分析,P0.05認(rèn)為有統(tǒng)計(jì)學(xué)差異。 結(jié)果 給藥后125mg/kg組血液呈現(xiàn)酸性,pH值與陰性對(duì)照相比有顯著性差異(P0.05)。重復(fù)給藥后25、50mg/kg組體重增長(zhǎng)減少,肺臟體系數(shù)升高,與陰性對(duì)照組存在顯著性差異(P0.05)。單次及連續(xù)5天給藥后,各劑量組都能引起灌洗液中總細(xì)胞計(jì)數(shù)上升,差異有顯著性(p0.05)。75mg/kg組作用24小時(shí)和50mg/kg組連續(xù)5天給藥肺泡灌洗液蛋白含量增加,與陰性對(duì)照相比差異有顯著性(p0.05)。重復(fù)給藥25、50mg/kg引起肺組織勻漿LDH升高,10、25、50mg/kg引起AKP升高,與陰性對(duì)照相比差異有顯著性(p0.05)。重復(fù)給藥各劑量組中IL-1β水平與陰性對(duì)照相比升高,差異有顯著性(p0.05),而5、25、50mg/kg均引起了TNF-α水平升高,差異有顯著性(p0.05)。各組SP-A表達(dá)水平與陰性對(duì)照相比減少,其中10、25、50mg/kg組存在顯著性差異(P0.05)。HE染色顯示50mg/kg的肺泡結(jié)構(gòu)受到破壞。電鏡顯示50mg/kg組的肺泡上皮細(xì)胞凋亡比正常組增多,Ⅱ型上皮細(xì)胞特有的板層體結(jié)構(gòu)空泡化增多。 結(jié)論 1.小鼠單次或重復(fù)尾靜脈注射CSO-SA后,主要靶器官為肺,能引起肺部炎癥和出血,這可能是小鼠急性死亡的主要原因。 2.反復(fù)尾靜脈注射CSO-SA,能誘導(dǎo)小鼠肺部細(xì)胞凋亡,破壞肺泡Ⅱ型上皮細(xì)胞結(jié)構(gòu),減少表面活性物質(zhì),可能影響肺泡表面張力和通氣功能。
[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- 偽 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，

本文編號(hào)：2206520