姜黄素固体脂质纳米粒干粉吸入剂的体外释药、体内急性毒性及对哮喘模型小鼠炎症反应的影响
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摘要
目的:研究姜黄素(Cur)固体脂质纳米粒(SLN)干粉吸入剂(DPI)的体外释药性能、小鼠体内急性毒性及对哮喘模型小鼠炎症反应的影响。方法:采用喷雾干燥法将微乳法制备的Cur-SLN混悬液微粉化后与乳糖(200目)等充分混匀制成Cur-SLN-DPI。采用动态膜透析法考察体外释药情况,比较Cur原料药、Cur-SLN、Cur-SLN-DPI分别在3种释放介质[含1.0%十二烷基硫酸钠(SDS)的磷酸盐缓冲液(PBS,pH 7.4)、含0.2%聚山梨酯80的PBS(pH 7.4)、生理盐水-20%乙醇溶液]中释放5、15、30 min和1、1.5、3、6、8、12、18、24、36、48 h的释放曲线,并筛选合适的释药模型。按急性毒性试验要求考察尾静脉注射最大剂量2 000 mg/kg Cur-SLN-DPI对KM小鼠的影响。将KM小鼠随机分为正常对照组、模型组、阳性药物组(布地奈德3 mg)和Cur-SLN-DPI高、低剂量组(100、50 mg/kg),每组7只,以卵蛋白(OVA)为致敏原复制哮喘模型,每周1、3、5雾化OVA激发哮喘前30 min雾化给药,连续3周。末次激发24 h内,计数肺泡灌洗液(BALF)中白细胞总数、淋巴细胞数、中性粒细胞数和嗜酸性粒细胞数,观察支气管和肺组织的病理学变化。结果:与Cur原料药比较,Cur-SLN和Cur-SLN-DPI均具有缓释作用,且Cur-SLN-DPI在3种释放介质中的缓释更平稳,释药特征均符合Weibull模型。尾静脉注射2 000 mg/kg Cur-SLN-DPI对小鼠无明显的急性毒性。与正常对照组比较,模型组小鼠BALF中白细胞总数、淋巴细胞数、中性粒细胞数、嗜酸性粒细胞数均明显增加(P<0.01),支气管黏膜上皮被覆假复层纤毛柱状细胞,周围炎症细胞浸润严重,肺淤血,中度间质性肺炎;与模型组比较,各给药组小鼠BALF中上述细胞数均明显降低(P<0.01),Cur-SLN-DPI低、高剂量组小鼠气管病变均改善,肺部淤血减轻,且高剂量组减轻更明显。结论:Cur-SLN-DPI具有体外缓释作用,对小鼠无明显急性毒性,可改善哮喘模型小鼠的气管炎症反应和肺部淤血程度。
OBJECTIVE:To study in vitro drug release and acute toxicity in vivo of Curcumin(Cur)solid lipid nanoparticles(SLN)dry powder inhaler(DPI)and its effects of inflammatory response in asthmatic model mice. METHODS:Cur-SLN-DPI was obtained with spray-drying method by micronizing the Cur-SLN suspension prepared by the microemulsion method and thoroughly mixing with lactose(200 mesh) etc. The drug release in vitro was investigated by dynamic membrane dialysis.Accumulative release rates(Q)of Cur raw material,Cur-SLN and Cur-SLN-DPI in 3 kinds of release mediums [phosphate buffer solution(PBS,pH 7.4)containing 1.0% sodium dodecyl sulfate(SDS),PBS(pH 7.4)containing 0.2% tween 80,normal saline-20% ethanol solution] were compared 5,15,30 min and 1,1.5,3,6,8,12,18,24,36,48 h after releasing. Drug release model was fitted. The effects of intravenous injection of maximal dose 2 000 mg/kg Cur-SLN-DPI via tail vein on KM mice were investigated by acute toxicity test. KM mice were randomly divided into normal control group,model group,positive drug group(budesonide 3 mg),Cur-SLN-DPI high-dose and low-dose groups(100,50 mg/kg),with 7 mice in each group. The ovalbumin(OVA)was used as sensitizer to induce asthma model;the model mice were given relevant medicine with aerosol administration30 min before aerosol administration of OVA inducing asthma on Monday,Wednesday and Friday per week,for consecutive 3 weeks. Within 24 h after last induction,total number of leukocyte,the number of lymphocyte,neutrophil and eosinophil were counted in broncho alveolar lavage fluid(BALF);the pathological changes of bronchus and lung tissue were observed. RESULTS:Compared with Cur raw material,Cur-SLN and Cur-SLN-DPI showed good sustained-release effect,and Cur-SLN-DPI had more stable sustained release in 3 kinds of release mediums. The characteristics of drug release conformed to the Weibull model.Intravenous injection of 2 000 mg/kg Cur-SLN-DPI via tail vein had no significant acute toxicity in mice. Compared with normal control group,total number of leukocyte,the number of lymphocyte, neutrophil and eosinophil were increased significantly(P<0.01);bronchial mucosal epithelium was covered with pseudostratified ciliated columnar cells,with severe infiltration of inflammatory cells,pulmonary congestion and moderate interstitial pneumonia. Compared with model group,the number of above cells in BALF of mice were decreased significantly in administration group(P<0.01);tracheal lesions of mice were improved in Cur-SLN-DPI low-dose and high-dose groups;pulmonary congestion of them were alleviated,and that of highdose group was alleviated more significantly. CONCLUSIONS:Cur-SLN-DPI shows sustained-release effect in vitro and has no obvious acute toxicity to mice. Cur-SLN-DPI can improve the inflammatory response of the airway and the degree of pulmonary congestion in asthmatic model mice.
OBJECTIVE:To study in vitro drug release and acute toxicity in vivo of Curcumin(Cur)solid lipid nanoparticles(SLN)dry powder inhaler(DPI)and its effects of inflammatory response in asthmatic model mice. METHODS:Cur-SLN-DPI was obtained with spray-drying method by micronizing the Cur-SLN suspension prepared by the microemulsion method and thoroughly mixing with lactose(200 mesh) etc. The drug release in vitro was investigated by dynamic membrane dialysis.Accumulative release rates(Q)of Cur raw material,Cur-SLN and Cur-SLN-DPI in 3 kinds of release mediums [phosphate buffer solution(PBS,pH 7.4)containing 1.0% sodium dodecyl sulfate(SDS),PBS(pH 7.4)containing 0.2% tween 80,normal saline-20% ethanol solution] were compared 5,15,30 min and 1,1.5,3,6,8,12,18,24,36,48 h after releasing. Drug release model was fitted. The effects of intravenous injection of maximal dose 2 000 mg/kg Cur-SLN-DPI via tail vein on KM mice were investigated by acute toxicity test. KM mice were randomly divided into normal control group,model group,positive drug group(budesonide 3 mg),Cur-SLN-DPI high-dose and low-dose groups(100,50 mg/kg),with 7 mice in each group. The ovalbumin(OVA)was used as sensitizer to induce asthma model;the model mice were given relevant medicine with aerosol administration30 min before aerosol administration of OVA inducing asthma on Monday,Wednesday and Friday per week,for consecutive 3 weeks. Within 24 h after last induction,total number of leukocyte,the number of lymphocyte,neutrophil and eosinophil were counted in broncho alveolar lavage fluid(BALF);the pathological changes of bronchus and lung tissue were observed. RESULTS:Compared with Cur raw material,Cur-SLN and Cur-SLN-DPI showed good sustained-release effect,and Cur-SLN-DPI had more stable sustained release in 3 kinds of release mediums. The characteristics of drug release conformed to the Weibull model.Intravenous injection of 2 000 mg/kg Cur-SLN-DPI via tail vein had no significant acute toxicity in mice. Compared with normal control group,total number of leukocyte,the number of lymphocyte, neutrophil and eosinophil were increased significantly(P<0.01);bronchial mucosal epithelium was covered with pseudostratified ciliated columnar cells,with severe infiltration of inflammatory cells,pulmonary congestion and moderate interstitial pneumonia. Compared with model group,the number of above cells in BALF of mice were decreased significantly in administration group(P<0.01);tracheal lesions of mice were improved in Cur-SLN-DPI low-dose and high-dose groups;pulmonary congestion of them were alleviated,and that of highdose group was alleviated more significantly. CONCLUSIONS:Cur-SLN-DPI shows sustained-release effect in vitro and has no obvious acute toxicity to mice. Cur-SLN-DPI can improve the inflammatory response of the airway and the degree of pulmonary congestion in asthmatic model mice.
引文
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[13]李楠,李锡晶,王倩,等.微乳法制备姜黄素固体脂质纳米粒[J].中国药房,2015,26(19):2698-2702.
[14]李楠,于美丽.姜黄素固体脂质纳米粒干粉吸入剂的制备工艺研究[J].中国药房,2016,27(28):3979-3981.
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[16]陈德,刘意,范凯燕,等.姜黄素微球中药物存在形式与释药行为的关系研究[J].药学学报,2016,51(1):140-146.
[17]国家药典委员会.中华人民共和国药典:四部[S].2015年版.北京:中国医药科技出版社,2015:120-124.
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[20]DAL MAGRO R,ORNAGHI F,CAMBIANICA I,et al.ApoE-modified solid lipid nanoparticles:a feasible strategy to cross the blood-brain barrier[J].J Control Release,2017.DOI:10.1016/j.jconrel.2017.01.039.
[21]陈亚媛,赵娣,李宁.缓释型肺吸入制剂的研究进展[J].药学研究,2018,37(8):469-481.
[2]CHEN ZH,WANG PL,SHEN HH.Asthma research in China:a five-year review[J].Respirology,2013.DOI:10.1111/resp.12196.
[3]张改,王莹.支气管哮喘治疗的研究进展[J].科技信息,2014(9):262-263.
[4]BARNES PJ.Glucocorticosteroids:current and future directions[J].Br J Pharmacol,2011,163(1):29-43.
[5]叶翔.支气管哮喘的最新治疗研究进展[J].实用心脑肺血管病杂志,2014,22(7):5-7.
[6]CHEE C,SELLAHEWA L,PAPPACHAN JM.Inhaled corticosteroids and bone health[J].Open Respir Med J,2014,8(Suppl 1):85-92.
[7]薛海英,蒋国英,张宝辉.姜黄素对哮喘小鼠气道炎症和肺内诱导型一氧化氮合酶的影响[J].解剖科学进展,2018,24(2):163-165.
[8]KARAMAN M,FIRINCI F,BAGRIYANIK A,et al.Effects of curcumin on lung histopathology and fungal burden in a mouse model of chronic asthma and oropharyngeal candidiasis[J].Arch Med Res,2011,42(2):79-87.
[9]JANG DJ,KIM ST,LEE K,et al.Improved bioavailability and antiasthmatic efficacy of poorly soluble curcuminsolid dispersion granules obtained using fluid bed granulation[J].Bio-Medical Materials and Engineering,2014,24(1):413-429.
[10]JANG DJ,KIM ST,OH E,et al.Enhanced oral bioavailability and antiasthmatic efficacy of curcumin using redispersible dry emulsion[J].Bio-Medical Materials and Engineering,2014,24(1):917-930.
[11]CHAUHAN PS,SUBHASHINI,DASH D,et al.Intranasal curcumin attenuates airway remodeling in murine model of chronic asthma[J].Int Immunopharmacol,2014,21(1):63-75.
[12]王风秀,于锋英.姜黄素药物递送系统研究进展[J].中国药业,2014,23(18):120-121.
[13]李楠,李锡晶,王倩,等.微乳法制备姜黄素固体脂质纳米粒[J].中国药房,2015,26(19):2698-2702.
[14]李楠,于美丽.姜黄素固体脂质纳米粒干粉吸入剂的制备工艺研究[J].中国药房,2016,27(28):3979-3981.
[15]李楠,刘佩莉,孔令钰,等.姜黄素固体脂质纳米粒干粉吸入剂的质量评价及初步稳定性考察[J].中国药房,2016,27(34):4838-4841.
[16]陈德,刘意,范凯燕,等.姜黄素微球中药物存在形式与释药行为的关系研究[J].药学学报,2016,51(1):140-146.
[17]国家药典委员会.中华人民共和国药典:四部[S].2015年版.北京:中国医药科技出版社,2015:120-124.
[18]魏伟,吴希美,李元建.药理实验方法学[M].4版.北京:人民卫生出版社,2010:383-393.
[19]WEBER S,ZIMMER A,PARDEIKE J.Solid lipid nanooparticles(SLN)and nanostructured lipid carriers(NLC)for pulmonary application:a review of the state of the art[J].Eur J Pharm Biopharm,2014,86(1):7-22.
[20]DAL MAGRO R,ORNAGHI F,CAMBIANICA I,et al.ApoE-modified solid lipid nanoparticles:a feasible strategy to cross the blood-brain barrier[J].J Control Release,2017.DOI:10.1016/j.jconrel.2017.01.039.
[21]陈亚媛,赵娣,李宁.缓释型肺吸入制剂的研究进展[J].药学研究,2018,37(8):469-481.