青藤碱干预多柔比星肾病大鼠的实验研究

目的:观察白介素-6(IL-6)在多柔比星肾病大鼠肾组织及尿液中的表达及青藤碱(SIN)对其表达的影响.方法:雄性Wistar大鼠120只.随机分为正常组32只,肾病组88只.肾病组大鼠单次尾静脉注射多柔比星5mg/kg,制备微小病变型肾病(MCNS)模型.采用酶联免疫吸附法(ELISA)检测各组大鼠不同时期肾组织及尿...     

The Anti-Angiogenic Effect of Sinomenine

Sinomenine is an alkaloid extracted from the Chinese medicinal plant, Sinomenium acutum, which has been utilized to treat rheumatoid arthritis (RA) in China for over 2000 years. Sinomenine has been shown to mediate a wide range of pharmacological actions which includes anti-inflammatory and anti-rheumatic effects. RA has been classified as a chronic immune-mediated disease that exhibits overlapping manifestation of inflammatory, abnormal cellular and hormonal immune responses with synovial hyperplasia. Since, angiogenesis is recognized to play a critical role in the development of RA and anti-angiogenic therapy has been proposed as a new therapeutic strategy for treatment of RA, we would like to see if sinomenine possesses anti-angiogenic property. In this study, sinomenine inhibited bFGF-induced proliferation of human umbilical vein endothelial cells (HUVEC) and arrested its cell cycle in G1 phase. Sinomenine disrupted tube formation of HUVEC on Matrigel and suppressed the chemotaxis of HUVEC. In addition, sinomenine reduced neovascularization in Matrigel plug assay as well as microvascular outgrowth in rat aorta ring sprouting assay. These results suggest that sinomenine inhibited bFGF-induced angiogenesis in vitro and in vivo. As the leukocytes–endothelial adhesive interactions also play an important role in inflammation, we found that sinomenine reduced the transmigration of granulocytic differentiated HL60 cells across IL-1β activated HUVEC monolayer. Therefore, the inhibition of leukocytes migration across blood vessel walls and the anti-angiogenic effect of sinomenine may contribute towards its therapeutic mechanisms in alleviating the pathogenesis of RA.     

盐酸青藤碱醇质体的制备及其体外经皮渗透研究

目的 制备盐酸青藤碱醇质体并考察醇质体作为盐酸青藤碱经皮给药载体的渗透特性。方法 采用注入法制备盐酸青藤碱醇质体,并对其形态、粒径及包封率进行分析;采用透皮扩散仪,以小鼠皮肤进行体外透皮试验,比较盐酸青藤碱在水溶液、脂质体以及醇质体中的透皮行为。结果 制得的盐酸青藤碱醇质体外形圆整,平均粒径为88.7 nm,包封率为62.8%,累积透皮量和透皮速率均明显高于盐酸青藤碱脂质体和水溶液。结论 醇质体能明显促进盐酸青藤碱的经皮渗透,有望成为盐酸青藤碱透皮给药的新载体。     

盐酸青藤碱微囊的制备及其质量评价

目的 研究盐酸青藤碱微囊的制备工艺,并考察其体外释药特性。方法 用复凝聚法制备盐酸青藤碱微囊,以包封率为指标,用正交试验设计法对盐酸青藤碱的制备工艺进行研究,用高效液相色谱法研究制备的微囊体外释药特性,并且对其形态、稳定性等进行研究。结果 盐酸青藤碱微囊的囊心囊材比、搅拌速率及成囊温度对微囊的包封率均有显著影响,当囊心与囊材比为1∶3、搅拌速率为200 r/min、成囊温度为60 ℃时,制得的盐酸青藤碱微囊囊形圆整光滑,囊壁清晰,粒径均匀,平均包封率可高达85%,载药量平均为16.5%,囊径为4～12 μm。结论 复凝聚法制备盐酸青藤碱微囊工艺简单、可靠,产品稳定性好。微囊作为一种新兴剂型,具有一定的缓释效果。     

Influence of sinomenine on protein profiles of peripheral blood mononuclear cells from ankylosing spondylitis patients: a pharmacoproteomics study

Background Ankylosing spondylitis （AS） is a common inflammatory rheumatic disease which lacks satisfactory treatment so far.Sinomenine （SIN） is an alkaloid and has recently been utilized in treating multiple rheumatic diseases including AS in China,but its exact mechanism remains to be explored.This study investigated the alteration of proteome in peripheral blood mononuclear cells （PBMCs） from AS patients.Methods Thirty AS patients were enrolled in this study.PBMCs from each AS patient were cultured in medium with or without SIN respectively.Then PBMCs proteins from both groups were separated by two-dimensional electrophoresis （2-DE） and analyzed by mass spectrometry （MS）.Two differentially expressed proteins were then chosen to be verified using Western blotting.Results Seven proteins,including α-synuclein （SNCA）,calmodulin （CALM）,acidic leucine-rich nuclear phosphoprotein 32 family member A （ANP32A）,chloride intracellular channel protein 1 （CLIC1）,guanine nucleotide-binding protein G（I）/G（S）/ G（T） subunit beta-1 （GNB1）,gelsolin （GSN） and histone H2B type 1-M （HISTH2BM）were over-expressed,while coronin1A （CORO1A） was under-expressed in the SIN-treated PBMCs.Further bioinformatics search indicated that the changes of SNCA,ANP32A and CLIC1 pertained to apoptosis,while changes of GSN and CORO1A were associated with both apoptosis and inhibition of immunological function.Subsequently GSN and CORO1A were selected to validate by Western blotting and the results were consistent with those of 2-DE.Conclusion There were 8 differentially expressed proteins in the SIN-treated PBMCs,which might shed some light on the mechanism of SIN in the treatment of AS.     

青风藤化学成分和药理作用的研究进展及其质量标志物（Q-Marker）的预测分析

青风藤主要含有生物碱、挥发油、脂和甾体等活性成分。以青藤碱为代表的异喹啉类生物碱目前研究居多,其在风湿痹痛、关节肿痛、麻痹瘙痒中显示出良好的临床效果。青风藤在临床中应用越来越广泛,其质量评价方法亟需建立。根据中药质量标志物(quality marker,Q-Marker)新概念,从植物亲缘学和化学成分特有性、有效性、药性相关性、化学成分可测性等方面对青风藤Q-Marker进行预测分析,青藤碱、木兰花碱、蝙蝠葛任碱、左旋千金藤啶碱、千金藤宁碱、去甲乌药碱、尖防己碱等多种生物碱及挥发油、甾醇、蒽醌、菲及酚类物质可作为青风藤的主要Q-Marker,为青风藤质量评价体系的建立提供依据和相关新药研发提供参考。     

关节腔注射剂青藤碱纳米晶自稳定Pickering乳液的制备及药效学研究

目的 通过制备青藤碱纳米晶自稳定Pickering乳液（sinomenine nanocrystals self-stabilized Pickering emulsions,Sin- NSSPE）关节腔注射剂,在关节腔内形成药物贮库,缓慢释放以提高生物利用度,降低药物毒副作用,以青藤碱纳米晶（sinomenine nanocrystal,Sin-NCs）自身为稳定剂制备Sin-NSSPE关节腔注射剂,用药效学实验考察其对类风湿性关节炎（rheumatoid arthritis,RA）的治疗效果。方法 通过药物质量浓度、油水相比例和水相pH值等因素对Sin-NSSPE进行优化,运用福氏佐剂关节炎方法建立RA大鼠模型,以关节肿胀度、关节炎指数、脾指数、酶联接免疫吸附剂测定（enzyme-linked immunosorbnent assay,ELISA）、组织病理学检查、Western blotting法作为指标进行检测。结果 结果显示Sin-NSSPE的粒径均匀,载药量良好,Sin-NCs的平均粒径为（121.49±18.26）nm、Sin-NSSPE的粒径为（1 159.60±160.15）nm、载药量4.92 mg/mL;药效学实验结果显示,与青藤碱ig组相比,Sin-NSSPE关节腔注射组能有效降低关节肿胀度和炎症指数,改善滑膜组织的病变、抑制关节炎症。青藤碱关节腔注射组和Sin-NSSPE关节腔注射组均能明显改善大鼠关节肿胀,降低滑膜组织中炎症因子白细胞介素-1（interleukin-1,IL-1）、IL-6、肿瘤坏死因子-α（tumor necrosis factor-α,TNF-α）表达（P＜0.05、0.01）,且Sin-NSSPE关节腔注射组治疗效果优于青藤碱关节注射组,同时剂型稳定性提高并形成药物贮库,使药效更加平稳。结论 制备的Sin-NSSPE性质稳定,Sin-NSSPE关节腔注射对大鼠RA有良好的疗效。     

青藤碱通过线粒体途径诱导肺癌NCI-H460细胞凋亡的初步研究

目的:研究青藤碱(sinomenine,SIN)对人肺癌NCI-H460细胞株的生长抑制及诱导凋亡作用及其机制。方法:四甲基偶氮噻蓝（MTT）法检测细胞增殖的抑制作用,流式细胞仪AnnexinV/PI双染法检测细胞凋亡,TdT酶介导的dUTP缺口末端标记（TUNNEL）方法观察细胞的凋亡,罗丹明123(Rhodamine123)染色流式细胞仪检测线粒体膜电位(ΔΨm)。结果:SIN对NCI-H460细胞株生长具有抑制作用并诱导凋亡。AnnexinV/PI双染检测细胞的凋亡可见随SIN浓度增加,细胞凋亡增加呈浓度依赖性。TUNNEL阳性的凋亡细胞呈棕黄色,细胞核片段化改变。罗丹明染色检测线粒体膜电位的结果提示在SIN作用于NCI-H460细胞48h后,线粒体膜电位下降,SIN浓度越高,膜电位下降越显著。结论:SIN具有明显的细胞毒作用,能诱导NCI-H460细胞凋亡,SIN通过线粒体途径诱导NCI-H460细胞凋亡。     

星点设计-效应面法优化盐酸青藤碱脉冲片处方的研究

目的采用星点设计-效应面法优化盐酸青藤碱脉冲片处方。方法采用干法压制包衣法制备盐酸青藤碱脉冲片,以片芯中羧甲淀粉钠的用量、包衣处方中凝胶材料比例(羟丙甲纤维素∶卡拉胶)和凝胶材料用量为考察因素,以释药时滞为考察指标,采用星点设计安排实验,并对指标与因素进行多元线性和二项式数学拟合,通过效应面法预测最佳处方,并对其进行验证。结果指标与因素之间可采用二项式拟合,相关系数为0.993 7;采用优化条件制备的3种处方的脉冲片均经一定时滞(6 h左右)后药物呈脉冲释放,释药时滞实测值与预测值偏差均在±4.43%以内。结论盐酸青藤碱脉冲片具有体外定时脉冲释药特性,星点设计-效应面优化法可用于本制剂的处方优化,所建立的数学模型预测性良好。     

青藤碱水凝胶贴剂的微针经皮给药的研究

目的在微针作用下,考察青藤碱水凝胶贴剂透皮给药的特点与规律。方法制备不同长度的微针,并制备青藤碱水凝胶贴剂;离体小鼠皮肤经不同针形微针预处理相同时间、相同针形微针预处理不同时间后贴敷青藤碱水凝胶贴剂,用改进的Franz扩散池研究青藤碱的透过规律并与未处理皮肤的被动扩散进行比较;高效液相色谱法测定青藤碱的含量。结果 100μm微针、200μm微针预处理皮肤后青藤碱透皮速率分别是未经微针预处理的40.7、52.4倍;200μm微针预处理皮肤7 min后青藤碱的透皮速率是未经微针预处理的142.0倍。200μm微针经不同的力预处理后,青藤碱的透皮速率随着力的增大而增大,当力大于5 N时透皮速率趋于平衡。结论微针与水凝胶贴剂结合经皮给药时,透皮速率显著提高,并且微针针形、预处理时间和所受的力对药物的经皮渗透具有重要影响。