二妙散对巨噬细胞分化的调控作用研究

目的 研究二妙散对脂多糖（lipopolysaccharide,LPS）诱导巨噬细胞分化的影响。方法 噻唑蓝（methyl thiazolyltetrazolium,MTT）法测定二妙散水煎液对细胞活力的影响;将LPS （100 ng·mL^-1）及不同浓度（0.1,1.0,10 mg·mL^-1）的二妙散水煎液共同作用RAW264.7细胞后,酶联免疫吸附法（enzyme-linked immune sorbent assay,ELISA）检测上清液中肿瘤坏死因子-α（tumor necrosis factor-α,TNF-α）、白介素-10（interleukin 10,IL-10）的含量,实时荧光定量PCR （Real-timefluorescent quantitative PCR,qPCR）检测RAW264.7细胞中白介素-1β（Interleukin 1β,IL-1β）,IL-6的mRNA水平,Westernblot检测RAW264.7细胞NOS2的蛋白表达情况。结果 二妙散水煎液在0.01~100 mg·mL^-1内对RAW264.7细胞无明显毒性。与LPS诱导的模型组相比,二妙散水煎液能明显降低细胞上清液中TNF-α的含量（P<0.05）、升高IL-10的含量（P<0.05）,且可显著抑制RAW264.7细胞中M1分化标记物IL-1β、IL-6的mRNA水平（P<0.05）及NOS2的蛋白表达（P<0.05）。结论 二妙散可通过抑制巨噬细胞向M1促炎方向分化,从而发挥抗炎作用。     

消乳散结胶囊联合他莫昔芬治疗乳腺增生症的疗效观察

目的 研究瑞香狼毒Stellera chamaejasme花中黄酮和木脂素类化学成分及其抗氧化活性,分析构效关系。方法 利用大孔吸附树脂、正反相硅胶、Sephadex LH-20等色谱分离材料,通过柱色谱和高效液相色谱等分离方法进行分离纯化,运用核磁共振（NMR）、质谱（MS）等波谱技术鉴定化合物结构,并采用FRAP法、DPPH法和ABTS法对分离得到的化合物进行体外抗氧化活性测试。结果 从瑞香狼毒花甲醇提取物中分离鉴定了12个化合物,分别鉴定为艾黄素（1）、槲皮素（2）、isoscutellarein-8-O-β-D-glucuronopyranoside（3）、槲皮素-3-O-β-D-葡萄糖苷（4）、紫云英苷（5）、hypolaetin-8-O-β-D-glucuronopyranoside（6）、kaempferol 3-O-β-D-glucopyranosyl-（1→2）-O-α-L-xylopyranoside（7）、rel-（3R,3''S,4R,4''S）-3,3'',4,4''-tetrahydro-6,6''-dimethoxy[3,3''-bi-2H-benzopyran]-4,4''-diol（8）、马台树脂醇（9）、乌拉尔醇（10）、环黄芪醇（11）、松脂醇（12）。抗氧化活性实验表明,黄酮和木脂素类化合物均显示了较强的抗氧化活性。结论 化合物1、3、5～7和10为首次从该植物中分离得到;化合物2、4、5和10表现出显著的抗氧化活性,其中黄酮类化合物C-3或C-8位连有糖链会降低其抗氧化活性。     

基于Nrf2/HO-1信号通路研究橄榄苦苷的体外抗炎作用

目的 探讨橄榄苦苷对脂多糖（LPS）诱导的小鼠腹腔巨噬细胞（RAW264.7）炎症的保护作用及机制。方法 MTT法检测橄榄苦苷（0、10、20、40 μmol/L）对RAW264.7细胞活性的影响;用橄榄苦苷（10、20、40 μmol/L）预处理细胞1 h后,LPS诱导炎症模型,Griess试剂检测细胞内Nitrite释放;Western blotting方法检测细胞iNOS、COX-2、Nrf2、Keap-1、HO-1、NQO1蛋白表达水平;流式细胞仪检测细胞内NO、ROS、Ca²⁺的水平;荧光显微镜检测线粒体膜电位（MMP）水平;ELISA法检测细胞上清液中肿瘤坏死因子-α（TNF-α）、白介素-6（IL-6）的释放。结果 与模型组比较,橄榄苦苷组Nitrite释放水平显著降低（P<0.05、0.01、0.001）,iNOS、COX-2蛋白的表达显著降低（P<0.001）,NO的产生显著减少（P<0.05、0.01）;TNF-α、IL-6的释放受到显著抑制（P<0.001）;Ca²⁺释放受到显著抑制（P<0.01）;ROS生成受到显著抑制（P<0.01）;JC-1单体降低,恢复聚合物状态（红色荧光变多）,MMP稳定性增强;Keap1蛋白表达显著降低, Nrf2、HO-1、NQO1蛋白的表达均显著升高（P<0.05、0.01、0.001）。结论 橄榄苦苷对LPS诱导的RAW264.7细胞炎症具有抑制作用,其作用机制可能与激活Nrf2/HO-1信号通路相关。     

三七总皂苷调节iNOS-NO-NF-κB信号通路抑制LPS诱导的RAW246.7细胞炎症研究

目的 探讨三七总皂苷（PNS）对脂多糖（LPS）诱导的RAW264.7细胞iNOS-NO-NF-κB信号通路相关分子表达及活性的影响。方法 MTT比色法检测PNS对RAW264.7细胞增殖的抑制作用;不同浓度PNS（25、50、100 μg/mL）干预体外培养LPS诱导的RAW264.7细胞24、48 h后,Griess法检测NO变化;PNS干预24 h后,Western blotting检测iNOS、NF-κB、p-NF-κB、IKKα、p-IKKα、p-ΙκΒα蛋白表达量的变化;PNS干预4 h后,激光共聚焦显微镜检测NF-κB转位入核情况。结果 PNS浓度大于150 μg/mL时,才表现出显著抑制细胞增殖的作用。25、50、100 μg/mL PNS作用于RAW264.7细胞24和48 h后,与模型组比较,NO生成量均显著降低（P<0.001）。50、100 μg/mL PNS作用于RAW264.7 24 h后,iNOS、NF-κB蛋白表达量显著降低,磷酸化蛋白p-NF-κB、p-IKKα、p-ΙκΒα表达水平也显著降低（P<0.01、0.001）。与模型组比较,PNS 25、50、100 μg/mL组核因子p65入核荧光强度显著降低（P<0.01、0.001）。结论 PNS能够显著抑制LPS诱导的RAW264.7细胞iNOS-NO-NF-κB信号通路的活性,降低细胞炎症反应。     

平炎舒消膏抗炎作用及其机制研究

目的 研究平炎舒消膏（PSO）的抗炎作用并初步探讨其作用机制。方法 采用小鼠腹腔毛细血管通透性模型、大鼠足肿胀模型及大鼠棉球肉芽肿模型,观察PSO的在体抗炎作用。采用酶联免疫吸附法（ELISA）和生物测定法,观察PSO对脂多糖（LPS）诱导的巨噬细胞（RAW264.7）释放的白细胞介素-1（IL-1β）和肿瘤坏死因子-α（TNF-α）含量及生物学活性的影响,探索PSO的抗炎作用机制。结果 体内实验中,PSO显著抑制醋酸所致小鼠腹腔毛细血管通透性升高,明显降低大鼠自身血所致足跖肿胀,减轻大鼠棉球肉芽肿。体外实验中,PSO浓度相关性地抑制LPS诱导的RAW264.7释放IL-1β及TNF-α,降低IL-1β对胸腺细胞的增殖效应及TNF-α对L-929细胞的杀伤作用。结论 平炎舒消膏可减轻各期炎症形成,其机制可能与抑制巨噬细胞释放炎性因子有关。     

基于抗高尿酸血症活性的鸡矢藤物质基础研究

目的 研究鸡矢藤提取物抗高尿酸作用的药效物质基础。方法 次黄嘌呤复制高尿酸小鼠模型,观察鸡矢藤提取物对高尿酸血症小鼠血清尿酸水平的影响;观察对脂多糖诱导的小鼠巨噬细胞RAW264.7的NO水平的影响,对DPPH自由基的清除率。采用HPLC-LTQ/Orbitrap MS法快速鉴定鸡矢藤提取物的化学成分。结果 鸡矢藤提取物可显著降低次黄嘌呤诱导的高尿酸血症小鼠血清尿酸水平（P<0.05或P<0.01）,可显著降低脂多糖诱导的小鼠巨噬细胞RAW264.7的NO水平（P<0.05或P<0.01）,对DPPH自由基清除的IC₅₀为1.4 mg·mL^-1。采用HPLC-LTQ/Orbitrap MS法从鸡矢藤提取物中分析鉴定出14个成分,其中包括京尼平龙胆二糖甙、鸡矢藤次苷甲酯、鸡矢藤苷、京尼平苷、7,8-Dihyroiridoid、鸡矢藤次苷、天竺葵素-3-葡萄糖苷、芦丁、异槲皮素、紫云英苷、飞燕草素、大黄酚、大黄素、绿原酸。结论 应用高尿酸小鼠模型及HPLC-LTQ/Orbitrap MS法快速筛选鸡矢藤提取物中可能的活性成分,可为鸡矢藤抗高尿酸药效物质基础研究提供一种快速、高效的方法学借鉴。     

菥蓂正丁醇部位抑制TLR-4/NF-κB信号通路抗脂多糖诱导的RAW264.7细胞炎症反应作用研究

目的 研究菥蓂Thlaspi arvense水提醇沉部位、正丁醇部位的抗炎活性以及正丁醇部位的作用机制。方法 将菥蓂水提物经过乙醇、正丁醇萃取后制得菥蓂水提醇沉部位、正丁醇部位。采用MTT法确定水提醇沉部位、正丁醇部位（15.625、31.25、62.5、125、250、500、1 000 μg/mL）对RAW264.7细胞的毒性。采用脂多糖（LPS）1 μg/mL诱导RAW264.7细胞产生炎性反应,建立体外炎症细胞模型。Griess法检测水提醇沉部位、正丁醇部位（7.812 5、15.625、31.25、62.5、125、250、500、1 000 μg/mL）对LPS诱导RAW264.7细胞后NO释放量的影响;ELISA法检测水提醇沉部位、正丁醇部位（125、250、500 μg/mL）对白细胞介素-6（IL-6）、肿瘤坏死因子-α（TNF-α）释放量的影响;Western Blotting法检测正丁醇部位（125、250、500 μg/mL）对环氧合酶-2（COX-2）、一氧化氮合酶（iNOS）、白细胞介素1β（IL-1β）、核因子-κB（NF-κB）p-P65、Toll样受体4（TLR-4）、磷酸化NF-κB抑制蛋白（p-IκBα）蛋白表达的影响。结果 菥蓂水提醇沉部位、正丁醇部位质量浓度在1 000 μg/mL以下时对RAW264.7细胞几乎无毒性作用。与模型组比较,菥蓂水提醇沉部位、正丁醇部位62.5、125、250、500、1 000 μg/mL组NO释放量显著下降（P<0.01）;菥蓂水提醇沉部位、正丁醇部位125、250、500 μg/mL组IL-6、TNF-α的分泌量显著降低（P<0.01）,且正丁醇部位作用优于水提醇沉部位;菥蓂正丁醇部位500 μg/mL组的iNOS和250、500 μg/mL组COX-2、IL-1β、NF-κB p-P65、TLR-4、p-IκBα蛋白表达量显著降低（P<0.01）,作用均呈浓度相关性。结论 菥蓂正丁醇部位显著抑制LPS诱导的RAW264.7细胞NO的释放,IL-1β、IL-6、TNF-α的分泌以及COX-2、iNOS蛋白的表达,其抗炎作用机制可能与抑制TLR-4/NF-κB信号通路相关。     

苹果多糖的制备及对脂多糖诱导RAW 264.7细胞凋亡的影响

摘 要 目的：观察苹果多糖（AP）对脂多糖（LPS）诱导RAW 264.7细胞凋亡的影响及其机制。方法: 采用水提醇沉法从苹果果渣中提取苹果多糖,并测定其糖含量和分子量。采用流式细胞仪考察AP对LPS诱导RAW 264.7细胞凋亡的作用。采用Western Blot法考察AP对LPS诱导RAW 264.7细胞中Bcl 2、Bax蛋白表达的影响。结果: 经水提醇沉法提取纯化得到苹果多糖的糖含量为91.3%,重均分子量为184613 Da。流式细胞仪检测结果显示,LPS诱导RAW 264.7细胞凋亡明显高于正常对照组（P<0.01）;与LPS组比较,浓度为0.5 mg·mL^-1的AP作用72 h后RAW 264.7细胞凋亡率显著降低（P<0.01）,浓度为1.0 mg·mL^-1的AP作用48,72 h后RAW 264.7细胞凋亡率显著降低（P<0.01）。Western Blot检测结果显示,与正常对照组比较,LPS诱导RAW 264.7细胞中的Bcl 2蛋白在48,72 h表达明显升高,差异具有统计学意义（P<0.01）;与LPS组比较,浓度为1 mg·mL^-1的AP作用后,升高了LPS诱导的RAW 264.7细胞Bcl 2蛋白的表达,降低其Bax蛋白的表达,差异具有统计学意义（P<0.01）。结论:AP抑制LPS诱导RAW 264.7细胞凋亡,其机制可能是通过提高RAW 264.7细胞中Bcl 2蛋白的表达,降低其Bax蛋白表达。     

平消胶囊联合左甲状腺素钠治疗结节性甲状腺肿的临床研究

目的 对狭叶薰衣草Lavandula angustifolia中的木脂素类化合物进行研究。方法 运用RP-HPLC、TLC、硅胶、凝胶、MCI-gel树脂等方法进行分离纯化,并根据理化性质和波谱数据鉴定化合物的结构。结果 从狭叶薰衣草中分离得到11个木脂素类化合物,分别鉴定为松脂醇（1）、丁香树脂醇（2）、fraxiresinol-4''-O-β-D-glucopyranoside（3）、syringaresinol-4''-O-β-D-glucopyranoside（4）、8-hydroxypinoresinol-4-O-β-D-glucopyranoside（5）、rel-（2α,3β）-7-O-methylcedrusin（6）、落叶松脂醇-4''-O-β-D-葡萄糖苷（7）、（2S,3R）-2,3-dihydro-2-（4-hydroxy-3-methoxyphenyl）-3-hydroxymethyl-7-methoxybenzofuran-5-（trans） propen-1-ol-3-O-β-glucoside（8）、（7S,8R）-dihydrodehydrodiconiferyl alcohol-9-β-D-glucopyranoside（9）、（7R,8R）-7,8-dihydro-9''-hydroxyl-3''-methoxyl-8-hydroxymethyl-7-（4-hydroxy-3-methoxyphenyl）-1''-benzofuranpropanol-9''-O-β-D-glucopyranoside（10）、（E）-3-（（2S,3S）-2-（4-hydroxy-3-methoxyphenyl）-3-hydroxymethyl-7-methoxy-2,3-dihydrobenzofuran-5-yl） allyl-2-hydroxyacetate（11）。结论 11个化合物均首次从狭叶薰衣草中分离得到。     

RAW 264.7细胞炎症模型优化及异喹啉类生物碱抗炎活性研究

目的 对RAW 264.7细胞炎症模型进行优化,并考察异喹啉类生物碱的抗炎活性。方法 采用内毒素脂多糖（LPS）诱导RAW264.7细胞炎症反应,以细胞上清液中炎症因子分泌水平为指标,优化LPS诱导浓度、时间及阳性药地塞米松（DEX）孵育时间,并考察异喹啉类生物碱的抗炎活性。结果 RAW 264.7细胞上清液中肿瘤坏死因子-α（TNF-α）分泌水平随LPS诱导浓度、时间增加而升高,但当LPS质量浓度超过100 ng/mL、诱导时间超过12 h后,TNF-α分泌水平趋于平缓;LPS为20 ng/mL、诱导12 h的TNF-α分泌水平明显升高。DEX对TNF-α分泌抑制作用随孵育时间延长而增强,但孵育4 h后,TNF-α分泌抑制作用趋于平缓。异喹啉类生物碱呈现不同程度TNF-α分泌抑制作用,其抑制活性与其季胺型母核结构及R基团烷氧基取代等密切相关。结论 LPS为20 ng/mL、诱导12 h能较好诱导RAW 264.7细胞炎症反应;异喹啉类生物碱的TNF-α分泌抑制作用与其季胺型母核结构、R基团取代等密切相关。     

包公藤甲素类似物的合成

包公藤甲素(简称包甲素)系从包公藤茎中分离的一个新生物碱,其结构为2β-羟基-6β-乙酰氧基-N-去甲托品烷(1),为一强效M-胆碱受体激动剂,已用于青光眼治疗。项中等已报道了不同的合成方法,合成品为外消旋体,合成步骤长、收率很低。     

EFFECTS OF 2-GUANIDINE-4-METHYLQUINAZOLINE ON GASTRIC ACID SECRETION IN RATS

In this study 2-guanidine-4-methylquinazoline (2-GMQ) appeared to decrease basal and stimulated gastric acid secretion, while structurally related compounds as dimethyl- biguanide, cyanoguanidine and 2-cyanoamino-4-methylpyrymidine did not. Thus, there is an antisecretory effect when the biguanide group is associated with a lipophilic structure. The antisecretive effects exerted by 2-GMQ are associated with anti H₂-histamine activity.The anti H₂-histamine nature of the effects of 2-GMQ was confirmed by the capacity of this compound of depressing the chronotropic activity of the isolated guinea pig auricle increased by histamine, as well as relaxant activity in rat uterus contracted by histamine, since both preparations are rich in H₂-histamine receptors.     

Synthesis and evaluation of 2,6-piperidinedione derivatives as potentially novel compounds with analgesic and other CNS activities

New 2,6-piperidinediones 2_a–g and 4_a–d were prepared by initial condensation of aromatic aldehydes or cycloalkanones with cyanoacetamide to give α-cyanocinnamides l_a–g or cycloalkylidenes 3_a,b which underwent Michae1 addition with ethyl cyanoacetate or diethylmalonate. Compounds 4_a–d were alkylated by various alkyl halides to produce the N-alkylated 2,6-piperidinedione derivatives 5_a–m. Some new selected compounds 2_a–c,f, 4_a–d & 5_e,h,j were pharmacologically evaluated for potential anticonvulsant, sedative and analgesic activities. These compounds exhibited significant anticonvulsant and analgesic effects after a single I.P. administration 100 mg/kg b.wt. . On the other hand all the investigated compounds induced hypnotic activity and prolonged the phenobarbital sodium- induced sleep as compared with the control group and the most potent compound was found to be 2_f.     

Investigation of the prevalence of tetQ, tetX and tetX1 genes in Bacteroides strains with elevated tigecycline minimum inhibitory concentrations

In this study, the antibiotic susceptibilities to tigecycline and tetracycline of 35 selected Bacteroides fragilis group strains were determined by Etest, and the presence of tetQ, tetX, tetX1 and ermF genes was investigated by polymerase chain reaction (PCR). tetQ was detected in all 12 B. fragilis group isolates (100%) exhibiting elevated tigecycline minimum inhibitory concentrations (MICs) (≥8 μg/mL) as well as the 8 strains (100%) with a tigecycline MIC of 4 μg/mL, whilst tetX and tetX1 were present in 15% and 75% of these strains, respectively. All of these strains were fully resistant to tetracycline (MIC ≥ 16 μg/mL). On the other hand, amongst the group of strains with tigecycline MICs < 4 μg/mL (15 isolates), tetQ, tetX and tetX1 were found less frequently (73.3%, 13.3% and 46.7%, respectively). All but two strains harbouring the tetQ gene in this group were non-susceptible to tetracycline, with a MIC > 4 μg/mL. These data suggest that in most cases tigecycline overcomes the tetracycline resistance mechanisms frequently observed in Bacteroides strains. However, the presence of tetX and tetX1 genes in some of the strains exhibiting elevated MICs for tigecycline draws attention to the possible development and spread of resistance to this antibiotic agent amongst Bacteroides strains. The common occurrence of ermF, tetX, tetX1 and tetQ genes together predicted the presence of the CTnDOT-like Bacteroides conjugative transposon in this collection of Bacteroides strains.     

国产醋酸甲地孕酮的质量研究:6-羟基-6-甲基-17α-乙酰氧基黄体酮差向异构体的分离与鉴定

用柱层析及薄层层析对国产醋酸甲地孕酮的杂质进行了分析,共分离出7个杂质点:Ⅰ₁,Ⅰ₂,Ⅰ₃,Ⅰ₄,Ⅰ₅,Ⅰ₆,Ⅰ₇,并对其中Ⅰ₃和Ⅰ₄进行分离纯制及光谱分析,确定其结构为6β-羟基-6α-甲基-17α-乙酰氧基黄体酮及其差向异构体6α-羟基-6β-甲基-17α-乙酰氧基黄体酮。本文报道了薄层层析条件(硅胶GF₂₅₄,展开剂:醋酸乙酯-甲苯=7:3)以及光谱鉴别的依据。     

EFFECT OF POLICOSANOL SUCCESSIVE DOSE INCREASES ON PLATELET AGGREGATION IN HEALTHY VOLUNTEERS

Policosanol is a cholesterol-lowering drug with hypocholesterolemic effects demonstrated in experimental models, healthy volunteers and type II hypercholesterolemic patients. In addition, antiplatelet effects of policosanol have been shown in experimental models and healthy volunteers. The effect of successively increasing doses of policosanol on platelet aggregation was investigated in a randomized, placebo-controlled, double-blind study conducted in 37 healthy volunteers. The volunteers were on a placebo-baseline period (two tablets per day) for 7 days and thereafter they received randomly, under double-blind conditions, placebo or policosanol (10mgday⁻¹) for 7 days. After this period dosage was doubled to 20mgday⁻¹for the next 7 days and then again doubled to 40mgday⁻¹, while the control group received placebo tablets all the time. Platelet aggregation as well as coagulation time was measured at baseline and after each dosing step. Results showed that antiplatelet effects of policosanol were successfully enhanced throughout the study, thus suggesting a dose-dependent relationship. No significant effect was reached during the first dosing period, but significant reductions of epinephrine and ADP-induced platelet aggregation were observed after the second one. Finally, a significant inhibition of platelet aggregation induced by all the agonists was observed at the last dosing step. Coagulation time remained unchanged during the trial.     

β-丁香烯代谢产物和结构改造的研究

从β-丁香烯代谢产物中分离得到一种有药理活性的化合物,经光谱分析及化学反应证明其结构为β-丁香酮(8)。根据代谢产物的化学结构以及构-效关系理论,对β-丁香烯进行了结构改造研究,结果得到一种活性较强的半合成产物,为无定形长丝状物,其结构被鉴定为β-丁香烯醇(3)。药理实验表明化合物(8)及(3)对豚鼠离体气管平滑肌均有较强的松弛作用,化合物(3)还能明显抑制组织胺和乙酰胆碱诱发的豚鼠哮喘,其作用持久而毒性低,可望成为一种新型的平喘药物。     

NEURAMIDE STIMULATES ADRENOCORTICOTROPIN BUT NOT PROLACTIN RELEASE FROM RAT PITUITARY

Neuramide (NMD), a substance found in crude preparations of porcine stomach extract, is a viral inhibitor that also has putative immunostimulatory effects. The effects of NMD on stress-hormone (ACTH and prolactin—PRL) release were assessed inin vivoandin vitrostudies. In the former, blood levels of corticosterone and PRL were measured in NMD-treated male rats.In vitroexperiments were performed to evaluate the effects of NMD and three of its fractions (obtained with high performance liquid chromatography) on ACTH and PRL release from perfused rat pituitary slices. NMD increased plasma corticosterone levelsin vivoand produced dose-dependent increases inin vitropituitary release of ACTH. No effects on PRL secretion were observedin vivoorin vitro. The stimulatory effects on ACTH release were caused by the NMD fraction with a molecular weight of >5000<10000Da.     

7α-和7β-甲基-10β,17β-二乙酰氧基-△4-雌甾烯-3酮的抗早孕作用

7α-和7β-甲基-10β,17β-二乙酰氧基-△⁴-雌甾烯-3酮(简称7α-和7β-甲-乙氧雌酮)对小鼠抗早孕ED₅₀分别为1.6和5.5 mg/kg。7α-甲-乙氧雌酮在大鼠也有抗早孕作用并使血浆孕酮浓度降低,应用10 μg/ml浓度能抑制离体妊娠大鼠卵巢孕酮合成。7α-和7β-甲-乙氧雌酮与兔子宫胞浆雌二醇受体的相对结合亲和力(RBA)分别为10.8和1.5,与孕酮受体的RBA均<1.7α-和7β-甲-乙氧雌酮都有较弱的雌激素和抗雌激素活性。