小檗碱合用厚朴酚类对THP-1泡沫细胞胆固醇外流的影响

目的 研究小檗碱合用厚朴酚类作用于THP-1源性泡沫细胞后对细胞内胆固醇含量及ABCAl mRNA表达的影响.方法 采用HPLC-MS法测定总胆固酵,抗干扰Lowery法测定总蛋白质,以油红O染色法镜下现察泡沫化程度,实时荧光定量RT-PCR法检测ABCAl mRNA的表达.结果 小檗碱合用厚朴酚类作用于泡沫细胞后,细胞内总胆固醇明显减少,总蛋白质明显降低,泡沫化程度减轻,ABCA1 mRNA表达升高,是对照组近2倍,是模型组近5倍.结论 小檗碱合用厚朴酚类通过上调ABCA1表达促进细胞内胆固醇外流.     

HPLC测定不同来源姜制厚朴中厚朴酚及和厚朴酚的含量

目的比较按国标和广东地标生产的姜制厚朴中厚朴酚与和厚朴酚的含量差异。方法采用HPLC测定不同来源姜制厚朴中厚朴酚及和厚朴酚的含量。结果厚朴酚质量浓度在1.38~69.00μg.mL-1与峰面积有良好线性关系,r=0.9996,平均回收率为100.66%,RSD=1.15%(n=5);和厚朴酚质量浓度在0.49~24.40μg.mL-1与峰面积有良好线性关系,r=0.9998,平均回收率为100.04%,RSD=1.05%(n=5)。结论国标和广东地标姜制厚朴中厚朴酚与和厚朴酚总含量总体差别不大,各地姜制厚朴质量差别大。     

炮制对厚朴饮片中厚朴酚、和厚朴酚含量变化规律的影响

目的:采用高效液相法对湖北恩施GAP示范基地厚朴炮制前后的酚性活性成分的含量进行测定,探讨其含量的变化规律;同时与不同产地来源的厚朴饮片的厚朴酚、和厚朴酚的含量进行比较。方法:应用HPLC测定厚朴中酚性成分的含量。结果:①总结厚朴炮制前后厚朴酚、和厚朴酚的变化规律,GAP示范基地的厚朴经姜炙后其厚朴酚、和厚朴酚的含量都明显增加,和厚朴酚增加了约40%,厚朴酚增加了约140%。②不同产地厚朴饮片中其酚性成分的含量各不相同,GAP示范基地的厚朴饮片中其厚朴酚、和厚朴酚的含量相对最高,湖北房县厚朴中含量最低。结论:此方法准确,操作简单,能快速有效的测定厚朴饮片中厚朴酚、和厚朴酚的含量。     

厚朴不同炮制品中厚朴酚与厚朴酚的气相色谱测定

采用气相色谱法对厚朴生品及１１种不同制品中厚朴酚与和厚朴酚进行含量测定，实验表明不同炮制品中厚朴酚与和厚朴酚的含量各异，以生品含量最高，炮制后均有所下降。     

厚朴酚与和厚朴酚抗鼻咽癌作用机制研究

目的探讨厚朴中小分子化合物厚朴酚与和厚朴酚对鼻咽癌细胞增殖、迁移、侵袭的影响。方法分别采用MTT比色法、细胞划痕实验、Transwell实验,观察厚朴酚与和厚朴酚对鼻咽癌细胞HONE1增殖、迁移、侵袭的影响。结果厚朴酚与和厚朴酚对鼻咽癌细胞HONE1有较显著的抑制增殖、迁移和侵袭效应,而且这种效应呈显著的剂量依赖性。结论厚朴酚与和厚朴酚能够抑制鼻咽癌肿瘤细胞增殖、迁移和侵袭,厚朴酚与和厚朴酚是具有开发潜力的抗癌小分子化合物。     

微波辅助提取厚朴中厚朴酚及和厚朴酚的研究

目的:进行微波辅助提取厚朴中厚朴酚及和厚朴酚的研究.方法:采用正交实验用微波辅助进行提取方法的研究;用HPLC法测定厚朴酚及和厚朴酚含量,流动相:甲醇-水-冰醋酸(85∶15∶0.25),流速为1.0 ml/min,检测波长为294nm,柱温为23℃.结果:用中等强度的微波功率,酒精度75%,提取时间为4min,料液比1∶40的提取条件可很好地提取厚朴药材中的厚朴酚及和厚朴酚,和厚朴酚、厚朴酚以及总酚的提取率分别为92.33%、73.59%和83.32%;RP-HPLC方法较好,结果理想.结论:本研究得到了微波辅助提取厚朴中厚朴酚及和厚朴酚的方法.     

HPLC法测定厚朴制剂中厚朴酚与和厚朴酚的含量

建立一种采用HPLC法测定厚朴中厚朴酚与和厚朴酚含量的测定方法。结果显示,本测定方法简便、灵敏、准确、重现性好,可以作为含有厚朴制剂的定量方法,可用于有关制剂的质量控制。所测样品中的厚朴酚、和厚朴酚的含量分别为0.60％和0.21%。     

厚朴中厚朴酚与和厚朴酚总量测定方法的简化实验

厚朴中厚朴酚与和厚朴酚总量测定方法的简化实验孙立华曹志胜李欣荣(青岛市药品检验所青岛266071)孙国才(青岛市卫生学校青岛266071)《中国药典》1985～1995年版中收载的厚朴酚、和厚朴酚总量的测定方法,是以相同的方法分别测定了厚朴酚、和厚朴酚两种成分的含量,然后将两者的量加起来即为总量。本文在原药典法的基础上,不需将两者分离,直接测定两者的总吸收度,求知总量。1测定方法精密量取供试品溶液50μl(根皮25μl),刮取厚朴....     

丹皮酚对RAW264.7巨噬细胞源性泡沫细胞形成的影响

目的探讨丹皮酚对RAW264.7巨噬细胞源性泡沫细胞形成的影响。方法以氧化型低密度脂蛋白诱导RAW264.7巨噬细胞泡沫化为模型,油红O染色检测细胞内脂质,荧光探针Dil标记氧化低密度脂蛋白(oxidized low-densitylipoprotein,oxLDL)并结合高内涵筛选成像系统检测oxLDL摄取,实时荧光定量PCR检测CD36和清道夫受体A(scaven-ger receptor A,SR-A)表达。结果成功建立了RAW264.7巨噬细胞源性泡沫细胞模型,经不同浓度丹皮酚作用后,细胞内脂质减少,oxLDL摄取下降,SR-A基因表达下降,但CD36基因表达无显著改变。结论丹皮酚可抑制RAW264.7巨噬细胞源性泡沫细胞形成,其作用机制可能与下调SR-A表达有关。     

厚朴温中丸中厚朴酚与和厚朴酚含量测定的研究

建立了厚朴温中丸中厚朴酚与和厚朴酚的含量测定方法。方法：采用HPLC测定厚朴酚与和厚朴酚的含量。结果：厚朴酚与厚朴酚含量测定方法的线性范围分别为0.488-2.440μg（Ch i-Square：27.44）和0.362-1.810μg（Ch i-Square：36.07）,平均回收率分别为97.0%,RSD=1.3%（n=5）和96.8%,RSD=1.4%（n=5）。结论：所建立的方法准确可行,重复性好,可作为厚朴温中丸中控制厚朴含量的质量标准。     

清热、活血类中药有效成分对THP-1巨噬泡沫细胞形成及分泌IL-1β、TNF-α的影响

目的探讨清热、活血类中药有效成分抑制巨噬细胞泡沫化的作用及机制。方法以氧化型低密度脂蛋白(ox-LDL)和细菌脂多糖(LPS)诱导THP-1细胞成为巨噬泡沫细胞,采用油红O染色脂质半定量分析法和ELISA法检测中药有效成分对THP-1巨噬泡沫细胞形成及培养上清中白细胞介素1β(IL-1β)和肿瘤坏死因子α(TNF-α)含量的影响。结果20μmol/L的黄连素、黄芩苷、大黄素、丹参酮ⅡA和100 mg/L的三七总皂苷可显著降低THP-1巨噬泡沫细胞油红O染色阳性率;20μmol/L的大黄素、丹参酮Ⅰ、丹参酮ⅡA可显著降低THP-1巨噬细胞的IL-1β分泌量;20μmol/L的黄连素可显著降低THP-1巨噬细胞的TNF-α分泌量。结论清热、活血类中药有效成分可抑制巨噬细胞泡沫化及其炎症因子IL-1β和/或TNF-α的分泌,抑制炎症因子的释放可能是其抗泡沫细胞形成的机制之一。     

Aged garlic extract inhibits homocysteine-induced scavenger receptor CD36 expression and oxidized low-density lipoprotein cholesterol uptake in human macrophages in vitro

Aim of the study

Expression of CD36 scavenger receptors on macrophages is involved in oxidized low-density lipoprotein (OxLDL) uptake and foam cell formation during atherosclerotic lesion development. It has been shown previously in vitro and in vivo that the atherosclerotic risk factor homocysteine (Hcy) stimulates macrophage CD36 expression and OxLDL uptake. We now examined the effects of aged garlic extract (AGE), a garlic preparation enriched in water-soluble organic sulfur-containing compounds, on Hcy-induced CD36 expression and foam cell formation in human monocytes/macrophages.

Results

Incubation with Hcy (200 μM for 72 h) in THP-1-derived macrophages and primary human macrophages caused a 37.8 ± 5.2% and 60.7 ± 4.2% increase in CD36 expression compared to control, respectively. Coincubation with AGE (5 mg/ml) significantly suppressed CD36 expression in THP-1 derived macrophages by 48.6 ± 9.0% compared to Hcy-incubated cells only. AGE (1-5 mg/ml) dose dependently inhibited Hcy-induced CD36 expression in primary human macrophages, and decreased binding of nuclear proteins to a PPARγ response element. Preincubation with AGE significantly inhibited DiI-labeled OxLDL uptake.

Conclusions

These data indicate that AGE inhibits CD36 expression and OxLDL uptake in human macrophages by modulating the PPARγ pathway, and suggest that the extract could be useful for the prevention of atherosclerotic lesions.     

化瘀祛痰方药对巨噬细胞源性泡沫细胞ABCA1、CD36表达的影响

目的:观察化瘀祛痰方药及其拆方对THP-1源性泡沫细胞腺苷三磷酸结合盒转运体A1(ABCA1)、CD36表达的影响,探讨该方药的作用机制,并比较方药全方及各拆方的疗效及调控作用。方法:体外培养THP-1细胞,佛波酯(PMA)使之巨噬化,氧化型低密度脂蛋白(ox-LDL)使巨噬细胞泡沫化。细胞随机分为空白对照组、模型组、补气组、化瘀组、祛痰组和全方组。油红O染色观察细胞浆脂滴变化,酶法测定细胞内总胆固醇(TC)、游离胆固醇(FC)、胆固醇酯(CE)的含量,并计算CE/TC值。反转录-聚合酶链反应(RT-PCR)定量分析ABCA1、CD36 mRNA水平。结果:模型组细胞TC、FC、CE含量及CE/TC值均显著高于对照组(P<0.01);与模型组相比,全方组和祛痰组细胞TC、FC、CE含量及CE/TC值均显著降低。RT-PCR结果提示模型组细胞ABCA1、CD36 mRNA水平显著升高;而化瘀祛痰方药及祛痰拆方含药血清可显著升高ABCA1 mRNA,显著降低CD36 mRNA水平。结论:化瘀祛痰方药及祛痰方可抑制ox-LDL诱导的THP-1细胞泡沫化,该作用可能与其上调ABCA1基因、下调CD36基因表达,从而减弱THP-1细胞对ox-LDL摄取有关。     

Aged garlic extract inhibits CD36 expression in human macrophages via modulation of the PPARγ pathway

Expression of CD36 scavenger receptors on macrophages is involved in oxidized low‐density lipoprotein uptake and foam cell formation during atherosclerotic lesion development. We examined the effects of aged garlic extract (AGE), a garlic preparation enriched in water‐soluble cysteinyl moieties that increases cellular total thiols and glutathione concentrations, on CD36 expression in human monocytes/macrophages (THP‐1 cells and primary human monocytes). Compared to control, AGE (1–5 mg/mL) dose‐dependently and significantly suppressed CD36 expression up to by 61.8 ± 7.4% in THP‐1‐derived macrophages and up to 50.5 ± 7.1% in primary human macrophages, respectively. Furthermore, AGE prevented induction of CD36 expression by the peroxisome proliferator activator receptor (PPAR) γ agonist troglitazone, and decreased binding of nuclear proteins to a PPARγ response element. AGE showed a stronger inhibitory effect on CD36 expression in THP‐1 cells during simultaneous incubation with phorbol 12‐myristate 13‐acetate (PMA) compared to cells that had been pre‐incubated with PMA. Furthermore, AGE decreased CD11b expression in a dose‐dependent manner. These data indicate that AGE inhibits CD36 expression by modulating the PPARγ pathway in human macrophages and monocytes differentiation into macrophages, and suggests that the extract could be useful for the prevention of atherosclerotic lesions. Copyright © 2010 John Wiley & Sons, Ltd.     

三重四级杆串联质谱(LC-MS/MS)法同时测定复方广痛消水提物中5种有效成分的含量

目的:建立同时测定广痛消水提物中延胡索乙素、芍药苷、小檗碱、厚朴酚、和厚朴酚含量的方法.方法:采用LC-MS/MS联用方法.结果:芍药苷、延胡索乙素、小檗碱、厚朴酚、和厚朴酚线性范围分别为0.882 8 ～ 113(r=0.999 8),1.5438～49.4(r =0.999 5),0.904 7～115.8(r=0.9997),0.8813～112.8 (r=0.9994),0.1656～21.2mg.L-1(r=0.9999).加样回收率分别为1.998％,1.14％,3.117％,1.828％,1.596％.结论:该法专属性强、快速灵敏,可用于广痛消水提物中延胡索乙素、芍药苷、小檗碱、厚朴酚、和厚朴酚的含量测定.     

艳山姜挥发油通过调控巨噬细胞CD36和ABCA1表达抑制泡沫细胞的形成

目的:观察艳山姜挥发油对氧化低密度脂蛋白(ox-LDL)诱导巨噬细胞转化为泡沫细胞的抑制作用,并探索其机制。方法:使用佛波酯(PMA,100μg·L^-1)诱导人白血病单核细胞(THP-1)24 h后形成巨噬细胞,实验分为4组,分别为空白组(无血清RPMI 1640),模型组(80 mg·L^-1ox-LDL),艳山姜挥发油低剂量组(80 mg·L^-1ox-LDL+4μg·L^-1艳山姜挥发油),艳山姜挥发油高剂量组(80 g·L^-1ox-LDL+20μg·L^-1艳山姜挥发油)。噻唑蓝(MTT)比色法检测艳山姜挥发油对巨噬细胞的活性的影响,蛋白免疫印迹法(Western blot)检测巨噬细胞中白细胞分化抗原36(CD36)和三磷酸腺苷结合盒转运体A1(ABCA1)的表达,酶联免疫吸附测定(ELISA)检测巨噬细胞内胆固醇酯含量,油红O染色法检测巨噬细胞中脂质小滴的含量。结果:艳山姜挥发油对巨噬细胞无毒性。与空白组比较,模型组的巨噬细胞内脂滴和胆固醇酯的含量显著增加(P<0.01),CD36蛋白表达显著上升(P<0.01),ABCA1蛋白表达无显著变化;与模型组比较,艳山姜挥发油显著抑制巨噬细胞中脂滴和胆固醇酯的含量(P<0.01),下调CD36的蛋白表达(P<0.01),上调ABCA1蛋白的表达(P<0.01),艳山姜挥发油可抑制巨噬细胞向泡沫细胞的转化。结论:艳山姜挥发油对ox-LDL诱导的巨噬细胞向泡沫细胞的形成具有抑制作用,该药理作用与艳山姜挥发油下调巨噬细胞CD36和上调ABCA1蛋白的表达有关。     

猫耳刺皂苷ilexpernoside C抑制低密度脂蛋白多聚体诱导泡沫细胞形成的活性及机制研究

巨噬细胞来源的泡沫细胞在血管壁上的堆积,是动脉粥样硬化的主要成因。该实验构建了低密度脂蛋白多聚体(LDL aggregates)诱导的泡沫细胞模型,对冬青属植物猫耳刺分离的化合物进行体外活性研究,发现ilexpernoside C(IC1)能显著抑制泡沫细胞的形成。通过对泡沫细胞形成相关受体的分析发现,IC1能显著抑制低密度脂蛋白相关受体1(LRP1)的表达。因此,该研究认为皂苷IC1通过抑制巨噬细胞的胞吞作用,从而抑制泡沫细胞的形成,使其可能成为一种抗动脉粥样硬化的潜在药物。     

Application of the elevated plus-maze test in mice for evaluation of the content of honokiol in water extracts of magnolia.

In our previous study using an improved elevated plus-maze in mice, the oriental herbal medicine Saiboku-to prolonged the time spent in open arms, showing an anxiolytic effect, and the effect was mainly caused by honokiol derived from magnolia. This study was carried out to compare the anxiolytic potentials of honokiol and water extracts of three magnolia samples; two being Kara-koboku (Magnolia officinalis) (KA: from Zhejiang-sheng, China; honokiol 0.25% and magnolol 1.16%, and KB: from Sichuan-sheng, China; honokiol 1.72% and magnolol 1.71%), and one being Wa-koboku (Magnolia obovata) (WA: from Iwate-ken, Japan; honokiol 0.32% and magnolol 0.81%). Seven daily treatments with 0.1-1 mg/kg honokiol, but not 0.2 and 1 mg/kg magnolol, revealed an anxiolytic effect with the peak potential at 0. 2 mg/kg. The anxiolytic potentials of 40 and 80 mg/kg KA, which contained the highest amount of magnolol, were almost equivalent to those of 0.1 and 0.2 mg/kg honokiol, respectively. KB, at 11.6 mg/kg, and 62.5 mg/kg WA resulted in almost the same anxiolytic potential as that of 0.2 mg/kg honokiol. No significant change in the ambulatory activity was produced by any drug treatment. These results suggest that honokiol is the chemical responsible for the anxiolytic effect of the water extract of magnolia and that the other chemicals including magnolol in magnolia scarcely influence the effect of honokiol. It is also considered that the elevated plus-maze test is applicable for evaluation of the content of honokiol in magnolia.     

黄连素对糖尿病大鼠坐骨神经损伤的保护作用

目的通过观察黄连素对糖尿病模型大鼠痛阈、坐骨神经传导速度(NCV)和脊髓组织病理学及血清、坐骨神经内白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)、高敏C反应蛋白(hs-CRP)等炎症因子的影响,探讨黄连素对糖尿病大鼠坐骨神经损伤保护作用的机制。方法以链脲佐菌素(STZ)腹腔注射诱导形成糖尿病周围神经病变(DPN)模型,分别以100 mg/kg或187.5 mg/kg黄连素灌胃治疗,每天1次,持续8周。采用酶联免疫吸附测定(ELISA)法分别测定各组大鼠血清及坐骨神经内IL-1β、TNF-α、hs-CRP的浓度。结果与对照组比较,DPN模型组大鼠IL-1β、TNF-α、hs-CRP含量显著增高(P均0.05),黄连素治疗8周后,与DPN模型组比较IL-1β、TNF-α、hs-CRP含量明显降低(P均0.05)。结论黄连素对DPN大鼠痛阈、NCV有明显改善,抗炎作用可能是黄连素对糖尿病大鼠坐骨神经损伤保护作用的机制之一。