Anti-tumor molecular targets of intestinal metabolite IH-901 of Ginseng saponin in human hepatocellular carcinoma cells

摘要肝癌（hetatocellularcarcinoma，HCC）是一种恶性程度高、浸润和转移性强，治疗难度大的恶性肿瘤，至今仍无有效抗肝癌药物。人参（PanaxginsengC.A.Meyer）是我国传统名贵中药，具有明显抗肿瘤效果，其主要活性成分为人参皂甙。最新药理学研究表明，人参发挥抗肿瘤作用的最终活性物质不是天然人参皂甙，而是其一系列肠道菌代谢产物。其中，人参皂甙肠道菌代谢物20-O-β-D-吡喃葡萄糖苷-20(S)-原人参二醇（简称IH-901，M1或CompoundK）为天然二醇组人参皂甙肠道细菌最终代谢产物，是人参皂甙抗肿瘤作用的最终主要活性形式之一。然而，迄今为止，有关IH-9...Abstract Hepatocellular carcinoma (HCC) is a common malignant tumor which threatens seriously human life and health. As HCC has a high degree of malignancy and is easy to metastasize and invade, there are still no effective anti-hepatoma agents so far. Panax ginseng C. A. Meyer, a rare Chinese traditional medicine, has an obvious action to anti-tumor, and its main effective component is gi...学位：理学博士院系专业：生命科学学院生物医学科学系_细胞生物学学号：B20042601

Microbial Transformation of Gypenoside from Gynostemma pentaphyllum by Aspergullus glaucus and Its Biological Activities

以福建绞股蓝[Gynostemma pentaphyllum(Thunb)Makino]为材料,经乙醇提取和正丁醇萃取获得绞股蓝皂苷,利用灰绿曲霉(Aspergullus glaucus)微生物转化绞股蓝皂苷,通过测定绞股蓝皂苷和灰绿曲霉转化产物的抗癌、抗酪氨酸酶和抗氧化活性,比较绞股蓝皂苷经微生物转化修饰前后生物活性的差异.结果表明:灰绿曲霉转化产物对肝癌细胞SMMC7721有体外抑制作用,半抑制质量浓度(IC50)为91.66μg/m L,而绞股蓝皂苷抗癌效果不强;绞股蓝皂苷对蘑菇酪氨酸酶活力具有较强的抑制作用,IC50为0.14 mg/m L,其抑制作用属于可逆抑制,抑制类型为混合型抑制作用,而灰绿曲霉转化产物抗蘑菇酪氨酸酶活性较弱;绞股蓝皂苷和灰绿曲霉转化产物分别在0.2和2 mg/m L质量浓度下对DNA氧化损伤有保护作用.研究结果说明绞股蓝皂苷经微生物修饰后,抗癌效果增强,对蘑菇酪氨酸酶抑制作用和DNA氧化损伤保护作用减弱.该研究结果可为利用微生物转化法筛选抗癌绞股蓝皂苷奠定基础.Gynostemma pentaphyllum in Fujian Province was used as the material in this research.Gypenoside was extracted with ethanol and n-butyl alcohol. Gypenoside was microbially transformed by Aspergullus glaucus. The abilities of anticancer,anti-tyrosinase and anti-oxidation in gypenoside and transformation products were measured in our study.The difference of bioactivity between gypenoside and microbially transformed gypenoside was compared.The result showed that the products of transformation by A. glaucus affected the growth of hepatocellular carcinoma cells SMMC7721 with half maximal inhibitory concentration( IC50) values of 91. 66 μg/m L,but the anticancer activity of gypenoside was very weak.Gypenoside had strong inhibition to the mushroom tyrosinase. The value of IC50 was 0. 14 mg / m L. Moreover,the products of transformation by A. glaucus showed little anti-tyrosinase acti-vity.The kinesis study showed that the inhibition of gypenoside to mushroom tyrosinase was reversible and inhibition types were mixed. Gypenoside exhibited the active protective effect on H2O2-induced oxidative-stress damage at the concentration of 0. 2 mg / m L,and the products of transformation by A. glaucus were at 2 mg / m L. This research indicated that the anticancer effect of gypenoside modified by microorganisms was enhanced,but the activity of anti-tyrosinase and H2O2-induced oxidativestress damage became weakened.This study laid the foundation of screening anticancer gypenoside by means of microbial transformation.国家自然科学基金(81274149);; 厦门市科技创新项目(3502Z20132009,3502Z20154083);; 厦门市对台科技合作项目(3502Z20141041

Apoptosis induced by a novel intestinal metabolite of ginseng saponin in human hepatocellular carcinoma BEL-7402 cells

目的探讨一种新的人参稀有皂苷20-O-(β-D-吡喃葡萄糖)-20(S)-原人参二醇皂苷(IH-901)对人肝癌BEL-7402细胞的增殖抑制效果和诱导凋亡作用。方法以0、6.25、12.5、25、50、75、100μmol/L的IH-901作用于体外培养的BEL-7402细胞,通过MTT法检测IH-901对细胞生长的抑制作用,相差显微镜观察细胞形态变化,AO/EB的荧光核染色观察凋亡细胞形态,流式细胞术检测细胞周期和细胞凋亡。结果MTT实验显示IH-901抑制BEL-7402细胞的生长,呈时间及浓度依赖的方式;显微镜下观察到典型的凋亡形态变化和生化特征:细胞固缩,核染色质凝聚,出现凋亡小体;AO/EB的荧光核染色后,观察到典型的凋亡细胞;流式细胞仪可以检测到凋亡峰,且细胞被阻滞在G0/G1期。结论IH-901能通过诱导细胞凋亡的方式有效地抑制人肝癌BEL-7402细胞的生长,稀有人参皂苷IH-901可能成为一种潜在的抗肝癌药物。Objective To investigate the effects of 20-O-(β-D-glucopyranosyl)-20(S)-protopanaxadiol(IH-901) on the human hepatocellular carcinma BEL-7402 cells in terms of inhibition of proliferation and induction of apoptosis.Methods BEL-7402 Cells in cultural medium in vitro were given 0,6.25,12.5,25,50,75,and 100 μmol/L IH-901.The inhibitory rate of cells was measured by MTT assay,morphology of cell apoptosis was observed by AO/EB fluorescent staining,cell apoptotic rate and cell phase were detected by flow Cytometry(FCM).Results The results indicated that IH-901 could inhibit the proliferation of BEL-7402 cells significantly.The suppression was both in a time-and dose-dependent manner.Morphological examination of IH-901-treated samples showed cells with chromatin condensation,cell shrinkage,and all typical characteristics of apoptotic cells.Marked morphological changes of cell apoptosis was observed very clearly by AO/EB fluorescent staining.The increase in the apoptotic sub-G1 fraction in a concentration-dependent manner and cell cycle arrest at the G0/G1 phase were detected by FCM.Conclusion The results demonstrate that IH-901 dramatically suppresses BEL-7402 cell growth by inducing programed cell death.These results may provide a pivotal mechanism for the use of IH-901 in the prevention and treatment of human hepatocellular carcinoma BEL-7402 cells,the antiproliferation,and the apoptosis厦门市科技创新基金重点项目(3502Z20062008

Promotion of proliferation of luminal B breast cancer cells by mesenchymal stem cells and its underlying molecular mechanisms

目的分析人脐带间充质干细胞(hUC-MSCs)对Luminal B型乳腺癌细胞生长增殖的影响,并初步探讨其可能的分子机理。方法绿色荧光蛋白(GFP)和荧光素酶共表达慢病毒感染人Luminal B型乳腺癌细胞BT474,并经嘌呤霉素筛选两周后,于荧光显微镜下观察GFP的表达情况,IVIS Kinetic成像系统拍照以观察和记录慢病毒感染后BT474细胞荧光素酶的表达情况;荧光显微镜下直接观察,结合MTS实验分析hUC-MSCs共培养或其浓缩上清处理对GFP和荧光素酶共表达BT474细胞生长增殖的影响;Western blot法检测hUC-MSCs浓缩上清处理对BT474细胞Akt和MAPK信号通路激活情况以及下游细胞周期调控蛋白Cyclin D1表达的影响;常规RT-PCR法检测hUC-MSCs中NRG-1、NRG-2、IGF-Ⅰ、IGF-Ⅱ和EGF等配体的表达。荧光素酶表达强度与细胞数量的相关性经由Excel软件行统计学分析,MTS实验数据则经由SPSS13.0统计软件行统计学分析。结果荧光显微镜和IVIS Kinetic成像系统的观察结果分别证实,GFP和荧光素酶经慢病毒载体系统的介导可在BT474细胞中成功地共表达,且荧光素酶的表达强度与细胞数量呈直线相关。MSCs共培养或其浓缩上清处理均可显著促进Luminal B型乳腺癌细胞BT474的生长增殖,其细胞存活比例分别为各自对照组的148.06%(P<0.005)和147.99%(P<0.001);MSCs浓缩上清处理同时激活BT474细胞内Akt和MAPK信号通路,并上调细胞周期调控蛋白Cyclin D1表达。此外,RT-PCR结果显示,hUC-MSCs中NRG-1和EGF的mRNAs水平呈高表达,而NRG-2、IGF-Ⅰ和IGF-Ⅱ等配体的mRNAs表达也可见。结论 MSCs可通过表达并分泌NRG-1等配体,从而激活Luminal B型乳腺癌细胞BT474的下游Akt和MAPK信号转导通路以上调细胞周期调控蛋白Cyclin D1的表达,进而促进其生长增殖。Objective To investigate the effect of human umbilical cord mesenchymal stem cells(hUC-MSCs) on proliferation of luminal B breast cancer cells and its underlying molecular mechanisms. Methods Human luminal B breast cancer cells BT474 were infected with GFP and luciferase co-expressing lentiviruses and then subjected for selection with Puromycin for 2 weeks. The expression of GFP and luciferase was detected by fluorescent microscopy and IVIS Kinetic image system, respectively. The effect of coculture or treatment with conditioned medium of hUCMSCs on proliferation of BT474 was analyzed with MTS assay. Western blot was carried out to detect the effect of treatment with conditioned medium of hUC-MSCs on the activation of both Akt and MAPK signalings in BT474, as well as the expression of downstream cell cycle regulator Cyclin D1. Regular RT-PCR was applied to analyze the mRNAs expression of ligands such as NRG-1, NRG-2, IGF-Ⅰ, IGF-Ⅱ, and EGF in hUC-MSCs. The correlation between relative luciferase activity and cell number was analyzed with Excel software, while MTS assay data was statistically analyzed with SPSS 13.0 software. Results The co-expression of GFP and luciferase in BT474 via lentiviral expression system was visualized by fluorescent microscopy and IVIS Kinetic image system. The linear correlation between relative luciferase activity and cell number was determined by curve fitting analysis. Coculture or treatment with conditioned medium of hUC-MSC significantly promoted the proliferation of BT474, with survival rates being 148.06 %(P < 0.005)and 147.99 %(P < 0.001)of control, respectively. In addition, treatment with conditioned medium of hUC-MSC was shown to induce activation of both Akt and MAPK signalings, which further upregulated the expression of Cyclin D1. Moreover, high mRNAs expression levels of both NRG-1 and EGF, as well as moderate mRNAs expression levels NRG-2, IGF-Ⅰ, and IGF-Ⅱ were showed by RT-PCR. Conclusion Our results here demonstrated that MSCs may promote the proliferation of luminal B breast cancer cells through paracrine of ligands such as NRG-1, which in turn results in the activation of both Akt and MAPK signalings and upregulation of the expression of Cyclin D1.国家自然科学基金面上项目(81272922);; 福建省自然科学基金面上项目(2016J01577);; 福州总医院院内课题国际合作研究专项(2015G01

药用植物对蘑菇酪氨酸酶激活作用的研究

以采集自厦门周边地区的15种药用植物为材料,研究药用植物的提取分离物对蘑菇酪氨酸酶的效应.结果表明:绞股蓝乙醇提取物、金毛狗脊乙醇提取物、爵床乙醇提取物、大青乙醇提取物、溪黄草石油醚和乙酸乙酯萃取物、千里光乙酸乙酯和正丁醇萃取物、翅柄马兰石油醚和乙酸乙酯萃取物,以及金酸枣正丁醇萃取物具有轻微激活作用,激活率分别为3.4%、20%、20%、40%、38.3%、27.6%、36%、42%、20%、50%和30%;扭序花正丁醇萃取物、野慈姑乙醇提取物、毛麝香乙醇提取物、使君子乙酸乙酯和正丁醇萃取物、冬青氯仿萃取物具有较好的激活作用,激活率分别为55%、60%、68.2%、62%、60%和80%;而金酸枣石油醚萃取物、冬青乙酸乙酯萃取物、斑鸠菊正丁醇萃取物和白面风乙醇提取物具有明显的激活作用,激活率分别为120%、150%、170%、和327%.动力学分析激活机理主要有混合性、竞争性和反竞争性3种激活类型

结肠镜对急性不全肠梗阻的诊断及治疗作用探讨

目的　探讨结肠镜在急性不全肠梗阻患者中的诊断与治疗作用。方法　对我院 1994年 5月～ 2 0 0 3年 4月共 5 933例肠镜检查中的 12 2例 (占 2 .0 6 % )急性不全肠梗阻患者进行回顾性分析 ,就其发病年龄分布、梗阻病因及内镜下诊断进行分析探讨。结果　本组 12 2例急性不全肠梗阻的原因包括肠癌、息肉、粪块、炎症性肠病等多种 ,其中结直肠癌共 5 0例 ,发病率最高 ,占 4 0 .98% ,且以左半结肠癌为主。肠粪石梗阻在高龄患者中发病率仅次于结肠癌。全部患者的肠镜检查结果中无假阴性或假阳性。同时结肠镜可解除非肿瘤病变如粪石、肠粘连等所致的肠梗阻。结论　结肠镜检查对急性不全肠梗阻有重要的诊断价值并可对治疗起积极指导作用

盐封后碳化钼电催化剂的制备及其氢析出性能（英文）

碳化钼具有低廉的价格、优越的催化性能以及良好的稳定性而被人们认为是极好的可以替代Pt等贵金属的氢析出反应（HER）催化剂。本工作采用钼酸钠和2,6-二氨基吡啶为反应原料,之后不断进行...supported by the National Key Research and Development Program of China(2017YFA0206500);; the National Natural Science Foundation of China(21773198,U1705253)~

福建绞股蓝皂苷的优化提取及其抗肝癌活性

绞股蓝素有"南方人参"美誉,含有与人参皂苷相同骨架达玛烷型结构的三萜皂苷。通过不同体积分数乙醇溶液优化提取绞股蓝皂苷,测定其抗癌活性,为后续活性化合物的分离奠定基础。利用HPLC分析绞股蓝的皂苷成分并确定提取绞股蓝皂苷的乙醇体积分数,正交实验优化提取方法,进而MTT法测定提取的绞股蓝皂苷抗肝癌活性。HPLC分析确定用体积分数75%乙醇溶液提取绞股蓝总皂苷(GYP),体积分数45%乙醇提取极性绞股蓝皂苷(GYP1),体积分数90%乙醇提取弱极性绞股蓝皂苷(GYP2);正交实验结果表明在70℃水浴中以30质量的体积分数45%乙醇加热提取12 h为最佳条件提取GYP1,在60℃水浴中以30质量的体积分数90%乙醇加热提取6 h为最佳条件提取GYP2;利用MTT法测定GYP、GYP1和GYP2对肝癌细胞的抑制作用,GYP和GYP2对肝癌细胞SMCC7721具有抗癌活性,半抑制率IC50值分别为(165.83±14.12)μg/mL和(113.97±9.26)μg/mL,GYP1没有抗癌活性,表明绞股蓝抗癌药效成分主要是由弱极性绞股蓝皂苷发挥功效

Challenge by Head Transplant

范瑞平：诸位好!受《中国医学伦理学》杂志王明旭主编所托,组织一篇＂换头术的挑战＂争鸣笔谈,特邀各位参与。请踊跃发表意见,观点不拘,长短不限,畅所欲言,各抒己见,若能针对已发观点形成争论,则更能增加读者兴趣

Experimental study on reperfusion of intraocular lens

作者简介: 祁明信, 男, 1945 年7 月 出生, 教授、主任医师、博士研究生 导师, 主要从事白内障的基础与临 床研究。联系电话: 0591-83570887; E-mail:qihuang@netease. com 通讯作者: 黄秀榕,E-mail:[email protected][中文文摘]目的开展晶状体再灌注的离体和动物实验研究,并对再灌注人工晶状体技术进行评价。方法采用新鲜离体幼兔眼、离体猪眼、新西兰白兔眼,应用自行研制的人工晶状体材料,进行以下实验:(1)体外固化实验;(2)晶状体前囊膜微型撕囊及其稳固性实验;(3)经微型前囊膜开口超声乳化吸出晶状体内容物实验;(4)活的新西兰白兔眼内人工晶状体再灌注实验。结果(1)按硅酮聚合物与固化剂50:1的比例可获得柔软、弹性好、固化时间短(完全固化时间为60min)的注入材料;(2)晶状体前囊膜1.8～2.0mm的连续环形撕囊口具有较好的稳定性,可经该微型开口吸出晶状体内容物并灌注材料;(3)超声能量18%、流量25mL·min-1、负压120mmHg(1kPa=7.5mmHg)为晶状体内容物经微型前囊膜开口吸出的最佳条件;(4)注入灌注材料后可形成由晶状体囊膜包裹的、置换原晶状体皮质和核的、新的再灌注人工晶状体。结论采用再灌注人工晶状体的方法可进行新型人工晶状体再灌注,可为治疗白内障和老视提供参考。[英文文摘]Objective To carry out the experimental study on reperfusion of intraocular lens(IOLs) in vitro or in animal,and to assess the technique of IOLs reperfusion.Methods The following experiments were performed by using self-developed materials in fresh rabbit eyes and pig eyes in vitro,as well as in eyes of alive New-Zea-land rabbits:(1)Solidification study of self-developed material in vitro;(2)Continuous circular capsulorhexis(CCC) in anterior capsule of lens and its stability;(3)Draw of lens contents via phaco through mini-CCC;(4)IOLs ref illing in the eyes of alive New-Zea land rabbits. Results(1) Thematerialwhich was soft, springy and short-term solidification(full solidification time was 60 minutes) were obtained in certain proportion of geland solidified agent(50:1) in vitro; ( 2)The CCC in anterior capsule of lens with 1.8-2.0 mm diameter had very good stability. The lens contents were drawn and the materialwere refilled through themini-CCC; (3) The best conditions of drawing out lens contents through m ini-CCC were phaco energy 18% , flow 25 mL·min- 1, and negative pressure 120 mmH g (1kPa=7.5 mmHg);(4) The new refilled IOLs, which were wrapped by capsule of lens and were replaced original cortex and nucleus of lens, were obtained after thematerial refillied. Conc lusion. New IOLs are refilled through this method, which can prov ide reference for the treatment of cataract and presbyopia.福建省科技三项费用;教育厅重点资助项目基金资助(编号:K98041