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首页> 外文学位 >Effects of Vernonia amygdalina Leaf Extracts and Alkaloids on Cell Proliferation and Tubulin Assembly and Disassembly in MCF-7 Breast and DU-145 Prostate Carcinoma Cell Lines.
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Effects of Vernonia amygdalina Leaf Extracts and Alkaloids on Cell Proliferation and Tubulin Assembly and Disassembly in MCF-7 Breast and DU-145 Prostate Carcinoma Cell Lines.

机译：扁桃紫叶提取物和生物碱对MCF-7乳腺癌和DU-145前列腺癌细胞系细胞增殖和微管蛋白组装和拆卸的影响。

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摘要

Vernonia amygdalina (VA) is a tropical African plant of the Asteraceae family used as a dietary supplement and medicinal herb. In this study, liquid-liquid extraction, chromatographic and spectroscopic techniques were used to extract, fractionate and characterize the major active fractions in VA. In addition, the antiproliferative activities, cellular and molecular mechanisms of action of fractions on cancer cell lines were investigated. Immunocytochemical and fluorescence techniques were used to examine tubulin assembly, cell proliferation, apoptosis, and cancer cell death after exposure to active VA fractions. The antiproliferative and antitubulin actions of VA fractions were compared to actions of Taxol and Nocodazol.;Our results indicate that sample fractionation by solvent partitioning followed by repeated chromatographic and spectroscopic analyses revealed presence of fatty acid esters, fatty acid amide, triterpene, diterpene alcohols, and phytostanols as the major chemical groups in the aqueous and organic fractions of VA leaf extracts. Their structures were elucidated, on the basis of GC-MS data, as hexanedioic acid, bis (2-ethylhexyl) ester (2a), erucamide (2b) and squalene (2c) in the water fraction; hexadecanoic acid methyl ester (3a), 9(Z)12(Z)15(Z)-octadecatrienoic acid, methyl ester (3b) and phytol (3c) in methanol fraction; phytol (4a), squalene (4b) and 7,22-ergostadienol (4c) in the petroleum ether fraction. Acid-base extraction of alkaloids and FPLC-purification of protein fractions revealed Topotecan, and fractions A, B, C, and D, respectively.;In vitro evaluation of anti-proliferative activities of sample extracts on human estrogen-receptor negative breast cancer (MDA-MB-231), estrogen receptor positive breast cancer (MCF-7), and androgen-independent prostate cancer (DU-145) cell lines revealed significant growth inhibition by organic fractions (P0.001) compared to aqueous fraction (P0.05). Alkaloidal compound, Topotecan, also induced significant growth inhibition on DU-145 (IC50 2.512e-007g/mL), and MCF-7 (P0.0001) cells. Immunocytochemical analysis of tubulin design indicates that organic and aqueous fractions, as well as Topotecan abrogated the steady state-microtubule pattern to a defused disassembled form normally characterized with Nocodazol treatment. In contrast, Taxol induced aggregation and crystallization of tubulin. Further examination of cellular mechanism of action reveals dose-related induction of apoptotic and necrotic deaths in MCF-7 and DU-145 cells treated with alkaloidal, aqueous and organic fractions of VA extracts, with greater quantities of apoptotic phenotypes observed in MCF-7 than in DU-145 cells. FPLC-purified VA protein fractions also induced apoptotic death, with significant effects observed for fractions A and B (47%) compared to fractions C and D (41% and 4%), in MCF-7 cells. Manifestations of apoptosis were marked by membrane blebs, cell shrinkage, nuclear fragmentation, chromatin condensation, DNA fragmentation, and formation of apoptotic bodies. Taken together, these observations demonstrate that VA contains biologically active components capable of inhibiting growth and proliferation of cancer cells; and that these active constituents exert their antineoplastic properties via mechanisms that target and trigger disruption of microtubule organization, effectively causing apoptotic and necrotic cell death. Commercially available Topotecan, a prominent alkaloid in the VA fraction, proved to be a potent antiproliferative and anticancer agent with antitubulin and apoptotic-inductive actions similar to Nocodazol and Taxol.
机译：紫花杏（Vernonia amygdalina）（VA）是菊科（Asteraceae）的一种热带非洲植物，用作膳食补充剂和药用植物。在这项研究中，使用液-液萃取，色谱和光谱技术来提取，分级和表征VA中的主要活性成分。另外，还研究了级分对癌细胞系的抗增殖活性，细胞和分子作用机理。免疫细胞化学和荧光技术被用来检查微管蛋白组装，细胞增殖，细胞凋亡和暴露于活性VA级分后的癌细胞死亡。将VA馏分的抗增殖和抗微管蛋白作用与紫杉醇和Nocodazol的作用进行了比较;我们的结果表明，通过溶剂分配对样品进行分级分离，然后进行反复的色谱和光谱分析，发现存在脂肪酸酯，脂肪酸酰胺，三萜烯，二萜醇，和植物甾烷醇是VA叶提取物中水和有机部分的主要化学基团。根据GC-MS数据阐明了其结构，其中的水部分为己二酸，双（2-乙基己基）酯（2a），芥酸酰胺（2b）和角鲨烯（2c）；十六烷酸甲酯（3a），9（Z）12（Z）15（Z）-十八碳三烯酸，甲酯（3b）和植醇（3c）在甲醇馏分中;石油醚馏分中的植醇（4a），角鲨烯（4b）和7,22-麦角甾烷二烯醇（4c）。生物碱的酸碱提取和蛋白质部分的FPLC纯化分别显示托泊替康以及部分A，B，C和D .;样品提取物对人雌激素受体阴性乳腺癌的抗增殖活性的体外评估（ MDA-MB-231），雌激素受体阳性乳腺癌（MCF-7）和非雄激素非依赖性前列腺癌（DU-145）细胞系显示，与水性成分（P <0.001）相比，有机成分具有显着的生长抑制作用（P <0.001）。 0.05）。生物碱化合物Topotecan也对DU-145（IC50 2.512e-007g / mL）和MCF-7（P <0.0001）细胞具有显着的生长抑制作用。微管蛋白设计的免疫细胞化学分析表明，有机物和水相部分以及托泊替康将稳态微管模式废除为通常以Nocodazol处理为特征的解散分解形式。相反，紫杉酚诱导微管蛋白的聚集和结晶。进一步研究细胞的作用机制，发现用生物碱，水和有机成分的VA提取物处理的MCF-7和DU-145细胞与剂量相关地诱导了凋亡和坏死性死亡，在MCF-7中观察到的凋亡表型数量更多。在DU-145细胞中FPLC纯化的VA蛋白组分也诱导了细胞凋亡，在MCF-7细胞中，与组分C和D（41％和4％）相比，组分A和B（47％）具有显着效果。细胞凋亡表现为膜泡，细胞收缩，核碎裂，染色质浓缩，DNA碎裂和凋亡小体形成。综上所述，这些观察结果表明，VA含有能够抑制癌细胞生长和增殖的生物活性成分。并且这些活性成分通过靶向并触发微管组织破坏的机制发挥其抗肿瘤作用，从而有效地导致凋亡和坏死性细胞死亡。商购的Topotecan是VA组分中的突出生物碱，被证明是一种有效的抗增殖和抗癌剂，具有类似于Nocodazol和Taxol的抗微管蛋白和凋亡诱导作用。

著录项

  • 作者

    Oyugi, Daniel Arap.;

  • 作者单位

    Howard University.;

  • 授予单位 Howard University.;
  • 学科 Biology Cell.;Chemistry Organic.;Health Sciences Oncology.;Chemistry Analytical.
  • 学位 Ph.D.
  • 年度 2012
  • 页码 159 p.
  • 总页数 159
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

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