【作者】 王璇；

【导师】 王晓岚；

【作者基本信息】 江南大学 ， 营养与食品卫生学， 2009， 硕士

【摘要】 目的:采用纳米技术制备番茄红素纳米分散体并研究其体内外抗肝肿瘤作用及初步探讨作用机制,为开发纳米级别番茄红素抗肿瘤功能性食品提供理论依据。方法:参照乳化-蒸发工艺,制备番茄红素纳米分散体。采用分光光度法研究番茄红素纳米分散体体外清除活性氧能力。建立小鼠H22肝癌移植瘤模型,结合体内抗氧化实验、病理技术、免疫组化方法研究番茄红素纳米分散体体内抗肿瘤功效及初步探讨机制。体外培养人肝癌HepG2细胞和人肝L-02细胞,结合四甲基偶氮唑盐(MTT)法、生化检测、免疫组化方法等研究番茄红素纳米分散体对人肝癌HepG2细胞细胞增殖的影响及初步探讨机制。结果:制备的番茄红素纳米分散体Z-均粒径为261nm,水中分散性好。体外抗氧化结果表明番茄红素纳米分散体对活性氧(过氧化氢、羟自由基)具有明显的清除效果,相同浓度下番茄红素纳米分散体的清除率明显高于番茄红素四氢呋喃溶液。荷瘤小鼠抗肿瘤实验结果显示番茄红素纳米分散体对小鼠H22肝癌移植瘤具有显著的体内抗肿瘤作用,1.95、3.9mg·kg-1剂量番茄红素纳米分散体处理后抑瘤率分别为32.37%、54.34%,均比3.9mg·kg-1剂量油溶番茄红素组处理后抑瘤率高;体内抗氧化实验结果表明番茄红素纳米分散体可显著提高荷瘤小鼠血清GSH-Px活性,显著降低血清MDA含量,同时可显著提高小鼠肝组织T-AOC、SOD活性,显著降低肝组织H2O2、MDA含量;病理检测结果表明用药组肿瘤组织周边结缔组织炎细胞浸润;免疫组化结果显示番茄红素纳米分散体处理荷瘤小鼠后,肿瘤细胞PCNA表达水平明显下调。体外细胞培养实验,MTT结果显示番茄红素纳米分散体和空白乳化剂对人肝癌细胞都有相对选择性毒性作用。检测不同制剂作用HepG2细胞24h后细胞上清液LDH活性,结果显示番茄红素纳米分散体和乳化剂通过影响细胞膜通透性,使细胞受损;免疫组化结果显示2μmol/L番茄红素纳米分散体处理细胞24h后PCNA表达明显下降,而乳化剂并不影响PCNA的表达。结论:番茄红素纳米分散体体内外均具有较强抗肝肿瘤作用,其作用机制可能与提高体内抗氧化能力、降低PCNA表达、直接损伤肿瘤细胞等有关。更多还原

【Abstract】 Objective: To prepare lycopene nanodispersion and evaluate the anti-hepatoma effect of lycopene nanodispersion in vitro and in vivo and its mechanism.Methods: Lycopene nanodispersion was prepared by the emulsification-evaporation process. Scavenging effects of lycopene nanodispersion on reactive oxygen species (·OH and H2O2) were investigated by spectrophotometric methods in vitro. Mice transplanted with H22 liver tumor were established and antitumor effect of lycopene naonodispersion in vivo was analyzed. Study of antioxidant activity, pathological technology and immunohistochemical technology were utilized to detect the antitumor mechanism in vivo. The MTT assay was used to examine the proliferation arrest of carcinoma cell line (HepG2) and normal cell line (L-02) in vitro. And cell morphological observation, biochemical test and immunohistochemical technology were used to detect the antitumor mechanism in vitro.Results: Lycopene nanodispersion was obtained and the particle size was 261nm. The dispersive property was effectively improved. Lycopene nanodispersion had strong scavenging effects in both radical systems and the anti-radical activities. At the same concentration, the scavenging capabilities of lycopene nanodispersion were stronger than the lycopene dissolved in THF. Mice transplanted with H22 liver tumor were treated with 1.95 mg·kg-1and 3.9 mg·kg-1 lycopene nanodispersion for 10 days respectively, the tumor growth inhibition rate were 32.37% and 54.34%, which were higher than the inhibiton rate caused by 3.9 mg·kg-1 unprocessed lycopene. Lycopene nanodispersion could significantly increase the activities of GSH-Px, and decrese the content of MDA of blood serum of mice. It also could significantly increase the activities of T-AOC and SOD activity, and the reduction of the content of MDA and H2O2 of liver of mice. The results of pathological technology demonstrated that inflammatory cells infiltrated in the surrounding connective tissue of tumors. Lycopene nanodispersion could down-regulate the expression of proliferating cell nuclear antigen (PCNA) in Hepatocarcinoma H22. The MTT assay showed that lycopene nanodispersion had the selective toxicity on HepG2 cells. Tween-20, as emulsifier for lycopene nanodispersion, also had the same selective toxicity. The activities of LDH in HepG2 cells culture supernatant significantly increased after 24h exposure to lycopene nanodispersion and emulsifier, which indicated that they may induce some damages on HepG2 cells membrane. Lycopene nanodispersion could down-regulate the expression of PCNA in HepG2 cells and tween-20 didn’t have the same effect.Conclusion: Lycopene nanodispersion was able to inhibit potently the gowth of hepatoma in vitro and in vivo. The mechanism may be associated with increase of anti-oxidation activity, down regulation PCNA expression and direct cytotoxicity.更多还原