【作者】 王琪；

【导师】 许冰； 许爱娥；

【作者基本信息】 浙江大学 ， 皮肤病与性病学， 2004， 硕士

【摘要】 一、研究背景和目的 白癜风是一种获得性黑素细胞破坏所致皮肤和毛发色素脱失，其发病机制目前尚不清楚，国内外大量研究表明，该病发病机制可能与遗传、免疫、神经等方面有关。近年来越来越多研究发现在白癜风发生和发展过程中有很多细胞因子参与。黑素细胞与周围的角质形成细胞密切相邻，调节人黑素细胞的生长因子部分是由邻近角质形成细胞产生和分泌的。研究证实围绕在黑素细胞周围的角质形成细胞能分泌内皮素-1(ET-1)和碱性成纤维细胞生长因子(bFGF)，ET-1可通过受体介导的信号传导途径刺激黑素细胞的增殖，黑素生成及增加酪氨酸酶的活性。bFGF是黑素细胞生长分化的重要因子，是黑素细胞的内源性促分裂剂。因此，研究这些细胞因子对于探索白癜风的发病机制及寻找白癜风治疗的新方法具有一定意义。迄今为止，国内外尚未见有中药对角质形成细胞分泌ET-1和bFGF影响研究的报道。 本实验以门诊手术室提供的儿童包皮为材料，培养人角质形成细胞。通过观察本科室临床用于治疗白癜风的中药(加味桃红四物汤，又名皮康口服液)对角质形成细胞分泌的细胞因子ET-1和bFGF的变化，探讨皮康口服液对角质形成细胞分泌ET-1和bFGF的影响，从而为皮康口服液治疗白癜风提供实验理论依据，为进一步探讨中药治疗白癜风的作用机理奠定基础。浙江大学硕士学位论炙一几、研究对象和方法 1、研究对象 角质形成细胞:由本院外科门诊手术室提供的正常儿童环切包皮经分离培养后获得，用于本次研究的为第4代细胞。 中药:加味桃红四物汤(又名皮康口服液)，由黄茂、生晒参、当归、丹参、生地、红_花、桃仁、赤芍、川芍、_女贞子、旱莲草等组成。本院制剂室用水提醇沉法制备，lomL/支，相当于」:药12.29/支。 2、力‘法 中药对角质形成细胞增殖影响的测定采用MTT法。 「}一‘药对角质形成细胞分泌ET-1和bFGF影响的测定采用ELISA法。 3、统计学处理 应用SPSSIO.0统计软件对实验数据进行处理。两个样本均数的比较采用t检验。三、结果 】、，高浓度的皮康口服液(4 .27、5.49、6.lmg/mL)细胞增殖指数明显高于对照组 (只0.05)，而低浓度的皮康口服液(l .22、1 .83、3.05mg/mL)细胞增殖指数与 对照组无明显差异(乃0.05)。 2、高浓度(5.49mg/mL)或低浓度(l .83mg/mL)皮康口服液组角质形成细胞分 泌ET-1的量均明显高于不加皮康日服液的对照组(只0.01)，而加入皮康口服 液组角质形成细胞分泌bFGF的量与不加皮康口服液的对照组无明显差异 (乃().()5)。四、结论 1、高浓度的皮康口服液对角质形成细胞增殖有促进作用，而低浓度的皮康口服 液对角质形成细胞增拍无促进作用。 2、皮康口服液能促进角质形成细胞分泌ET-1，而不能促进角质形成细胞分泌 bFGF。 3、皮康口服液在体外可刺激角质形成细胞分泌ET-1，提示其治疗白瘫风的作用浙江大学硕士学位论义机理可能是通过促进角质形成细胞分泌ET-1，进而促进黑素细胞的增殖、黑素生成及增加酪氨酸酶的活性这·途径。更多还原

【Abstract】 Background and ObjectiveVitiligo is a kind of disease characterized by the depigmentation of the skin and hair because of acquired melanocytes damage. At present, the pathogenesis of vitiligo remains unclear, but many domestic and abroad researches have proved that vitiligo may be related with heredity, immunity and nerval elements. Of late years, more and more studies find that the cytokines participate in the development of vitiligo. There are similarities between keratinocytes and melanocytes, and the cytokines secreted by keratinocytes play certain roles in the survival and activation of melanocytes. Human keratinocytes have been shown to synthesize and secrete endothelin-l(ET-l) and basic fibroblast growth factor (bFGF) in culture. Recently it has been shown that ET-1 is able to stimulate melanocytes proliferation and tyrosinase activity via a receptor-mediated signal transduction pathway, and bFGF is a potent mitogen for melanocytes. For these reasons, it is important to study the relationship between ET-1 and bFGF and vitiligo. As yet we haven’t seen any published article on the effect of Chinese traditional medicine on keratinocytes secreting ET-1 and bFGF.In order to explore the mechanism of Chinese traditional medicine treating vitiligo. we investigate the effect of Modified Taohong Decoction (Pikang oral liquid)on keratinocytes secreting ET-1 and bFGF. Materials and Methods1. MaterialsHuman Keratinocytes: the normal child foreskin was collected, and keratinocytes were separated and cultured. The fourth era cells were investigated.Chinese traditional medicine: Modified Taohong Siwu Decoction (Pikang oral liquid), which contains radix astragali, dry radix ginseng, radix angelicae sinensis, radix salviae militiorrhizae, radix rehmanniae, flos carthami, semen persicae, radix paeoniae rubra, rhizoma chuanxiong, fructus ligustri lucidi, herba hyperici and so on, provided by the pharmaceutical department of our hospital. 10mL Pikang oral liquid corresponds to 12.2 gram of raw.2. MethodsThe effects of Chinese traditional medicine on keratinocyte proliferation were measured by MTT method.The effects of Chinese traditional medicine on the ET-1 and bFGF secretion of keratinocytes were measured by ELISA technique using commercially available ELISA Kits according to the manufacturer’s instructions.3. Statistical analysisAll the data were analysized using SPSS 10.0 statistical software. Student’s Mest was used to analysis the statistical differences between Chinese traditional .medicine and control.Results1. The cells proliferation indexs were significantly higher in high concentration Pikang oral liquids (4.27, 5.49, 6.1mg/mL) than in of blank controls(P<0.05), and no significant differences were observed between low concentration Pikang oral liquids(1.22, 1.83, 3.05mg/mL) and blank controls (P>0.05) .2. ET-1 secreted by keratinocytes were significantly greater in both high concentration(5.49mg/mL) and low concentration (1.83mg/mL) Pikang oral liquid than that in blank controls(,P<0.01), and no significant differences were observed between different concentration Pikang oral liquids and blank controls in bFGF secreted by keratinocytes.(T>0.05) .Conclutions1. High concentration of Pikang oral liquid can enhance the proliferation of keratinocytes, and low concentration of Pikang oral liquid have no effect on the proliferation of keratinocytes.2. Pikang oral liquid can stimulate keratinocytes to secrete ET-1, and have no effect on the secretion of bFGF.3. The results suggest that the therapeutic effect of Pikang oral liquid on vitiligo may be related to promoting the ET-1 secretion of keratinocytes and stimulating the proliferation and melanogenesis of melanocytes.更多还原