铁皮石斛多糖对辣椒素激活BV2细胞内TRPV1的调控作用  

The regulation effect of Dendrobium officinalis polysaccharides on TRPV1 in CapsaicininducedBV2cells

作　　者：彭奕 敢小双 沈伟花 韩萍 林丽 杜志云 PENG Yi;GAN Xiao-shuang;SHEN Wei-hua;HAN Ping;LIN Li;DU Zhi-yun(Xinjiang Institute of Physics and Chemistry,Chinese Academy of Sciences,Urumqi,Xinjiang 830011,China;Infinitus(China)Co.,Ltd,Jiangmen,Guangdong 529156,China;School of Biomedicine,Guangdong University of Technology,Guangzhou,Guangdong 510062,China;Foshan Conney Allan Biotechnology Co.,Ltd.,Foshan,Guangdong 528200,China;University of Chinese Academy of Sciences,Beijing 100000,China)

机构地区：[1]中国科学院新疆理化技术研究所,新疆乌鲁木齐830011 [2]无限极(中国)有限公司,广东江门529156 [3]广东工业大学生物医药学院,广东广州510062 [4]佛山市康伲爱伦生物技术有限公司,广东佛山528200 [5]中国科学院大学,北京100000

出　　处：《日用化学品科学》2024年第12期18-24,共7页Detergent & Cosmetics

基　　金：国家自然科学基金(2217081416)。

摘　　要：优化铁皮石斛多糖的提取工艺,对多糖含量、分子量进行表征,研究其对辣椒素(Capsaicin)特异性激活BV2细胞内TRPV1的调控作用。通过正交试验优化提取料液比、提取温度、提取时间3个关键因素对多糖提取工艺的影响,采用苯酚硫酸法测定多糖含量,高效液相色谱联用多角度激光散射测定多糖分子量,利用Capsaicin特异性激活BV2细胞内TRPV1,建立神经高敏反应模型,采用提取制备的多糖处理BV2细胞24h,通过Elisa试剂盒测定和活细胞免疫荧光特异性标记TRPV1抗体,FITC染色研究TRPV1在BV2细胞内的表达情况。结果表明,铁皮石斛多糖提取最佳工艺为:料液比为40g/mL,提取温度为100℃,提取时间为3h,提取次数为2次,制备多糖的收率为13.09%,中性多糖含量为78.72%,保留时间为11~20min,其分子量范围为380~274500Da,其中大于200kDa分子量的多糖占82.75%。与空白对照组相比,Capsaicin刺激能显著提高BV2细胞中TRPV1的特异性表达(p<0.05),采用0.1mg/mL浓度的铁皮石斛多糖进行保护或修复,可以明显降低BV2细胞内TRPV1的活性(p<0.001),抑制BV2细胞中TRPV1的特异性表达,且前期保护效果显著。铁皮石斛多糖对Capsaicin特异性激活BV2细胞内TRPV1的神经高敏反应有明显的保护及修复作用。The study aimed to optimize the extraction process of polysaccharides from Dendrobium oficinale,characterize the content and the molecular weight of polysaccharides,and further study its regulatory effect on Capsaicin specific activation of TRPV1 in BV2 cells.Based on the orthogonal experiment,the effects of three key factors,namely the extraction liquid ratio,extraction temperature,and extraction time,on the polysaccharide extraction technology were optimized.The polysaccharide content was determined by phenol sulfuric acid method,and the polysaccharide molecular weight was measured by high performance liquid chromatography(HPLC)coupled to multi-angle laser light scattering method.Capsaicin was used to specifically induced TRPV1 in BV2 cell to establish a neural high-sensitivity reaction model.BV2 cells was treated with the polysaccharide for 24 h,and the expression of TRPV1 in BV2 cells was evaluated by ELISA kit,immunofluorescence and FITC staining.The results showed that the optimal extraction condition was as follows:a extraction liquid ratio of 40 g/mL,an extraction temperature of 100°C,an extraction time of 3 h,and the extraction times of 2 times.The yield of polysaccharides prepared was 13.09%,the content of neutral polysaccharides was 78.72%,and the retention time was 11~20 min.The molecular weight range was 380~274500 Da,with the polysaccharide with the molecular weight greater than 200 kDa accounted for 82.75%.Compared with the control group,Capsaicin stimulation significantly increased the expression of TRPV1 in BV2 cells(p<0.05).The activity of TRPV1 in BV2 cells was significantly decreased(P<0.001)and the expression of TRPV1 in BV2 cells was inhibited by using O.1 mg/mL Dendrobium officinale polysaccharide for protection or repair,with a significant protective effect in the early stage.Dendrobium officinalis polysaccharide has protective and repair effects markedly on the neuro-hypersensitivity of TRPV1 in Capsaicininduced BV2 cells.

关 键 词：铁皮石斛多糖 BV2细胞 TRPV1 神经高敏反应 保护作用 

分 类 号：TQ658[化学工程—精细化工]