雷公藤柱层析各段组分对胶原诱导性关节炎模型大鼠细胞因子表达的影响

Effect of Each Component of Tripterygium Wilfordii Column Chromatography on Expression of Cytokines in Collagen-induced Arthritis Rats

关键词:  雷公藤  柱层析  组分  胶原诱导性关节炎  肿瘤坏死因子  白细胞介素-1β  细胞凋亡

Key words:  Tripterygium wilfordii  column chromatography  component  collagen-induced arthritis  TNF-α  IL-1β  cell apoptosis

摘要:

目的 比较雷公藤柱层析各段组分对Ⅱ型胶原诱导性关节炎（collagen-induced arthritis，CIA）模型大鼠血清中肿瘤坏死因子-α（TNF-α）、白细胞介素-1β（IL-1β）表达的影响，初步探讨各段组分中有效成分与药效的关系。方法 HPLC测定各段组分中6个有效成分的含量。在SD大鼠背、尾根及左后足跖多点皮内注射牛Ⅱ型胶原蛋白（BCⅡ）乳液，建立CIA大鼠模型。选取80只成模大鼠随机分为模型对照组、阳性对照组和各段组分组，每组各8只。免疫20 d后，连续灌胃给药3周，记录大鼠关节炎指数（arthrits index，AI），通过酶联免疫法检测血清中TNF-α、IL-1β的表达，原位末端转移酶标记技术（TUNEL染色法）检测各段组分对肝细胞凋亡的影响，Western-blot检测大鼠肝组织中caspase-3蛋白表达。结果 中段和末段组分中有效成分的含量差异较大。与模型对照组比较，中段组分的高、低剂量组和末段组分的高剂量组给药3周后均能明显降低AI值（P<0.01），能明显抑制大鼠血清中TNF-α、IL-1β表达（P<0.01），也能明显诱导肝细胞凋亡（P<0.01），明显升高caspase-3活性（P<0.01）。结论 雷公藤柱层析各段组分治疗CIA可能与降低血清中TNF-α和IL-1β表达有关，灌胃后会诱导大鼠肝细胞发生凋亡，上调caspase-3活性，有效成分与药效存在量效关系。

Abstract:

OBJECTIVE To compare the effects of each component of Tripterygium wilfordii column chromatography on the expression of tumor necrosis factor-alpha(TNF-α) and interleukin-1β(IL-1β) in the serum of rats with type Ⅱ collagen- induced arthritis(CIA), and to explore the relationship between effective components of each component and their pharmacology effect. METHODS HPLC was used to determine the content of 6 effective components in each component. Bovine collagen Ⅱ(BCⅡ) emulsion was injected into the dorsal, tail and left posterior plantar plantaris of SD rats, and the rat model of CIA was established. After successful modeling, eighty CIA rats were selected and randomly divided into model control group, positive control group and groups of each component, with 8 rats in each group. After twenty days of immunization, rats were given intragastric administration for 3 weeks, the rats' arthritis index(AI) was recorded. The expression of TNF-α and IL-1β in serum was detected by enzyme-linked immunosorbent assay. In situ TUNEL assay was used to detect the effects of different components on hepatocyte apoptosis. Western-blot was used to detect the expression of caspase-3 protein in rats' liver. RESULTS The contents of effective components in the middle and the last components were quite different. Compared with the model control group, the high and low dose group of the middle components and the high dose group of the last components could obviously reduce the AI value three weeks after administration(P<0.01), inhibit the expression of TNF-α and IL-1β in the serum of rats(P<0.01), induce hepatocyte apoptosis(P<0.01), and increase the activity of caspase-3(P<0.01). CONCLUSION Each component of Tripterygium wilfordii column chromatography has therapeutie effects on CIA rats, its mechanisms may be related to the reduction of the expression of TNF-α and IL-1β in the serum, the rat hepatocyte apoptosis can be induced and the activity of caspase-3 can be up-regulated after gavage, there is a dose-effect relationship between effective components and pharmacology effect.