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目的观察大黄素对急性胰腺炎(AP)大鼠胰腺组织核因子-kB(NF-kB)活化的影响,初步阐明大黄素治疗 AP 的分子作用机制。方法雄性 SD 大鼠采用持续输注雨蛙素(5 μg·kg~(-1)·h~(-1))法建立急性水肿性胰腺炎模型,分为注射后5、30min 及1、2、4、6 h 组,停止输注后6h 组;造模前2h 大鼠腹腔注射大黄素(10、30、100mg/kg)组及生理盐水对照组。采用电泳迁移率改变分析实验检测 NF-kB DNA 结合活性,Western blot 检测 NF-kB 抑制蛋白(ⅠkBs)的ⅠkBα和ⅠkBβ降解水平,Northern blot 检测 NF-kB 信号下游效应分子单核细胞趋化蛋白-1(MCP-1)和细胞间粘附分子-1(ICAM-1)基因表达。结果超生理剂量的雨蛙素可诱导大鼠胰腺 NF-kB 发生双相活化,活化高峰出现在雨蛙素注射30min 和4h 后,其相对面积灰度值分别为正常对照的(3.9±2.0)和(7.7±3.2)倍(n=4,P 值均<0.01)。大黄素10、30和100mg/kg剂量治疗组大鼠胰腺 NF-kB 活性较 AP 模型组分别下降45%,62%和84%(30min)和28%,74%和80%(4h,n=4,P 值均<0.01)。同时,大黄素(100mg/kg)可明显抑制雨蛙素诱导的胰腺ⅠkBα和ⅠkBβ蛋白降解(n=4,P 值均<0.01)。与其对 NF-kB 活化的抑制作用一致,大黄素剂量依赖性地抑制雨蛙素诱导的胰腺 MCP-1和 ICAM-1基因表达。结论大黄素可抑制胰腺内ⅠkBs/NF-kB 信号转导通路活化,从而抑制炎性细胞因子和粘附分子基因表达,发挥其对 AP 的治疗作用。
Objective To observe the effects of emodin on pancreatic nuclear factor-kappa B (NF-kB) activation in rats with acute pancreatitis (AP), and to elucidate the molecular mechanism of emodin treatment of AP. Methods Male SD rats were treated with continuous infusion of cerulein (5 μg·kg -1 ·h -1) to establish an acute edematous pancreatitis model, which was divided into 5, 30 min and 1, 2, In the 4 and 6 h groups, the 6 h group was stopped after the infusion; the rats were intraperitoneally injected with emodin (10, 30, 100 mg/kg) and saline control groups 2 h before modeling. Electrophoretic mobility shift assay was used to detect NF-kB DNA binding activity. Western blot was used to detect IkBα and IkBβ degradation of NF-kB inhibitory protein (IkBs). Northern blot was used to detect NF-kB signaling downstream effector monocyte chemoattractant protein- 1 (MCP-1) and intercellular adhesion molecule-1 (ICAM-1) gene expression. Results The hyperphysiological dose of cerulein could induce biphasic activation of NF-kB in rat pancreas. The peak of activation appeared after injection of cerulein for 30 min and 4 h. Its relative area gray value was (3.9±2.0) and 7.7 ± 3.2) times (n = 4, P <0.01). The activity of NF-kB in the pancreas of rats treated with emodin at doses of 10, 30, and 100 mg/kg was lower than that in the AP model group by 45%, 62%, and 84% (30 min) and 28%, 74%, and 80%, respectively (4h, n= 4, P values are <0.01). At the same time, emodin (100 mg/kg) significantly inhibited the degradation of IkBα and IkBβ proteins induced by cerulein (n=4, P<0.01). Consistent with its inhibitory effect on NF-kB activation, emodin dose-dependently inhibited cerulein-induced pancreatic MCP-1 and ICAM-1 gene expression. Conclusion Emodin can inhibit the activation of IkBs/NF-kB signal transduction pathway in the pancreas, thereby inhibiting the expression of inflammatory cytokines and adhesion molecules, and exerting its therapeutic effect on AP.