白头翁醇提物对三硝基苯磺酸诱导大鼠结肠炎基质金属蛋白酶-3调控作用的研究

Regulation of radix pulsatillae alcohol extract in contols matrix metalloproteinase-3 in colonic mucosa of rat with trinitrobenzenesulfonic acid induced colitis

目的探讨白头翁醇提物对三硝基苯磺酸(Trinitrobenzene sulfonic acid,TNBS)诱导大鼠实验性结肠炎基质金属蛋白酶-3(Matrix metalloproteinase-3,MMP3)基因表达的调控作用。方法动物随机分为正常组、模型组、白头翁组3组。TNBS法制备大鼠结肠炎模型,造模后白头翁组予以白头翁醇提物水溶液(2g/kg.d)2 mL灌肠。于第8天评价各组大鼠疾病活动指数(DAI)和组织学损伤评分;ELISA法检测结肠组织TNF-α、IL-10含量;免疫组织化学法检测NF-κB p65的表达;实时荧光定量PCR(RT-PCR)法测定MMP3-mRNA的表达。结果相比模型组,白头翁组大鼠结肠粘膜DAI和组织学损伤评分、TNF-α含量、NF-κB p65的表达、MMP3-mRNA的表达均降低(P均<0.01或P<0.05),而IL-10含量增高(P<0.01);MMP3-mRNA的表达与NF-κBp65表达呈显著正相关(相关系数r=0.677,P<0.01)。结论白头翁醇提物对TNBS诱导大鼠实验性结肠炎有明显的抗炎效果,其作用机制可能是通过下调MMP3-mRNA的表达、抑制NF-κB的活性、平衡细胞因子网络等实现的。

Objective To explore the regulation of the radix pulsatillae alcohol extract on the gene expression of matrix metalloproteinase-3 （MMP3）, Methods The animals were randomized into 3 groups, including radix pulsatillae alcohol extract treated group, model group and normal group. Rat colitis models were established by trinitrobenzene sulfonic acid （TNBS） enema. The radix pulsatillae alcohoI extract treated group rats were perfused with 2 ml radix pulsatillae alcohol extract at the dosage of 2g/kg/d. On the eighth day, the disease activity index （DAI） and the score of the tissue damage of colonic mucosa was calculated. The level of tumor necrosis factor-α （TNF-α） and interleukin-10（IL-10） was detected by enzyme linked immune sorbent assay （ELISA）. The expression of NF-κB p65 in colonic tissue was observed by immunohistochemistry. The expression of MMP3-mRNA was detected by realtime fluorescent quantitive polymerase chain reaction （RT-PCR）. Results Compared that of model group, DAI and the score of the tissue damages, the levels of TNF-α, the expression of NF-κBp65 and the expression of MMP3-mRNA of radix pulsatillae alcohol extract treated group rats were significantly decreased （P〈0.01 or P〈0.05）, but IL-10 were significantly increased （P〈0.01）. The expression of MMP3-mRNA had a significant positive correlation with the expression of NF-κB p65 （r=0. 677, P〈 0. 01）. Conclusion Radix palstillae alcohol extract has an exact anti-inflammatory effects on TNBS induced rat colitis. The mechanism is possibly related to down-regulated the expression of MMP3-mRNA, inhibit the activation of NF-κBp65 and regulate network balance of eytokines.