芍药苷通过抑制NLRP3炎症小体治疗溃疡性结肠炎小鼠的研究  

Paeoniflorin Prevent Colonic Inflammation via Inhibiting NLRP3 Inflammasome

作　　者：刘琦[1] 罗霞[1] 罗爽[1] 何金榕 邓向亮[1] 陈扬[1] 周联[1] LIU Qi1, LUO Xia1, LUO Shuang1, HE Jinrong2, DENG Xiangliang1, CHEN Yang1, ZHOU Lian1(1. School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, 510006 Guangdong, China; 2. College of Basic Medicine, Guangzhou University of Chinese Medicine, Guanzzhou 510006 Guanzdong. Chin)

机构地区：[1]广州中医药大学中药学院,广东广州510006 [2]广州中医药大学基础医学院,广东广州510006

出　　处：《中药新药与临床药理》2018年第4期409-414,共6页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：国家自然科学基金资助项目(81603587;81673668;81703785);广东省自然科学基金资助项目(2017A020211016)

摘　　要：目的研究芍药苷(Paeoniflorin,PF)在溃疡性结肠炎(Ulcerative colitis,UC)小鼠结肠组织中的动态分布,并探讨其治疗UC的途径及其作用机制。方法 C57BL/6小鼠在实验第1~7天自由饮用3%葡聚糖硫酸钠(Dextran sulfate sodium,DSS)水溶液制备UC模型,第8天,UC小鼠灌服黄芩汤(含芍药苷),采用超高效液相色谱串联四极杆飞行时间质谱仪(UPLC-Q-TOF-MS)检测芍药苷在结肠中的含量变化,检测时间点为给药0.5,1,3,12 h。另取C57BL/6小鼠随机分为正常组、模型组、柳氮磺吡啶组(Sulfasalazine,SASP)和芍药苷组,每组8只。同时,SASP组(50 mg·kg^(-1))和芍药苷组(25 mg·kg^(-1))分别灌胃给药;正常组和模型组小鼠灌胃给予生理盐水,连续10 d,并记录体质量变化。实验第11天,取小鼠脾脏称质量;采用动物血细胞计数仪检测单核细胞数量;苏木素—伊红染色(HE)观察结肠形态变化;免疫组化法检测结肠和肠系膜中巨噬细胞的分布;超高分辨率激光共聚焦扫描显微镜(Confocal)检测结肠组织巨噬细胞和核苷酸结合寡聚化结构域样受体3(Nucleotide-binding oligomerization domain-like receptor family,pyrin domain-containing 3,NLRP3)蛋白共定位。ELISA试剂盒检测结肠匀浆上清白细胞介素-1β(IL-1β)的含量。结果 UPLC-Q-TOF-MS检测结果表明,芍药苷在结肠中可以检测到,并且芍药苷可显著减轻小鼠结肠溃疡症状并显著促进UC小鼠体质量(P<0.05)和脾指数恢复(P<0.001),降低UC小鼠外周血单核细胞比例(P<0.01)。减少结肠和肠系膜中巨噬细胞浸润的数量,并通过抑制结肠巨噬细胞中NLRP3炎症小体活化进而降低结肠组织上清中IL-1β含量(P<0.01)。结论芍药苷可以在结肠中检测到,并可改善UC小鼠病理症状,抑制肠系膜和结肠组织中巨噬细胞的浸润,抑制结肠巨噬细胞中NLRP3蛋白并抑制细胞因子IL-1β的释放。Objective To investigate the therapeutic effect of PF on UC and its possible pathways. Methods Theconcentrations of PF in DSS-induced UC mice after orally fed with HQT（0.5,1,3,12 h） were detected byUPLC-Q-TOF-MS. In vivo, animals were divided into 4 groups randomly： Control group, DSS group, SASPgroup,PF group（25 mg·kg^（-1））;apart from the normal group,the remaining groups were administrated with 3%DSS,there after provided with regular water for 3 days. PF and SASP were given intragastrically from day 1 to day10 and the mice were sacrificed on day 11. The basal body weight changes of each group, spleen index andmacroscopic appearances of colon were observed. The concentration of monocytes in peripheral blood was detected byPeripheral blood automatic analyzer and the distribution of macrophages（F4/80 ＋） in colon was detected byimmunohistochemistry. Protein co-localization of NLRP3 and colon macrophagocytes was observed by confocalmicroscope. In vitro experiment,RAW264.7 was stimulated with LPS（1 μg·m L-1） and treated with PF（12.5,25, 50 μmol · L-1）. Results PF was detect in the colon. PF could improve the disease symptom of mice（P〈0.05）,reduce the proportion of monocytes in peripheral blood（P〈0.01）,and reduce the number of macrophageinfiltration in the colon,inhibit protein expression of NLRP3 in the macrophage. In vitro experiments showed that PFpossessed anti-inflammatory effect by inhibit the secretion of NO（P〈0.001）. Conclusion PF can be absorbedinto colon and maybe a potential effective drug for ulcerative colitis.

关 键 词：黄芩汤 芍药苷 溃疡性结肠炎 巨噬细胞 NLRP3 白细胞介素-1Β 组织分布 

分 类 号：R285.5[医药卫生—中药学]