温肾降浊化瘀方对溃疡性结肠炎CD14/TLR4-NF-κB通路的影响

Effect of Wenshen Jiangzhuo Huayu decoction on the expression of CD14,TLR-4 and NF-κB in ulcerative colitis in mice

目的:观察溃疡性结肠炎(ulcerative colitis,UC)小鼠结肠黏膜CD14、TLR-4及NF-κBp65表达变化,探讨温肾降浊化瘀方干预UC的可能机制.方法:60只清洁级Balb/c小鼠随机分5组,每组12只,正常组、模型组、治疗组(低、中、高剂量组);除外正常组,其余各组采用自由饮用5%DSS法制备UC模型,模型组给予等剂量生理盐水,治疗组分别给予不同剂量温肾降浊化瘀类方药提取物干预6wk.分别应用光镜检测结肠黏膜组织形态学变化,免疫组织化学及实时荧光定量PCR检测结肠黏膜CD14、TLR-4以及NF-κBp65蛋白及其mRNA表达变化.结果:光镜示模型组小鼠结肠黏膜充血肿胀明显,上皮细胞脱落,伴溃疡明显,有大量中性细胞浸润;治疗组小鼠结肠黏膜充血、水肿、部分黏膜上皮脱落,未见溃疡形成;按结肠黏膜组织损伤评分,各治疗组评分(3.58±0.37)均低于模型组(7.36±0.27),有统计学意义(P<0.05).模型组CD14、TLR-4以及NF-Bp65蛋白及其mRNA表达呈上升趋势(3.17±0.55,3.86±0.75,4.75±0.52),治疗组上述指标表达(1.98±0.33,1.75±0.32,1.64±0.27)呈下降趋势,有统计学意义,无剂量依赖性关系.结论:温肾降浊化瘀方药可能通过介导CD14/TLR-4-NF-κB通路相关指标表达,发挥防治UC的效应.

AIM：To explore the mechanism by which Wenshen Jiangzhuo Huayu decoction prevents ulcerative colitis by detecting the expression of CD14,TLR-4 and NF-κB in colon mucosa of mice with ulcerative colitis.METHODS：Sixty Balb/c mice were randomly and equally divided into five groups：normal group,blank group,and three intervention groups （low,medium,high doses）.Except for the normal group,the other groups were given orally 5% DSS for four weeks.Mice in the intervention groups were intragastrically administered with different doses of Wenshen Jiangzhuo Huayu decoction for four weeks,and the blank group was intragastrically administered with normal saline for the same duration.Colon histomorphology was assayed by naked eyes and light microscopy.The expression of CD14,TLR-4 and NF-Bp65 was detected by immunohistochemistry and real-time PCR.RESULTS：Hyperemia,edema and ulcer were noted in the colon mucosa of mice in the blank group,while the changes in the intervention groups were characterized by hyperemia and edema.There was a significant difference in DAI index between the blank group and intervention groups （7.36 ± 0.27 vs 3.58 ± 0.37,P 0.05）.The expression of CD14,TLR-4 and NF-Bp6 was up-regulated in the blank group compared to the normal group.However,the expression of CD14,TLR-4 and NF-Bp6 was significantly down-regulated in the intervention groups compared to the blank group （1.98 ± 0.33 vs 3.17 ± 0.55,1.75 ± 0.32 vs 3.86 ± 0.75,1.64 ± 0.27 vs 4.75 ± 0.52,all P 0.05）.CONCLUSION：Wenshen Jiangzhuo Huayu decoction may prevent ulcerative colitis by regulating the CD14/TLR-4-NF-B signal pathway.