愈溃方对实验性溃疡性结肠炎大鼠肠黏膜Toll样受体4、髓样分化因子、核因子κB信号通路的影响

Effects of Yukui Formula on TLR4/MyD88/NF-κB Signaling Pathway in Intestinal Mucosa of Rats with Experimental Ulcerative Colitis

目的观察愈溃方对实验性大鼠溃疡性结肠炎(Ulcerative colitisi,UC)的有效性,并结合对大鼠肠黏膜核因子κB(Nuclear Factor kappa-B,NF-κB)、髓样分化因子(Myeloid differentiation factor 88,MyD88)和Toll样受体4(Toll-like Receptor 4,TLR4)的影响探讨其治疗机理。方法将70只雄性SD大鼠按体重随机分为正常对照组(C组)(8只)和模型组(M组)(62只)。C组不做特殊处置,M组进行结肠炎造模前(TNBS)联合乙醇造模。造模结束次日按体质量随机分为5组:模型对照组(MC组)、美沙拉嗪组(ASA组)、中药小剂量组(CS组)、中药中剂量组(CM组)和中药大剂量组(CL组)。C组和MC组给予0.5%羧甲基纤维素钠灌胃;ASA组给予美沙拉嗪缓释剂混悬液灌胃;3个中药组按不同药物浓度分别给予中药灌胃。灌胃剂量:10 ml/(kg·d),剂量相当于美沙拉嗪360 mg/kg、中药小剂量2.835 g/kg、中药中剂量5.67 g/kg、中药大剂量11.34 g/kg。于处置14 d后,依据疾病活动指数(Disease activity index,DAI)和结肠大体形态损伤指数(Colon macroscopic damage index,CMDI)评价药物疗效;ELISA法检测结肠组织NF-κB、MyD88、TLR4水平。结果模型对照组大鼠DAI评分值升高,与C组比较,差异有统计学意义(P<0.05)。干预后针对DAI、CMDI计分展开组间比较:中药大、中、小剂量组与ASA组DAI计分均有下降,与MC组比较,差异有统计学意义(P<0.05),但4组间两两比较,差异均无统计学意义(P>0.05)。NF-κB、My-D88和TLR4在MC组均明显升高,其水平与其余各组比较,差异有统计学意义(P<0.05)。经药物干预并随时间推移,各组NF-κB水平均有不同程度下降,依下降程度依次为ASA组>CL组>CM组>CS组>MC组。ASA组、CL组与CS组比较,差异有统计学意义(P<0.05),而CM组与CS组比较,差异无统计学意义(P>0.05),ASA组、CL组、CM组3组间比较,差异无统计学意义(P>0.05);My-D88和TLR4下降程度依次为CL组>ASA组>CM组>CS组>MC组。CL组、ASA组分别与CM组、CS组比较,差异有统计学意义(P<0.05),而CL组与ASA组之间、CM组与CS组之间比较,差异无统计学意义(P>0.05)。结论愈溃方治疗实验性大鼠UC有效,治疗机理可能与调控肠黏膜TLR4/My-D88/NF-κB通路有关。

Objective To observe the efficacy of Yukui Formula in the treatment of experimental ulcerative colitis(UC)in ratsand explore its therapeutic mechanism based on the effects on nuclear factor kappa-B(NF-κB),myeloid differentiation factor 88(MyD88),and Toll-like receptor 4(TLR4)in the intestinal mucosa of rats.Methods Seventy male SD rats were randomly divided into a normal control(C)group(n=8)and a model(M)group(n=62)according to body weight.The rats in the C group received no manipulations,while those in the M group underwent modeling by TNBS combined with ethanol.On the next day after modeling,rats were divided into a model control(MC)group,a 5-aminosalicylic acid(ASA)group,and Chinese medicine small,medium,and large dose(CS,CM,and CL)groups.Rats in the C groupand the MC group were given 0.5%sodium carboxymethyl cellulose at 10 ml/(kg·d)by gavage,and those in the ASA group received ASA suspension at 360 mg/kg by gavage.Rats in the CS,CM,and CL groups received Yukui Formula at 2.835 g/kg,5.67 g/kg,and 11.34 g/kg,respectively.After 14 days of administration,drug efficacy was evaluated according to the disease activity index(DAI)and colon macroscopic damage index(CMDI).NF-κB,MyD88,and TLR4 in colon tissues were detected by ELISA.Results The DAI score of the MC group was higher than that of the C group(P<0.05).After the intervention,the scores of DAI and CMDI were compared between groups.Specifically,the scores of DAI in the CS,CM,and CL groups and the ASA group decreased compared with that of the MC group(P<0.05),but there was no significant difference between the four groups(P>0.05).NF-κb,MyD88,and TLR4 in the MC group increased compared with those in the other groups(P<0.05).With the prolongation of time after drug intervention,the level of NF-κB in each group decreased,which was ranked as follows:ASA group>CL group>CM group>CS group>MC group.The ASA group,the CL group,and the CS group showed significant differences(P<0.05),but there was no significant difference between the CM group and the CS group(P>0.05).There was no significant difference among the ASA group,the CL group,and the CM group(P>0.05).In terms of decreasing degree of MyD88 and TLR4,the groups were ranked as follows:CL group>ASA group>CM group>CS group>MC group.The CL group and the ASA group were compared with the CM group and the CS group respectively,and the differences were statistically significant(P<0.05).There was no significant difference between the CL group and the ASA group,and between the CM group and the CS group(P>0.05).Conclusion Yukui Formula is effective in the treatment of experimental UC in rats,and the mechanism may be related to the regulation of the TLR4/MyD88/NF-κB signaling pathway in the intestinal mucosa.