大黄牡丹汤对脓毒症急性肠功能障碍大鼠肠道髓系细胞触发受体-1表达的影响

Effect of Dahuang Mudan Tang in Triggering Receptor Expressed on Myeloid Cells-1 in Intestinal Tract in Rats with Septic Acute Intestinal Dysfunction

目的:研究大黄牡丹汤对脓毒症大鼠急性肠功能障碍的治疗作用是否与调控肠道髓系细胞触发受体-1(TREM-1)表达有关。方法:雄性SD大鼠100只,腹腔注射4. 5 mg·kg^-1的脂多糖(LPS)进行造模,造模成功后随机分为5组:模型组,谷氨酰胺组(3. 75 g·kg^-1),大黄牡丹汤低、中、高剂量组(7. 5,15,30 g·kg^-1),另设正常大鼠10只作为正常组。每日1次。7 d后所有大鼠灌胃混悬液2 m L(乳果糖100 mg,甘露醇50 mg)。留取24 h尿液,运用带电化学检测器的高压液相色谱仪分析尿中乳果糖(lactulose)与甘露醇(mannitol)比值(L/M)。取血和回肠组织,运用高效液相色谱法检测血清瓜氨酸浓度;酶联免疫吸附法检测血清髓系细胞触发受体-1(TREM-1),肿瘤坏死因子-α(TNF-α),肠型脂肪酸结合蛋白(iFABP),D-乳酸浓度;实时荧光定量PCR检测回肠组织TREM-1,Toll样受体2(TLR2),Toll样受体4(TLR4),髓样细胞分化蛋白(MyD88),核转录因子-κB(NF-κB) mRNA的表达水平;电镜观察肠黏膜损伤的病理变化。结果:与正常组比较,模型组血清TREM-1,TNF-α浓度明显升高(P <0. 05,P <0. 01);回肠组织TREM-1,TLR2,TLR4,MyD88,NF-κB mRNA表达水平明显升高(P <0. 05,P <0. 01);i FABP,D-乳酸浓度,尿L/M明显升高(P <0. 05);血清瓜氨酸浓度明显降低(P <0. 05);回肠黏膜厚度、绒毛长度明显下降(P <0. 05)。与模型组比较,各给药组血清TREM-1,TNF-α浓度明显下降(P <0. 05,P <0. 01);回肠组织TREM-1,TLR2,TLR4,MyD88,NF-κB mRNA表达水平明显下降(P <0. 05,P <0. 01),4组之间无差异;谷氨酰胺组,大黄牡丹汤中、高剂量组血清i FABP,D-乳酸浓度,尿L/M明显下降(P <0. 05,P <0. 01);血清瓜氨酸浓度明显升高(P <0. 05,P <0. 01);绒毛长度及黏膜层厚度明显升高(P <0. 05);肠黏膜Chiu氏评分明显下降(P <0. 05),3组之间无差异。结论:大黄牡丹汤能有效治疗脓毒症大鼠急性肠功能障碍,其作用机制可能与通过调控肠道髓系细胞触发受体-1表达,减轻肠道炎症反应有关。

Objective: To investigate whether the therapeutic effect of Dahuang Mudan Tang on septic acute intestinal dysfunction in sepsis ratsis related to the regulation of expression of triggering receptor expressed on myeloid cells-1( TREM-1). Method: Totally 100 male SD rats were injected intraperitoneally with lipopolysaccharide( LPS) at a dose of 4. 5 mg·kg^-1 to build sepsis model. The sepsis model rats were randomly divided into five groups: model group,glutamine group( 3. 75 g·kg^-1),low,medium,high-dose Dahuang Mudan Tang group( 7. 5,15,30 g·kg^-1),and another 10 normal rats were selected as normal group. Seven days later,2 m L suspension( 100 mg lactulose and 50 mg mannitol) was orally administrated by gavage,and 24 h urinewas collected. The ratio of lactulose to mannitol in urine( L/M) was detected by HPLC with pulsed electrochemical detection( HPLC-PED). Serum citrulline concentrationsin blood and ileum were determined by HPLC. Enzyme linked immunesorbent assay( ELISA) was used to detect the concentrations of triggering receptor expressed on myeloid cells-1( TREM-1),tumor necrosis factor-α( TNF-α),intestinal fatty acid binding protein( i FABP) and D-lactic acid. Real-time PCR was used to detect the mRNA expressions of TREM-1,Toll-like receptors2( TLR2), Toll-like receptors 4( TLR4), myeloid cell differentiation protein( MyD88), nuclear transcription factor-κB( NF-κB). Electron microscopy was used to observe the pathological changes of intestinal mucosa injury. Result: Compared with normal group,the serum concentrations of TREM-1,TNF-α,i FABP,D-lactate;the ratio of lactulose to mannitol in urine( L/M) and the expressions of TREM-1,TLR2,TLR4,MyD88,NF-κB mRNA in model group were increased obviously( P < 0. 05,P < 0. 01);citrulline concentration was decreased obviously( P < 0. 05);the lengths of the villus and thicknesses of the mucosal layer were decreased obviously( P < 0. 05);the Chiu score was increased obviously( P < 0. 01). Compared with model group,the expressions of TREM-1,TLR2,TLR4,My D88,NF-κB mRNA,and the serum concentrations of TREM-1 and TNF-α in all medication administration groups were decreased obviously( P < 0. 05),with no difference between these groups. Compared with model group,the serum concentrations of i FABP,D-lactate,L/M,the Chiu scorein glutamine group,medium-dose Dahuang Mudan Tang group and high-dose Dahuang Mudan Tang group were decreased obviously( P < 0. 05),lengths of villus and thicknesses of mucosal layers were increased obviously( P <0. 05);and the citrulline concentrations were increased obviously( P < 0. 05). There was no difference between the three groups. Conclusion: Dahuang Mudan Tang can effectively treat SAID in rats,and its mechanism may be realized by regulating the expression of TREM-1 and relieving intestinal inflammation of intestinal tract.