大黄素和栀子苷配伍对全身炎症反应综合征大鼠肠黏膜屏障损伤及炎症反应的影响

Combined effects of Emodin and Geniposide on intestinal mucosal barrier damage and inflammatory response in rats with systemic inflammatory response syndrome

目的观察大黄素和子苷配伍对全身炎症反应综合征(SIRS)大鼠肠黏膜屏障功能损伤的保护和炎症反应的抑制作用。方法选择6~8周龄雄性SD大鼠,按随机数字表法分为正常对照组、模型组、大黄素组、子苷组、大黄素与子苷配伍组(配伍组)乌司他丁组。通过腹腔注射酵母多糖的方法构建大鼠SIRS模型,大黄素组、子苷组、乌司他丁组、配伍组于大鼠注射酵母多糖后即刻和12h后各给药1次。制模后24h,测定各组大鼠血清D-乳酸(D-LA)、二胺氧化酶(DAO)、内毒素(ET)、肿瘤坏死因子-α(TNF-α)、白细胞介素(IL-1β、IL-6、IL-10)含量;取小肠组织观察病理学改变;采用免疫组化法测定小肠组织核转录因子-kBp65(NF-kBp65)及Toll样受体4(TLR-4)的蛋白阳性表达水平。结果与正常对照组比较,模型组大鼠血清D-LA、DAO、ET、TNF-α、IL-1β、IL-6、IL-10均明显升高[D-LA(μmol/L):99.11±11.93比36.94±1.92,DAO(U/L):5018.80±759.00比2253.23±372.40,ET(μg/L):0.36±0.04比0.15±0.02,TNF-α(ng/L):66.61±20.88比9.47±0.78,IL-1β(ng/L):63.73±7.64比25.86±5.90,IL-6(ng/L):392.00±56.47比111.17±36.22,IL-10(ng/L):41.90±8.12比19.75±1.54,均P<0.05],组织病理学观察可见小肠黏膜水肿明显,黏膜上皮细胞排列紊乱并伴有细胞脱落,肠黏膜内炎症细胞浸润增多,杯状细胞减少,固有层松散、充血;小肠组织中TLR-4及NF-kB蛋白阳性表达明显增强[TLR-4蛋白阳性表达(A值):0.59±0.08比0.27±0.04,NF-kB蛋白阳性表达(A值):0.65土0.07比0.30土0.06,均P<0.05]。与模型组比较,大黄素组、子苷组、乌司他丁组、配伍组血清D-LA、DAO和ET水平均明显降低[D-LA(μmol/L):67.49±8.32、69.08±6.76、69.17±5.63、58.16±7.12比99.11±11.93,DAO(U/L):3659.38±563.90、3713.29±354.70、3575.30±444.40、3087.01±227.50比5018.80±759.00,ET(μg/L):0.27±0.04、0.24±0.03、0.23±0.03、0.20±0.02比0.36±0.04,均P<0.05],促炎因子TNF-α、IL-1β、IL-6含量亦均明显著降低[TNF-α(ng/L):44.34±10.63、39.23±11.74、35.80±11.49、28.74±9.56比66.61±20.88,IL-1β(ng/L):50.30±8.22、46.74±5.10、48.25±5.16、40.84±5.02比63.73±7.64,IL-6(ng/L):299.27±50.65、263.98±37.62、281.84±63.24、216.72±38.90比392.00土56.47,均P<0.05]。大黄素、子苷、大黄素与子苷配伍、乌司他丁均可升高血清抗炎因子IL-10水平(ng/L:92.63±32.83、87.34±30.79、71.66±16.82、133.70±39.40比41.90±8.12,均P<0.05)。大黄素组、子苷组、配伍组、乌司他丁组肠组织病理改变减轻,小肠组织中NF-kBp65和TLR-4蛋白阳性表达均显著降低[TLR-4蛋白阳性表达(A值):0.49±0.03、0.47±0.08、0.36±0.08、0.42±0.06比0.59±0.08,NF-kBp65蛋白阳性表达(A值):0.50±0.06、0.49±0.07、0.42±0.06、0.46±0.09比0.65±0.07,均P<0.05]。与大黄素组、子苷组比较,配伍组血清D-LA、DAO、ET、TNF-α、IL-1β、IL-6均显著降低,IL-10水平显著升高,肠组织病理改变减轻,小肠组织NF-kBp65和TLR-4蛋白阳性表达水平均明显降低(均P<0.05)。结论大黄素和子苷有助于缓解酵母多糖引发的SIRS,其作用与保护小肠黏膜屏障、抑制炎症反应有关,二者配伍有增效作用。

Objective To observe the protective and anti-inflammatory effects of Emodin and Geniposide compatibility on the intestinal mucosal barrier function damage in rats with systemic inflammatory response syndrome(SIRS).Methods Male Sprague-Dawley(SD)rats aged 6-8 weeks were randomly divided into normal control group,model group,Emodin group,Geniposide group,Emodin and Geniposide compatibility group(compatibility group),and Ulinastatin group.The SIRS model in rats was established by abdominal injection of yeast polysaccharide.The Emodin,Geniposide,Ulinastatin,and compatibility groups received administration immediately and 12 hours after the injection of yeast polysaccharide.After 24 hours of modeling,the contents of D-lactate(D-LA),diamine oxidase(DAO),endotoxin(ET),tumor necrosis factor-α(TNF-α),interleukins(IL-1β,IL-6,IL-10)in the serum of each group were measured;The small intestine was taken for histopathological examination,and the positive protein expression levels of nuclear factor-kB p65(NF-kB p65)and Toll-like receptor 4(TLR-4)in the small intestine tissue were determined by immuno histochemistry.Results Compared with the normal control group,the levels of D-LA,DAO,ET,TNF-α,IL-1β,IL-6 and IL-10 in the serum of the model group were significantly increased[D-LA(μmol/L):99.11±11.93 vs.36.94±1.92,DA0(U/L):5018.80±759.00 vs.2253.23±372.40,ET(μg/L):0.36±0.04 vs.0.15±0.02,TNF-α(ng/L):66.61±20.88 vs.9.47±0.78,IL-1β(ng/L):63.73±7.64 vs.25.86±5.90,IL-6(ng/L):392.00±56.47 vs.111.17±36.22,IL-10(ng/L):41.90±8.12 vs.19.75±1.54,all P<0.05],histopathological observations showed that the small intestine mucosa was significantly swollen,the arrangement of mucosal epithelial cells was disordered,and there was cell shedding,increased infiltration of inflammatory cells in the intestinal mucosa,decreased goblet cells,loose and congested lamina propria;the positive protein expression of TLR-4 and NF-B in the small intestine tissue was enhanced[TLR-4 positive protein expression(A value):0.59±0.08 vs.0.27±0.04,NF-kB positive protein expression(A value):0.65±0.07 vs.0.30±0.06,both P<0.05].Compared with the model group,the levels of D-LA,DA0,and ET in the serum of the Emodin group,Geniposide group,Ulinastatin group,and compatibility group were significantly decreased[D-LA(μmol/L):67.49±8.32,69.08±6.76,69.17±5.63,58.16±7.12 vs.99.11±11.93,DA0(U/L):3659.38±563.90,3713.29±354.70,3575.30±444.4,3087.01±227.50 vs.5018.80±759.0,ET(μg/L):0.27±0.04,0.24±0.03,0.23±0.03,0.20±0.02 vs.0.36±0.04,all P<0.05],and the contents of pro-inflammatory factors TNF-α,IL-1β,and IL-6 were significantly decreased[TNF-α(ng/L):44.34±10.63,39.23±11.74,35.80±11.49,28.74±9.56 vs.66.61±20.88,IL-1β(ng/L):50.30±8.22,46.74±5.10,48.25±5.16,40.84±5.02 vs.63.73±7.64,IL-6(ng/L):299.27±50.65,263.98±37.62,281.84±63.24,216.72±38.90 vs.392.00±56.47,all P<0.05].The levels of serum anti-inflammatory factor IL-10 were significantly increased(ng/L:92.63±32.83,87.34±30.79,71.66±16.82,133.70±39.40 vs.41.90±8.12,all P<0.05).The pathological changes in the intestinal tissue of the Emodin group,Geniposide group,compatibility group,and Ulinastatin group were reduced,and the positive expressions of NF-kB p65 and TLR-4 proteins in the small intestine tissue were significantly decreased[TLR-4 positive protein expression(A value):0.49±0.03,0.47±0.08,0.36±0.08,0.42±0.06 vs.0.59±0.08,NF-kB p65 positive protein expression(A value):0.50±0.06,0.49±0.07,0.42±0.06,0.46±0.09 vs.0.65±0.07,all P<0.05].Compared with the Emodin group and Geniposide group,the serum D-LA,DAO,ET,TNF-α,IL-1β,and IL-6 in the compatibility group were significantly decreased,the serum IL-10 level was significantly increased,the pathological changes in the intestinal tissue were reduced,and the expression levels of NF-kB p65 and TLR-4 in the small intestine tissue were significantly decreased(all P<0.05).Conclusions Emodin and Geniposide can help relieve SIRS induced by yeast polysaccharide,and their effect is related to protecting the intestinal mucosal barrier and inhibiting the inflammatory response.When combined,they exhibit a synergistic effect.