别旁茶苷通过PXR/NF-κB信号通路治疗小鼠溃疡性结肠炎机制研究
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摘要
目的探讨别旁茶苷Jasminoside(SC)对溃疡性结肠炎(UC)小鼠的治疗作用及其机制。方法将小鼠随机分为正常对照组,模型组,柳氮磺胺吡啶(SASP)组和SC高、中、低剂量组。除正常对照组外,其余各组采用2,4,6-三硝基苯磺酸(TNBS)建立小鼠结肠炎的模型。给药治疗7 d后取材,显微镜下观察结肠黏膜损伤情况,采用酶联免疫吸附剂测定(ELISA)法测定结肠中α肿瘤坏死因子(TNF-α)、白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)和转化生长因子-β1(TGF-β1)水平;采用逆转录-聚合酶链反应(RT-PCR)法检测结肠组织中细胞色素P4503A4酶(CYP3A4)、孕烷X受体(PXR)、TNF-α、IL-6和IL-1βmRNA表达;采用Western Blot法检测结肠组织中CYP3A4和核转录因子P65(NF-κBP65)表达。结果与正常对照组相比,模型组有明显的病理变化,小鼠结肠黏膜和粘膜下层的多形核细胞浸润情况较严重,出现溃疡和上皮杯状细胞缺失的情况;与TNBS组相比,SASP组和SC各给药组结肠组织的TNF-α、IL-6、IL-1β和TGF-β1的水平及TNF-α、IL-6和IL-1βmRNA的表达水平均有显著下降,并呈剂量依赖性,SASP和SC组中CYP3A4和PXR的mRNA水平显着上调;与正常对照组相比,TNBS组NF-κBP65蛋白表达显着增加,而SASP组和SC组中NF-κBP65表达下降。结论 SC对溃疡性小鼠结肠炎有显著疗效,其可能通过激活PXR、CYP3A,抑制NF-κB的表达,降低炎症因子TNF-α、IL-6、IL-1β和TGF-β1的水平来发挥保护作用。
Objective To investigate the therapeutic effect and mechanism of Jasminoside(SC)on Ulcerative colitis(UC)in mice. Methods The mice were randomly divided into normal group,model group,Sulfasalazine(SASP)group and SC high,middle and low dose groups. Except for normal group,mice in other groups were treated with2,4,6-trinitrobenzene sulfonic acid(TNBS)to establish IBD models. Colon mucosal lesions were observed after the mice were treated for 7 days. The levels of TNF-α,IL-6,IL-1β and TGF-β1 in colon were determined by enzyme-linked immunosorbent assay(ELISA). The expression of cytochrome P4503 A4(CYP3 A4),progesterone X receptor(PXR),TNF-α,IL-6 and IL-1β mRNA were detected by RT-PCR,and the expression of CYP3 A4 and nuclear transcription factor P65(NF-κBP65) were detected by Western Blot. Results Compared with the model group,the histopathology of colons were improved in the SASP and SC groups. SASP-and SC-treated mice had lower levels of TNF-α, IL-6, IL-1β, TGF-β1 and NF-κBP65 in the colonic tissues, and higher levels of CYP3 A4 and PXR mRNA expression than the model mice with dose dependence. Conclusion SC has beneficial effects on treating ulcerative colitis in mice mainly by activating PXR and CYP3 A and inhibiting the expression of NF-κBP65,and decreasing the production of inflammatory factors TNF-a,IL-6,IL-1β and TGF-β1.
Objective To investigate the therapeutic effect and mechanism of Jasminoside(SC)on Ulcerative colitis(UC)in mice. Methods The mice were randomly divided into normal group,model group,Sulfasalazine(SASP)group and SC high,middle and low dose groups. Except for normal group,mice in other groups were treated with2,4,6-trinitrobenzene sulfonic acid(TNBS)to establish IBD models. Colon mucosal lesions were observed after the mice were treated for 7 days. The levels of TNF-α,IL-6,IL-1β and TGF-β1 in colon were determined by enzyme-linked immunosorbent assay(ELISA). The expression of cytochrome P4503 A4(CYP3 A4),progesterone X receptor(PXR),TNF-α,IL-6 and IL-1β mRNA were detected by RT-PCR,and the expression of CYP3 A4 and nuclear transcription factor P65(NF-κBP65) were detected by Western Blot. Results Compared with the model group,the histopathology of colons were improved in the SASP and SC groups. SASP-and SC-treated mice had lower levels of TNF-α, IL-6, IL-1β, TGF-β1 and NF-κBP65 in the colonic tissues, and higher levels of CYP3 A4 and PXR mRNA expression than the model mice with dose dependence. Conclusion SC has beneficial effects on treating ulcerative colitis in mice mainly by activating PXR and CYP3 A and inhibiting the expression of NF-κBP65,and decreasing the production of inflammatory factors TNF-a,IL-6,IL-1β and TGF-β1.
引文
[1]HU D,WANG Y,CHEN Z,et al.The protective effect of piperine on dextran sulfate sodium induced inflammatory bowel disease and its relation with pregnane X receptor activation[J].Journal of ethnopharmacology 2015,169:109-123.
[2]DOU W,ZHANG J,LI H,et al.Plant flavonol isorhamnetin attenuates chemically induced inflammatory bowel disease via a PXR-dependent pathway[J].The Journal of nutritional biochemistry 2014,25(9):923-933.
[3]DE ALMEIDA A B A,SáNCHEZ-HIDALGO M,MARTíN A R,et al.Anti-inflammatory intestinal activity of Arctium lappa L.(Asteraceae)in TNBS colitis model[J].Journal of ethnopharmacology 2013,146(1):300-310.
[4]DRING M M,GOULDING C A,TRIMBLE V I,et al.The pregnane X receptor locus is associated with susceptibility to inflammatory bowel disease[J].Gastroenterology,2006,130(2):341-8;quiz 592.
[5]LEE E J,KIM C,KIM J Y,et al.Inhibition of LPS-induced inflammatory biomarkers by ethyl acetate fraction of Patrinia scabiosaefolia through suppression of NF-κB activation in RAW264.7 cells[J].Immunopharmacology and immunotoxicology,2012,34(2):282-291.
[6]孔令锋,李书渊.扭肚藤的生药学研究[J].广东药学院学报,2008,24(5):449-451.
[7]GAO Z H,YIN J Q,XIE X L,et al.Intracellular signaling mechanisms pharmacologiclaaction of Jasminum amplexicaule Buch.-Ham.(Oleaceae)on gastrointestinal secretion[J].Iran J Pharm Res,2014,13(3):959-965.
[8]宋雨鸿,黄洁春,韩亮,等.别旁茶提取物对实验性炎症性肠病大鼠的作用[J].广东药学院学报,2015,31(2):224-227.
[9]宋雨鸿,韩亮,王凤云.瑶药别旁茶总苷对溃疡性结肠炎小鼠模型干预的代谢组学初步研究[J].中药新药与临床药理,2016,27(3):399-403
[10]何影.JNK信号通路在小鼠TNBS结肠炎中的作用研究[D].四川大学,2007.
[11]BUSTIN S A,BENES V,GARSON J A,et al.The MIQE guidelines:minimum information for publication of quantitative real-time PCRexperiments[J].Clinical chemistry 2009,55(4):611-622.
[12]MEDINA-CONTRERAS O,GEEM D,LAUR O,et al.CX3CR1regulates intestinal macrophage homeostasis,bacterial translocation,and colitogenic Th17 responses in mice[J].The Journal of clinical investigation,2011,121(12):4787-4795.
[13]YAO J,WANG J Y,LIU L,et al.Anti-oxidant effects of resveratrol on mice with DSS-induced ulcerative colitis[J].Archives of Medical Research,2010,41(4):288-294.
[14]BLOKZIJL H,VANDER BORGHT S,BOK L I,et al.Decreased P-glycoprotein(P-gp/MDR1)expression in inflamed human intestinal epithelium is independent of PXR protein levels[J].Inflammatory bowel diseases 2007,13(6):710-720.
[15]LANGMANN T,MOEHLE C,MAUERER R,et al.Loss of detoxification in inflammatory bowel disease:dysregulation of pregnane X receptor target genes[J].Gastroenterology,2004,127(1):26-40.
[16]GOODWIN B,REDINBO M R,KLIEWER S A.Regulation of CYP3,a gene transcription by the pregnane X receptor[J].Annu Rev Pharmacol Toxicol,2002,42(1):1-23.
[17]ZHOU C,TABB M M,NELSON E L,et al.Mutual repression between steroid and xenobiotic receptor and NF-κB signaling pathways links xenobiotic metabolism and inflammation[J].The Journal of clinical investigation 2006,116(8):2280.
[18]ZHANG D K,CHENG L N,HUANG X L,et al.Tetrandrine ameliorates dextran-sulfate-sodium-induced colitis in mice through inhibition of nuclear factor-κB activation[J].International journal of colorectal disease,2009,24(1):5-12.
[19]XIE W,TIAN Y.Xenobiotic receptor meets NF-κB,a collision in the small bowel[J].Cell metabolism 2006,4(3):177-178.
[20]SHAH Y M,MA X,MORIMURA K,et al.Pregnane X receptor activation ameliorates DSS-induced inflammatory bowel disease via inhibition of NF-κB target gene expression[J].American Journal of Physiology-Gastrointestinal and Liver Physiology 2007,292(4):G1114-G1122.
[21]JOBIN C,BRADHAM C A,RUSSO M P,et al.Curcumin blocks cytokine-mediated NF-κB activation and proinflammatory gene expression by inhibiting inhibitory factor I-κB kinase activity[J].The Journal of Immunology 1999,163(6):3474-3483.
[22]SILVA M S D,SáNCHEZ-FIDALGO S,TALERO E,et al.Antiinflammatory intestinal activity of Abarema cochliacarpos(Gomes)Barneby&Grimes in TNBS colitis model[J].Journal of Ethnopharmacology 2010,128(2):467-475.
[23]QIDWAI T,KHAN F.Tumour necrosis factor gene polymorphism and disease prevalence[J].Scandinavian journal of immunology,2011,74(6):522-547.
[24]STEVENS C,WALZ G,SINGARAM C,et al.Tumor necrosis factor-α,interleukin-1β,and interleukin-6 expression in inflammatory bowel disease[J].Digestive diseases and sciences,1992,37(6):818-826.
[25]HEIDEMANN J,OGAWA H,DWINELL M B,et al.Angiogenic effects of interleukin 8(CXCL8)in human intestinal microvascular endothelial cells are mediated by CXCR2[J].The Journal of biological chemistry,2003,278(10):8508-8515.
[26]LI A,DUBEY S,VARNEY M L,et al.IL-8 directly enhanced endothelial cell survival,proliferation,and matrix metalloproteinases production and regulated angiogenesis[J].Journal of immunology(Baltimore,Md.:1950),2003,170(6):3369-3376.
[27]GHOSH A K,YUAN W,MORI Y,et al.Smad-dependent stimulation of type I collagen gene expression in human skin fibroblasts by TGF-beta involves functional cooperation with p300/CBP transcriptional coactivators[J].Oncogene,2000,19(31):3546-3555.
[28]HUME G,FOWLER E,LINCOLN D,et al.Angiotensinogen and transforming growth factorβ1:novel genes in the pathogenesis of Crohn’s disease[J].Journal of medical genetics 2006,43(10):e51-e51.
[29]TAMIZIFAR B,LANKARANI K,NAEIMI S,et al.Promoter polymorphism of transforming growth factor-β1 gene and ulcerative colitis[J].World journal of gastroenterology:WJG 2008,14(2):243.
[2]DOU W,ZHANG J,LI H,et al.Plant flavonol isorhamnetin attenuates chemically induced inflammatory bowel disease via a PXR-dependent pathway[J].The Journal of nutritional biochemistry 2014,25(9):923-933.
[3]DE ALMEIDA A B A,SáNCHEZ-HIDALGO M,MARTíN A R,et al.Anti-inflammatory intestinal activity of Arctium lappa L.(Asteraceae)in TNBS colitis model[J].Journal of ethnopharmacology 2013,146(1):300-310.
[4]DRING M M,GOULDING C A,TRIMBLE V I,et al.The pregnane X receptor locus is associated with susceptibility to inflammatory bowel disease[J].Gastroenterology,2006,130(2):341-8;quiz 592.
[5]LEE E J,KIM C,KIM J Y,et al.Inhibition of LPS-induced inflammatory biomarkers by ethyl acetate fraction of Patrinia scabiosaefolia through suppression of NF-κB activation in RAW264.7 cells[J].Immunopharmacology and immunotoxicology,2012,34(2):282-291.
[6]孔令锋,李书渊.扭肚藤的生药学研究[J].广东药学院学报,2008,24(5):449-451.
[7]GAO Z H,YIN J Q,XIE X L,et al.Intracellular signaling mechanisms pharmacologiclaaction of Jasminum amplexicaule Buch.-Ham.(Oleaceae)on gastrointestinal secretion[J].Iran J Pharm Res,2014,13(3):959-965.
[8]宋雨鸿,黄洁春,韩亮,等.别旁茶提取物对实验性炎症性肠病大鼠的作用[J].广东药学院学报,2015,31(2):224-227.
[9]宋雨鸿,韩亮,王凤云.瑶药别旁茶总苷对溃疡性结肠炎小鼠模型干预的代谢组学初步研究[J].中药新药与临床药理,2016,27(3):399-403
[10]何影.JNK信号通路在小鼠TNBS结肠炎中的作用研究[D].四川大学,2007.
[11]BUSTIN S A,BENES V,GARSON J A,et al.The MIQE guidelines:minimum information for publication of quantitative real-time PCRexperiments[J].Clinical chemistry 2009,55(4):611-622.
[12]MEDINA-CONTRERAS O,GEEM D,LAUR O,et al.CX3CR1regulates intestinal macrophage homeostasis,bacterial translocation,and colitogenic Th17 responses in mice[J].The Journal of clinical investigation,2011,121(12):4787-4795.
[13]YAO J,WANG J Y,LIU L,et al.Anti-oxidant effects of resveratrol on mice with DSS-induced ulcerative colitis[J].Archives of Medical Research,2010,41(4):288-294.
[14]BLOKZIJL H,VANDER BORGHT S,BOK L I,et al.Decreased P-glycoprotein(P-gp/MDR1)expression in inflamed human intestinal epithelium is independent of PXR protein levels[J].Inflammatory bowel diseases 2007,13(6):710-720.
[15]LANGMANN T,MOEHLE C,MAUERER R,et al.Loss of detoxification in inflammatory bowel disease:dysregulation of pregnane X receptor target genes[J].Gastroenterology,2004,127(1):26-40.
[16]GOODWIN B,REDINBO M R,KLIEWER S A.Regulation of CYP3,a gene transcription by the pregnane X receptor[J].Annu Rev Pharmacol Toxicol,2002,42(1):1-23.
[17]ZHOU C,TABB M M,NELSON E L,et al.Mutual repression between steroid and xenobiotic receptor and NF-κB signaling pathways links xenobiotic metabolism and inflammation[J].The Journal of clinical investigation 2006,116(8):2280.
[18]ZHANG D K,CHENG L N,HUANG X L,et al.Tetrandrine ameliorates dextran-sulfate-sodium-induced colitis in mice through inhibition of nuclear factor-κB activation[J].International journal of colorectal disease,2009,24(1):5-12.
[19]XIE W,TIAN Y.Xenobiotic receptor meets NF-κB,a collision in the small bowel[J].Cell metabolism 2006,4(3):177-178.
[20]SHAH Y M,MA X,MORIMURA K,et al.Pregnane X receptor activation ameliorates DSS-induced inflammatory bowel disease via inhibition of NF-κB target gene expression[J].American Journal of Physiology-Gastrointestinal and Liver Physiology 2007,292(4):G1114-G1122.
[21]JOBIN C,BRADHAM C A,RUSSO M P,et al.Curcumin blocks cytokine-mediated NF-κB activation and proinflammatory gene expression by inhibiting inhibitory factor I-κB kinase activity[J].The Journal of Immunology 1999,163(6):3474-3483.
[22]SILVA M S D,SáNCHEZ-FIDALGO S,TALERO E,et al.Antiinflammatory intestinal activity of Abarema cochliacarpos(Gomes)Barneby&Grimes in TNBS colitis model[J].Journal of Ethnopharmacology 2010,128(2):467-475.
[23]QIDWAI T,KHAN F.Tumour necrosis factor gene polymorphism and disease prevalence[J].Scandinavian journal of immunology,2011,74(6):522-547.
[24]STEVENS C,WALZ G,SINGARAM C,et al.Tumor necrosis factor-α,interleukin-1β,and interleukin-6 expression in inflammatory bowel disease[J].Digestive diseases and sciences,1992,37(6):818-826.
[25]HEIDEMANN J,OGAWA H,DWINELL M B,et al.Angiogenic effects of interleukin 8(CXCL8)in human intestinal microvascular endothelial cells are mediated by CXCR2[J].The Journal of biological chemistry,2003,278(10):8508-8515.
[26]LI A,DUBEY S,VARNEY M L,et al.IL-8 directly enhanced endothelial cell survival,proliferation,and matrix metalloproteinases production and regulated angiogenesis[J].Journal of immunology(Baltimore,Md.:1950),2003,170(6):3369-3376.
[27]GHOSH A K,YUAN W,MORI Y,et al.Smad-dependent stimulation of type I collagen gene expression in human skin fibroblasts by TGF-beta involves functional cooperation with p300/CBP transcriptional coactivators[J].Oncogene,2000,19(31):3546-3555.
[28]HUME G,FOWLER E,LINCOLN D,et al.Angiotensinogen and transforming growth factorβ1:novel genes in the pathogenesis of Crohn’s disease[J].Journal of medical genetics 2006,43(10):e51-e51.
[29]TAMIZIFAR B,LANKARANI K,NAEIMI S,et al.Promoter polymorphism of transforming growth factor-β1 gene and ulcerative colitis[J].World journal of gastroenterology:WJG 2008,14(2):243.