小檗碱与6-姜烯酚配伍对溃疡性结肠炎小鼠结肠上皮细胞Notch信号通路的影响
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摘要
目的探讨黄连、干姜有效成分小檗碱、6-姜烯酚单独和联合应用对溃疡性结肠炎小鼠结肠上皮细胞Notch信号通路的影响。方法 50只昆明小鼠随机选取10只为对照组,其余40只采用2%葡聚糖硫酸钠(DSS)诱导溃疡性结肠炎模型。成模后随机分为模型组、小檗碱组、6-姜烯酚组、小檗碱+6-姜烯酚组,ig给予相应药物,给药20 d后取结肠组织。免疫荧光法检测结肠上皮组织Hes-1、Math-1、MUC2、人类嗅素蛋白(olfm4)的表达,实时荧光定量PCR(qRT-PCR)、Western blotting法检测结肠上皮组织Notch-1、Hes-1、Math-1m RNA和蛋白的表达。结果与模型组比较,6-姜烯酚组、小檗碱组、小檗碱+6-姜烯酚组小鼠结肠上皮组织Notch-1、Hes-1蛋白和m RNA表达水平显著降低,Math-1蛋白和m RNA表达水平显著升高,olfm4蛋白表达显著降低,MUC2蛋白表达水平显著升高;3组间比较,小檗碱+6-姜烯酚组作用最明显。结论 6-姜烯酚、小檗碱均能够有效修复受损结肠黏膜,治疗溃疡性结肠炎,小檗碱和6-姜烯酚联合应用药效优于单独应用,其机制与抑制Notch通路的过度活化进而调节结肠上皮细胞增殖和分化相关。
Objective To investigate the effects of berberine and 6-shogaol on Notch signaling pathway in colonic epithelial cells of mice with ulcerative colitis. Methods A total of 50 Kunming mice were randomly selected as normal group, and the remaining 40 mice were treated with po 2% DSS to induce ulcerative colitis. The rats were randomly divided into model group, berberine group,6-shogaol group, and berberine + 6-shogaol group. The corresponding drugs were given by ig administration. Colon tissues were taken after 20 d of administration. The protein expression of Hes-1, Math-1, MUC2, and olfm4 in colon epithelium was detected by immunofluorescence method, and the gene and protein expression of Notch-1, Hes-1, Math-1 in colon epithelium were detected by q RT-PCR and Western blotting. Results Compared with the model group, the protein and mRNA expression of Notch-1, Hes-1, and olfm4 in colon epithelium of mice in 6-shogaol group, berberine group, and berberine + 6-shogaol group were significantly decreased,while the expression of Math-1 and MUC2 were significantly increased. Among three groups, berberine + 6-shogaol group showed the best effect. Conclusion Both 6-shogaol and berberine can effectively repair damaged colonic mucosa and treat ulcerative colitis. The combined use of berberine and 6-shogaol is more effective than the single application. Its mechanism is related to inhibiting the over-activation of Notch pathway and regulating the proliferation and differentiation of colonic epithelium.
Objective To investigate the effects of berberine and 6-shogaol on Notch signaling pathway in colonic epithelial cells of mice with ulcerative colitis. Methods A total of 50 Kunming mice were randomly selected as normal group, and the remaining 40 mice were treated with po 2% DSS to induce ulcerative colitis. The rats were randomly divided into model group, berberine group,6-shogaol group, and berberine + 6-shogaol group. The corresponding drugs were given by ig administration. Colon tissues were taken after 20 d of administration. The protein expression of Hes-1, Math-1, MUC2, and olfm4 in colon epithelium was detected by immunofluorescence method, and the gene and protein expression of Notch-1, Hes-1, Math-1 in colon epithelium were detected by q RT-PCR and Western blotting. Results Compared with the model group, the protein and mRNA expression of Notch-1, Hes-1, and olfm4 in colon epithelium of mice in 6-shogaol group, berberine group, and berberine + 6-shogaol group were significantly decreased,while the expression of Math-1 and MUC2 were significantly increased. Among three groups, berberine + 6-shogaol group showed the best effect. Conclusion Both 6-shogaol and berberine can effectively repair damaged colonic mucosa and treat ulcerative colitis. The combined use of berberine and 6-shogaol is more effective than the single application. Its mechanism is related to inhibiting the over-activation of Notch pathway and regulating the proliferation and differentiation of colonic epithelium.
引文
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[10]Zhang M,Xu C,Liu D,et al.Oral delivery of nanoparticles loaded with ginger active compound,6-shogaol,attenuates ulcerative colitis and promotes wound healing in a murine model of ulcerative colitis[J].J Crohns Colitis,2017,12(2):217-229.
[11]Zhang M,Viennois E,Prasad M,et al.Edible ginger-derived nanoparticles:A novel therapeutic approach for the prevention and treatment of inflammatory bowel disease and colitis-associated cancer[J].Biomaterials,2016,doi:10.1016/j.biomaterials.2016.06.018.
[12]Li Y H,Xiao H T,Hu D D,et al.Berberine ameliorates chronic relapsing dextran sulfate sodium induced colitis in C57BL/6 Mice by suppressing Th17 responses[J].Pharmacol Res,2016,doi:10.1016/j.phrs.2016.02.010.
[13]Wang J,Yao M,Wang Y,et al.6-Shogaol ameliorates injury to the intestinal mucosa and increases survival after high-dose abdominal irradiation[J].J Funct Foods,2017,doi:10.1016/j.jff.2017.06.054.
[14]Lange K M D,Moutsianas L,Lee J C,et al.Genome-wide association study implicates immune activation of multiple integrin genes in inflammatory bowel disease[J].Nat Gene,2017,49(2):256-261.
[15]中华医学会消化病学分会炎症性肠病学组.炎症性肠病诊断与治疗的共识意见[J].中国实用内科杂志,2018,38(9):796-813.
[16]Nandagopal N,Santat L A,Lebon L,et al.Dynamic ligand discrimination in the Notch signaling pathway[J].Cell,doi:10.1016/j.cell.2018.01.002.
[17]Siebel C,Lendahl U.Notch signaling in development,tissue homeostasis,and disease[J].Physiol Rev,2017,97(4):1235-1294.
[18]Zheng X,Tsuchiya K,Okamoto R,et al.Suppression of hath1 gene expression directly regulated by hes1 via notch signaling is associated with goblet cell depletion in ulcerative colitis[J].Inflamm Bowel Dis,2011,17(11):2251-2260.
[19]Hinoda Y,Akashi H,Suwa T,et al.Immunohistochemical detection of MUC2 mucin core protein in ulcerative colitis[J].J Clin Lab Anal,2015,12(3):150-153.
[20]Jang H,Park S,Lee J,et al.Rebamipide alleviates radiation-induced colitis through improvement of goblet cell differentiation in mice[J].J Gastroenterol Hepatol,2018,33(4):876-886.
[21]Radtke F,Macdonald H R,Tacchinicottier F.Regulation of innate and adaptive immunity by Notch[J].Nat Rev Immunol,2013,13(6):427-437.
[22]Thomas A.Wynn,Kevin M.Vannella.Macrophages in Tissue Repair,Regeneration,and Fibrosis[J].Immunity,2016,44(3):450-462.
[23]周萍,彭咏波,李萍.6-姜烯酚抗肿瘤机制研究进展[J].中南药学,2012,10(7):536-538.
[24]Yun K W,Hong Y J,Yong H Y,et al.6-Shogaol protects against oxidized LDL-induced endothelial injruries by inhibiting oxidized LDL-evoked LOX-1 signaling[J].Evid-Based Compl Alt,2013,doi:10.1155/2013/503521.
[25]Annamalai G,Suresh K.[6]-Shogaol attenuates inflammation,cell proliferation via modulate NF-κB and AP-1 oncogenic signaling in 7,12-dimethylbenz[a]anthracene induced oral carcinogenesis[J].Biomed Pharmacother,2018,doi:10.1016/j.biopha.2017.12.009.
[26]Poorrostami A,Farokhi F,Heidari R.Effect of hydroalcoholic extract of ginger on the liver of epileptic female rats treated with lamotrigine[J].Avicenna JPhytomed,2014,4(4):276-286.
[27]陈磊,胡子成,周毅,等.6-姜烯酚通过下调TLR4/NF-κB通路抑制血管平滑肌细胞增殖[J].第三军医大学学报,2016,38(3):245-250.
[28]Yu X T,Xu Y F,Huang Y F,et al.Berberrubine attenuates mucosal lesions and inflammation in dextran sodium sulfate-induced colitis in mice[J].PLoS One,2018,13(3):e0194069.
[29]盖晓红,刘素香,任涛,等.黄连的化学成分及药理作用研究进展[J].中草药,2018,49(20):4919-4927.
[2]李筱,沈秀云,高昳,等.老年与非老年溃疡性结肠炎患者临床特征对比研究[J].胃肠病学,2017,22(7):426-428.
[3]Zheng X,Tsuchiya K,Okamoto R,et al.Suppression of hath1 gene expression directly regulated by hes1 via notch signaling is associated with goblet cell depletion in ulcerative colitis[J].Inflamm Bowel Dis,2011,17(11):2251-2260.
[4]刘玉珠,徐芳,熊腊根.“通调气血,寒热并用”法结合西药治疗溃疡性结肠炎的疗效观察[J].中国中西医结合消化杂志,2015,23(7):493-495.
[5]貟登辉.基于知网文献研究中药治疗UC的临床用药规律[D].沈阳:辽宁中医药大学,2016.
[6]郭雁冰.中医药抗溃疡性结肠炎复发的研究[D].北京:北京中医药大学,2006.
[7]陈岩.干姜黄芩黄连人参汤治疗慢性结肠炎56例疗效观察[J].新中医,2010,42(10):38-39.
[8]李阳,郝艺照,傅熠俊,等.黄连-干姜药对预防DSS诱导的小鼠结肠炎作用及其机制[J].中国实验方剂学杂志,2017,23(15):162-167.
[9]王婷婷,钟凌云.不同姜汁制黄连抑制胃黏膜损伤方面的机制分析[J].中国实验方剂学杂志,2017,23(12):26-30.
[10]Zhang M,Xu C,Liu D,et al.Oral delivery of nanoparticles loaded with ginger active compound,6-shogaol,attenuates ulcerative colitis and promotes wound healing in a murine model of ulcerative colitis[J].J Crohns Colitis,2017,12(2):217-229.
[11]Zhang M,Viennois E,Prasad M,et al.Edible ginger-derived nanoparticles:A novel therapeutic approach for the prevention and treatment of inflammatory bowel disease and colitis-associated cancer[J].Biomaterials,2016,doi:10.1016/j.biomaterials.2016.06.018.
[12]Li Y H,Xiao H T,Hu D D,et al.Berberine ameliorates chronic relapsing dextran sulfate sodium induced colitis in C57BL/6 Mice by suppressing Th17 responses[J].Pharmacol Res,2016,doi:10.1016/j.phrs.2016.02.010.
[13]Wang J,Yao M,Wang Y,et al.6-Shogaol ameliorates injury to the intestinal mucosa and increases survival after high-dose abdominal irradiation[J].J Funct Foods,2017,doi:10.1016/j.jff.2017.06.054.
[14]Lange K M D,Moutsianas L,Lee J C,et al.Genome-wide association study implicates immune activation of multiple integrin genes in inflammatory bowel disease[J].Nat Gene,2017,49(2):256-261.
[15]中华医学会消化病学分会炎症性肠病学组.炎症性肠病诊断与治疗的共识意见[J].中国实用内科杂志,2018,38(9):796-813.
[16]Nandagopal N,Santat L A,Lebon L,et al.Dynamic ligand discrimination in the Notch signaling pathway[J].Cell,doi:10.1016/j.cell.2018.01.002.
[17]Siebel C,Lendahl U.Notch signaling in development,tissue homeostasis,and disease[J].Physiol Rev,2017,97(4):1235-1294.
[18]Zheng X,Tsuchiya K,Okamoto R,et al.Suppression of hath1 gene expression directly regulated by hes1 via notch signaling is associated with goblet cell depletion in ulcerative colitis[J].Inflamm Bowel Dis,2011,17(11):2251-2260.
[19]Hinoda Y,Akashi H,Suwa T,et al.Immunohistochemical detection of MUC2 mucin core protein in ulcerative colitis[J].J Clin Lab Anal,2015,12(3):150-153.
[20]Jang H,Park S,Lee J,et al.Rebamipide alleviates radiation-induced colitis through improvement of goblet cell differentiation in mice[J].J Gastroenterol Hepatol,2018,33(4):876-886.
[21]Radtke F,Macdonald H R,Tacchinicottier F.Regulation of innate and adaptive immunity by Notch[J].Nat Rev Immunol,2013,13(6):427-437.
[22]Thomas A.Wynn,Kevin M.Vannella.Macrophages in Tissue Repair,Regeneration,and Fibrosis[J].Immunity,2016,44(3):450-462.
[23]周萍,彭咏波,李萍.6-姜烯酚抗肿瘤机制研究进展[J].中南药学,2012,10(7):536-538.
[24]Yun K W,Hong Y J,Yong H Y,et al.6-Shogaol protects against oxidized LDL-induced endothelial injruries by inhibiting oxidized LDL-evoked LOX-1 signaling[J].Evid-Based Compl Alt,2013,doi:10.1155/2013/503521.
[25]Annamalai G,Suresh K.[6]-Shogaol attenuates inflammation,cell proliferation via modulate NF-κB and AP-1 oncogenic signaling in 7,12-dimethylbenz[a]anthracene induced oral carcinogenesis[J].Biomed Pharmacother,2018,doi:10.1016/j.biopha.2017.12.009.
[26]Poorrostami A,Farokhi F,Heidari R.Effect of hydroalcoholic extract of ginger on the liver of epileptic female rats treated with lamotrigine[J].Avicenna JPhytomed,2014,4(4):276-286.
[27]陈磊,胡子成,周毅,等.6-姜烯酚通过下调TLR4/NF-κB通路抑制血管平滑肌细胞增殖[J].第三军医大学学报,2016,38(3):245-250.
[28]Yu X T,Xu Y F,Huang Y F,et al.Berberrubine attenuates mucosal lesions and inflammation in dextran sodium sulfate-induced colitis in mice[J].PLoS One,2018,13(3):e0194069.
[29]盖晓红,刘素香,任涛,等.黄连的化学成分及药理作用研究进展[J].中草药,2018,49(20):4919-4927.