健脾理气方对功能性消化不良大鼠十二指肠MLCK和PGE2-cAMP通路的影响
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摘要
目的观察健脾理气方对功能性消化不良(functional dyspepsia,FD)模型大鼠十二指肠肌球蛋白轻链激酶(myosin light chain kinase,MLCK)和前列腺素E2 (prostaglandin E2,PGE2)-环磷酸腺苷(cyclic adenosine monophosphate,c AMP)通路的影响。方法 18只SD雄性大鼠采用数字表法随机分为正常组、模型组及中药治疗组(每组6只),以夹尾应激法建立功能性消化不良动物模型,正常组及模型组大鼠均给予0. 9%(质量分数)氯化钠注射液灌胃,中药治疗组给予健脾理气方水煎剂灌胃。取大鼠十二指肠组织进行HE染色,观察组织形态,同时应用定量聚合酶链式反应(quantitative real time polymerase chain reaction,qRTPCR)检测MLCK和PGE2受体EP1、EP4 mRNA含量,酶联免疫吸附(enzyme linked immunosorbent assay,ELISA)法测定PGE2和c AMP的表达。结果模型组大鼠十二指肠MLCK表达升高,PGE2、EP4、c AMP表达下降,EP1未见明显异常,健脾理气方治疗后可明显抑制MLCK表达,升高PGE2、EP4和c AMP表达。结论健脾理气方可以抑制MLCK mRNA表达,同时上调PGE2-EP4-c AMP信号通路表达,这可能是健脾理气方治疗FD的机制之一。
Objective To explore the effect of Jianpi Liqi formula( JPLQ) on myosin light chain kinase( MLCK) and prostaglandin E2-cyclic adenosine monophosphate( PGE2-cAMP) signaling pathway in the duodenum of functional dyspepsia( FD) rats. Methods Eighteen SD male rats were randomly divided into control group,FD model group and JPLQ group in a 1 ∶ 1 ∶ 1 ratio. Rats with FD were generated via stress-based tail clamping. Stressed rats were given JPLQ or saline in the drinking water,and the morphology of duodenum was measured by hematoxylin-eosin( HE) staining,mRNA expression of MLCK and PGE2 receptor subtype 1 and 4( EP1,EP4) were analyzed with quantitative real time polymerase chain reaction( qRT-PCR). PGE2 and c AMP expression was measured with enzymelinked immunosorbent assay( ELISA). Results MLCK mRNA expression in duodenum was decreased in the JPLQ-treated stressed rats as compared to the control-treated stressed rats. PGE2,EP4 and c AMP in duodenum was increased in the JPLQ-treated stressed rats as compared to the control-treated stressed rats. There was no difference of EP1 expression among the three groups. Conclusion The underlying mechanisms of JPLQ formula on FD are at least partially through the attenuation of MLCK signaling pathway and the improvement of PGE2-EP4-cAMP signaling pathway.
Objective To explore the effect of Jianpi Liqi formula( JPLQ) on myosin light chain kinase( MLCK) and prostaglandin E2-cyclic adenosine monophosphate( PGE2-cAMP) signaling pathway in the duodenum of functional dyspepsia( FD) rats. Methods Eighteen SD male rats were randomly divided into control group,FD model group and JPLQ group in a 1 ∶ 1 ∶ 1 ratio. Rats with FD were generated via stress-based tail clamping. Stressed rats were given JPLQ or saline in the drinking water,and the morphology of duodenum was measured by hematoxylin-eosin( HE) staining,mRNA expression of MLCK and PGE2 receptor subtype 1 and 4( EP1,EP4) were analyzed with quantitative real time polymerase chain reaction( qRT-PCR). PGE2 and c AMP expression was measured with enzymelinked immunosorbent assay( ELISA). Results MLCK mRNA expression in duodenum was decreased in the JPLQ-treated stressed rats as compared to the control-treated stressed rats. PGE2,EP4 and c AMP in duodenum was increased in the JPLQ-treated stressed rats as compared to the control-treated stressed rats. There was no difference of EP1 expression among the three groups. Conclusion The underlying mechanisms of JPLQ formula on FD are at least partially through the attenuation of MLCK signaling pathway and the improvement of PGE2-EP4-cAMP signaling pathway.
引文
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[14]林晓春,陈育尧,白殊同,等.甘草总黄酮对慢性浅表性胃炎大鼠胃粘膜损伤的保护作用[J].南方医科大学学报,2013,33(2):299-304.
[2] Vanheel H,Vicario M,Vanuytsel T,et al. Impaired duodenal mucosal integrity and low-grade inflammation in functional dyspepsia[J]. Gut,2014,63(2):262-271.
[3] Krug S M,Schulzke J D,Fromm M. Tight junction,selective permeability,and related diseases[J]. Semin Cell Dev Biol,2014,36:166-176.
[4] Zhang S S,Zhao L Q,Wang H B,et al. Efficacy of modified LiuJunZi decoction on functional dyspepsia of spleen-deficiency and qi-stagnation syndrome:a randomized controlledtrial[J]. BM C Complement Altern M ed,2013,13:54.
[5] Zhou H Y,Zhu H,Yao X M,et al. Metformin regulates tight junction of intestinal epithelial cells via M LCK-M LC signaling pathw ay[J]. Eur Rev M ed Pharmacol Sci,2017,21(22):5239.
[6] Gregory M,Cyr D G. Effects of prostaglandin E2 on gap junction protein alpha 1 in the rat epididymis[J]. Biol Reprod,2019,100(1):123-132.
[7]施新猷.现代医学实验动物学[M].北京:人民军医出版社,2000:333-335.
[8] Hong W,Nianhui Z,Ying C,et al. OTA induces intestinal epithelial barrier dysfunction and tight junction disruption in IPEC-J2 cells through ROS/Ca2+-mediated M LCK activation[J]. Environ Pollut,2018,242:106-112.
[9] Moeser A J,Haskell M M,Shifflett D E,et al. ClC-2chloride secretion mediates prostaglandin-induced recovery of barrier function in ischemia-injured porcine ileum[J].Gastroenterology,2004,127(3):802-815.
[10] Takeuchi K, Amagase K. Roles of cyclooxygenase,prostaglandin E2 and EP receptors in mucosal protection and ulcer healing in the gastrointestinal tract[J]. Curr Pharm Des,2018,24(18):2002-2011.
[11]Rodriguez-Lagunas M J,Martin-Venegas R,Moreno J J,et al. PGE2 promotes Ca2+-mediated epithelial barrier disruption through EP1 and EP4 receptors in Caco-2 cell monolayers[J]. Am J Physiol Cell Physiol,2010,299(2):C324-334.
[12] Su Y,Huang X,Raskovalova T,et al. Cooperation of adenosine and prostaglandin E2(PGE2)in amplification of c AM P-PKA signaling and immunosuppression[J]. Cancer Immunol Immunother,2008,57(11):1611-1623.
[13]尚秋辰.白术多糖对大肠杆菌腹泻模型小鼠肠道黏膜修复机理研究[D].扬州:扬州大学,2017.
[14]林晓春,陈育尧,白殊同,等.甘草总黄酮对慢性浅表性胃炎大鼠胃粘膜损伤的保护作用[J].南方医科大学学报,2013,33(2):299-304.