钙敏感受体及下游PLC/TRPM5介导苯丙氨酸刺激猪十二指肠分泌胆囊收缩素

L-phenylalanine-induced cholecystokinin secretion mediated by calcium-sensing receptor and downstream PLC/TRPM5 in swine duodenum

[目的]本文旨在探究苯丙氨酸(Phe)体外灌流对猪十二指肠组织胆囊收缩素(CCK)分泌的影响,以及钙敏感受体(CaSR)、鲜味受体(T1R1/T1R3)、下游磷脂酶C(PLC)和瞬时受体电位离子通道蛋白5(TRPM5)在该过程中的作用。[方法]以猪十二指肠为研究对象,利用免疫印迹技术检测CaSR在猪十二指肠上的表达;在此基础上,利用组织体外灌流系统探究Phe对猪十二指肠CCK分泌的影响,再通过抑制剂或激活剂揭示CaSR、T1R1/T1R3以及PLC和TRPM5对CCK分泌的作用。[结果]猪十二指肠表达CaSR;50 mmol·L^(-1) L-Phe而非D-Phe能够显著刺激CCK的分泌(P<0.05);加入CaSR抑制剂NPS 2143(25μmol·L^(-1))或calhex 231(20μmol·L^(-1))均可显著抑制L-Phe诱导的CCK分泌(P<0.05),但加入T1R1/T1R3激活剂肌苷酸(IMP,2.5 mmol·L^(-1))或抑制剂lactisole(5 mmol·L^(-1))后CCK的分泌并无显著变化(P>0.05);通路下游PLC抑制剂U-73122(10μmol·L^(-1))和TRPM5抑制剂氧化三苯基磷(TPPO,100μmol·L^(-1))均可显著抑制L-Phe诱导的CCK分泌(P<0.05)。[结论]L-Phe通过激活CaSR以及下游PLC/TRPM5刺激猪十二指肠组织分泌CCK。

[Objectives]This study aimed to investigate the effect of phenylalanine(Phe)on cholecystokinin(CCK)secretion in swine duodenal perfusion in vitro.In addition,the potential roles of the calcium-sensing receptor(CaSR),umami receptor(type 1 taste receptor 1/3,T1R1/T1R3)and downstream phospholipase C(PLC)and transient receptor potential subfamily M member 5(TRPM5)were also demonstrated.[Methods]Western blot was used to detect whether CaSR expressed in swine duodenum.Based on the above result,the effect of Phe on CCK secretion was explored by using in vitro perfusion system and the roles of the CaSR,T1R1/T1R3 and downstream PLC,TRPM5 on CCK secretion were investigated by the addition of inhibitors or agonists.[Results]CaSR protein was expressed in pig duodenum,and 50 mmol·L^(-1) L-Phe instead of D-Phe stimulated CCK secretion(P<0.05).CaSR inhibitors NPS 2143(25μmol·L^(-1))or calhex 231(20μmol·L^(-1))suppressed CCK secretion(P<0.05),however,T1R1/T1R3 agonist inosine monophosphate(IMP,2.5 mmol·L^(-1))or inhibitor lactisole(5 mmol·L^(-1))had no effect on CCK secretion(P>0.05).The addition of PLC antagonist U-73122(10μmol·L^(-1))or TRPM5 inhibitor triphenylphophine oxide(TPPO,100μmol·L^(-1))inhibited CCK secretion(P<0.05).[Conclusions]CaSR and downstream PLC/TRPM5 played an important role in L-Phe induced CCK secretion from swine duodenum in vitro.