YZG-330通过P38 MAPK信号通路调节TRPM8离子通道发挥降温作用  

YZG-330 regulates the TRPM8 ion channel through the P38 MAPK signaling pathway to reduce body temperature in mice

作　　者：朱子玉 虞远智 袁月 张建军 ZHU Zi-yu;YU Yuan-zhi;YUAN Yue;ZHANG Jian-jun(Beijing Key Laboratory of New Drug Mechanisms and Pharmacological Evaluation Study,Institute of Materia Medica,Chinese Academy of Medical Sciences and Peking Union Medical College,Beijing 100050,China)

机构地区：[1]中国医学科学院、北京协和医学院药物研究所,新药作用机制研究和药效评价北京市重点实验室,北京100050

出　　处：《药学学报》2022年第5期1336-1343,共8页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金项目资助(81773715);中国医学科学院创新工程项目(2021-I2M-1-028)。

摘　　要：实验室前期研究发现化合物YZG-330能够使小鼠降低体温,该作用可被腺苷A1受体(adenosine A1 receptor,A1R)拮抗剂DPCPX拮抗,本文基于A1R下游信号通路对YZG-330的作用机制进行进一步探究。通过测量小鼠肛温进行YZG-330药效学评价;采用Western blot技术检测小鼠下丘脑组织匀浆中瞬时受体电位(transient receptor potential,TRP)离子通道和P38蛋白及其磷酸化水平;通过Ca^(2+)荧光探针Fluo-3AM负载细胞,检测YZG-330对小鼠下丘脑细胞内Ca^(2+)含量的影响。YZG-330可剂量依赖性地降低小鼠体温,P38选择性抑制剂SB-203580(20 mg·kg^(-1),i.p.)可显著抑制YZG-330的降温作用。TRPM8拮抗剂(每只小鼠0.1μg,i.c.v.)可以部分拮抗YZG-330(0.25 mg·kg^(-1)和1 mg·kg^(-1),i.p.)的降温效果。腹腔注射给予小鼠YZG-330(2 mg·kg^(-1),i.p.)可显著上调小鼠下丘脑组织中P38磷酸化水平,A1R拮抗剂DPCPX(5 mg·kg^(-1),i.g.)可有效抑制该变化;YZG-330还可显著上调小鼠下丘脑组织中TRPM8表达量,SB-203580可非常显著地抑制YZG-330引起的TRPM8上调;YZG-330(0.1~10μmol·L^(-1))可剂量依赖性地升高小鼠下丘脑细胞内Ca^(2+)浓度,A1R抑制剂DPCPX(0.5和1μmol·L^(-1))与TRPM8拮抗剂(1μmol·L^(-1))均可抑制YZG-330的升高胞内Ca^(2+)浓度的作用。综上所述,YZG-330发挥降温作用的机制之一是通过激活A1R,促进P38蛋白磷酸化,进而上调TRPM8离子通道表达量并激活该通道开启,导致胞内Ca^(2+)含量上升,通过负调节下丘脑体温调定点导致体温下降。本文中所有动物实验均获得中国医学科学院药物研究所伦理委员会批准。Preliminary research in our laboratory found that compound YZG-330 can reduce mouse body temperature,which could be blocked by adenosine A1 receptor(A1R)antagonist DPCPX.Based on the downstream signaling pathway of the A1R,the mechanism by which YZG-330 lowers body temperature was further studied.The pharmacodynamics of YZG-330 was evaluated by measuring the rectal temperature;expression of the transient receptor potential(TRP)ion channel,the P38 protein and its phosphorylated form in mouse hypothalamic homogenate were detected by Western blotting.A Ca2+fluorescent probe,Fluo-3AM,was added to cells to detect the effect of YZG-330 on the Ca2+content of mouse hypothalamic cells.YZG-330 dose-dependently reduced the body temperature in mice,and the selective P38 inhibitor SB-203580(20 mg·kg^(-1),i.p.)significantly inhibited the hypothermic effect of YZG-330.A TRPM8 antagonist 2(0.1μg per mouse,i.c.v.)markedly attenuated the hypothermic effect of YZG-330(0.25 or 1 mg·kg^(-1),i.p.).YZG-330(2 mg·kg^(-1),i.p.)significantly increased the phosphorylation of P38,an effect that could be attenuated by the A1R antagonist DPCPX(5 mg·kg^(-1),i.g.)in mouse hypothalamus.In addition,YZG-330 also prominently enhanced the expression of TRPM8,which could be blocked by SB-203580;YZG-330(0.1-10μmol·L^(-1))increased intracellular Ca2+concetration in mouse hypothalamic cells in a dose-dependent manner,and was inhibited by the A1R inhibitor DPCPX(0.5 and 1μmol·L^(-1))and TRPM8 antagonist 2(1μmol·L^(-1)).In conclusion,YZG-330 exerts its hypothermic effect by activating the A1R to promote the phosphorylation of P38 protein and thereby up-regulating the expression and activity of the TRPM8 ion channel,resulting in increased intracellular Ca2+concentration to stimulate mouse hypothalamus cells to down-regulate body temperature.All animal experiments were approved by the Ethics Committee of the Institute of Materia Medica,Chinese Academy of Medical Sciences.

关 键 词：腺苷A1受体 丝裂原活化蛋白激酶P38 瞬时受体电位离子通道亚家族8 Ca^(2+)离子 

分 类 号：R966[医药卫生—药理学]