慢性束缚应激大鼠下丘脑ARC中Ob-R、AgRP及NPY的表达及逍遥散的调节作用
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摘要
目的:探讨慢性束缚应激大鼠下丘脑弓状核(ARC)瘦素受体(Ob-R)、刺鼠基因相关蛋白(AgRP)、神经肽Y(NPY)变化及中药逍遥散的影响。方法:120只SD实验大鼠随机分为正常组、7d模型组、21d模型组、中药组。采用束缚制动法复制应激大鼠模型,中药组在每日束缚制动前灌服中药逍遥散21d。观察实验大鼠摄食量及体质量变化;ELISA方法检测大鼠外周血Leptin含量;Western Blot和qRT-PCR法分别检测各组大鼠下丘脑ARC中Ob-R、AgRP和NPY蛋白及基因表达。结果:在应激第7、14、21天,21d模型组大鼠体质量、摄食量均显著性少于正常组(P<0.01);而中药组大鼠体质量增长明显优于21d模型组(尤其应激第7、21天)(P<0.05),摄食量在应激第14、21天较21d模型组增加明显(P<0.05)。与正常组比较,7d模型组和21d模型组大鼠血清Leptin含量、下丘脑ARC中Ob-R蛋白及基因表达显著性升高(P<0.05,P<0.01),而AgRP和NPY蛋白及基因表达则显著降低(P<0.05,P<0.01);中药组大鼠血清Leptin含量、下丘脑ARC中Ob-R蛋白及基因表达较21d模型组降低(P<0.05,P<0.01),而AgRP和NPY蛋白及基因表达则显著提高(P<0.01),中药组与7d模型组比较上述指标差异无统计学意义。结论:慢性束缚应激大鼠食欲下降、体质量增长缓慢等躯体不适症状可能与其下丘脑ARC中Ob-R表达增多,AgRP、NPY合成、分泌减少有关,中药逍遥散能通过调节ARC中Ob-R、AgRP和NPY蛋白及基因表达水平,有效改善慢性束缚应激大鼠上述躯体不适症状。
Objective: To investigate the changes of leptin receptor(Ob-R), Agouti gene-related protein(AgRP) and neuropeptide Y(NPY) in rat hypothalamic arcuate nucleus(ARC) after chronic immobilization stress and the effect of Xiaoyao Powder. Methods: One-hundred and twenty Sprague-Dawley(SD) rats were randomly divided into the following groups: normal control group(ZC group), 7-day model group(7-MX group), 21-day model group(21-MX group) and XYS-treated group(ZY group). The model was established by immobilization stress for 7 or 21 days, and the rats in the ZY group were intragastrically administered with Xiaoyao Powder before exposed to immobilization stress every day for 21 days. The food intake and body weight were observed. The concentration of leptin in serum was detected by ELISA method, the expressions of Ob-R, AgRP and NPY in the ARC were assayed by Western Blot and qRT-PCR methods. Results: On the 7 th, 14 th and 21 st days of stress, the body weight and food intake of rats in 21-MX group were lower than those in ZC group(P<0.01), while the growth of body weight of rats in ZY group was significantly better than those in 21-MX group(especially on the 7 th and 21 st days of stress)(P<0.05), and the food intake in ZY group increased significantly(especially on the 14 th and 21 st days of stress)(P<0.05). The serum leptin and expression levels of Ob-R protein and gene in 7-MX group and 21-MX group were obviously higher than those in ZC group(P<0.05, P<0.01), and the protein and gene expression levels of AgRP and NPY were significantly decreased(P<0.05, P<0.01). ZY group reduced serum leptin and Ob-R expression in ARC compared with 21-MX group(P<0.05, P<0.01), meanwhile, ZY group remarkably up-regulated AgRP and NPY expression in the ARC(P<0.01), but there was no difference between ZY group and 7-MX group. Conclusion: The somatic discomfort symptoms, such as loss of appetite and slowing growth in body weight in 21-MX group, may be related to increase in Ob-R and decrease in AgRP and NPY expression in hypothalamic ARC. Xiaoyao Powder can effectively improve the body discomfort symptoms of rats after chronic immobilization stress by regulating the protein and gene expression levels of Ob-R, AgRP and NPY in ARC.
Objective: To investigate the changes of leptin receptor(Ob-R), Agouti gene-related protein(AgRP) and neuropeptide Y(NPY) in rat hypothalamic arcuate nucleus(ARC) after chronic immobilization stress and the effect of Xiaoyao Powder. Methods: One-hundred and twenty Sprague-Dawley(SD) rats were randomly divided into the following groups: normal control group(ZC group), 7-day model group(7-MX group), 21-day model group(21-MX group) and XYS-treated group(ZY group). The model was established by immobilization stress for 7 or 21 days, and the rats in the ZY group were intragastrically administered with Xiaoyao Powder before exposed to immobilization stress every day for 21 days. The food intake and body weight were observed. The concentration of leptin in serum was detected by ELISA method, the expressions of Ob-R, AgRP and NPY in the ARC were assayed by Western Blot and qRT-PCR methods. Results: On the 7 th, 14 th and 21 st days of stress, the body weight and food intake of rats in 21-MX group were lower than those in ZC group(P<0.01), while the growth of body weight of rats in ZY group was significantly better than those in 21-MX group(especially on the 7 th and 21 st days of stress)(P<0.05), and the food intake in ZY group increased significantly(especially on the 14 th and 21 st days of stress)(P<0.05). The serum leptin and expression levels of Ob-R protein and gene in 7-MX group and 21-MX group were obviously higher than those in ZC group(P<0.05, P<0.01), and the protein and gene expression levels of AgRP and NPY were significantly decreased(P<0.05, P<0.01). ZY group reduced serum leptin and Ob-R expression in ARC compared with 21-MX group(P<0.05, P<0.01), meanwhile, ZY group remarkably up-regulated AgRP and NPY expression in the ARC(P<0.01), but there was no difference between ZY group and 7-MX group. Conclusion: The somatic discomfort symptoms, such as loss of appetite and slowing growth in body weight in 21-MX group, may be related to increase in Ob-R and decrease in AgRP and NPY expression in hypothalamic ARC. Xiaoyao Powder can effectively improve the body discomfort symptoms of rats after chronic immobilization stress by regulating the protein and gene expression levels of Ob-R, AgRP and NPY in ARC.
引文
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[15]旷湘楠,王少贤,方朝义,等.逍遥散调节慢性应激肝郁脾虚模型大鼠下丘脑弓状核ob-R、α-MSH变化机制.世界中医药,2017,12(3):488-493
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[18]李晓娟,白晓晖,李娜,等.逍遥散对慢性束缚应激抑郁模型大鼠JAK/STAT信号通路的影响.中华中医药杂志,2016,31(5):1700-1704
[2]王杰鹏,旷湘楠,张城浩,等.慢性应激肝郁脾虚模型大鼠行为学变化及逍遥散调节作用.河北中医药学报,2016,31(4):7-10,21
[3]Liang Y,Guo X L,Chen J X,et al.Effects of the chinese traditional prescription xiaoyaosan decoction on chronic immobilization stress-induced changes in behavior and ultrastructure in rat hippocampus.Evid Based Complement Alternat Med,2013:984797.doi:10.1155/2013/984797
[4]Luo H G,Chen J X,Zhang Q,et al.Comparative study on effects of Xiaoyao Powder and its modified prescription on plasma metabolomics of rats with chronic immobilization stress.Chinese Journal of Integrative Medicine,2013,19(8):610-615
[5]赵宏波,白晓晖,李晓娟,等.逍遥散治疗慢性束缚应激焦虑模型大鼠行为学评价.中华中医药杂志,2014,29(5):1669-1673
[6]姜幼明,蒙缜之,刘玥芸,等.逍遥散干预慢性应激焦虑状态概述.中医杂志,2015,56(3):259-261
[7]Wang S X,Chen J X,Yue G X,et al.Xiaoyaosan decoction regulates changes in neuropeptide Y and Leptin receptor in the rat arcuate nucleus after chronic immobilization stress.Evid Based Complement Alternat Med,2012,doi:10.1155/2012/381278
[8]Lenard N R,Berthoud H R.Central and peripheral regulation of food intake and physical activity:pathways and genes.Obesity(Silver Spring),2008,16(suppl 3):s11-22
[9]Li S F,Li X.Leptin in normal physiology and Leptin resistance.Sci.Bull,2016,61(19):1480-1488
[10]徐志波,陈芳,江劲.COPD营养不良大鼠下丘脑瘦素受体活化后STAT3、SOCS3的表达研究.浙江医学,2017,39(5):358-361
[11]魏萌,武俭,苏玮,等.侧脑室微量注射瘦素对大鼠下丘脑Gn IH表达的影响.承德医学院学报,2015,32(2):91-93
[12]赵艳军,马立新,吴瑞芹,等.侧脑室注射瘦素对大鼠迷走复合体阿黑皮素原的影响.河北医药,2017,39(22):3388-3391
[13]张善鹏,王军,娄玉杰.瘦素受体研究进展.中国畜牧兽医,2010,37(11):140-142
[14]陈家旭,杨维益.神经一内分泌一免疫网络研究概况及其与中医肝脏关系的探讨.北京中医药大学学报,1995,18(4):7-11
[15]旷湘楠,王少贤,方朝义,等.逍遥散调节慢性应激肝郁脾虚模型大鼠下丘脑弓状核ob-R、α-MSH变化机制.世界中医药,2017,12(3):488-493
[16]Jiang Y M,Li X J,Meng Z Z,et al.Effects of Xiaoyaosan on stressinduced anxiety-like behavior in rats:Involvement of CRF1receptor.Evid Based Complement Alternat Med,2016:1238426.doi:10.1155/2016/1238426
[17]Li X J,Ma Q Y,Jiang Y M,et al.Xiaoyaosan exerts anxiolytic-like effects by down-regulating the TNF-α/JAK2-STAT3 pathway in the rat hippocampus.Sci Rep,2017,7(1):353
[18]李晓娟,白晓晖,李娜,等.逍遥散对慢性束缚应激抑郁模型大鼠JAK/STAT信号通路的影响.中华中医药杂志,2016,31(5):1700-1704