两种气阴两虚证小鼠模型制备方法比较

Two Preparation Methods of Qi and Yin Deficiency Syndrome Mice Models

目的比较两种不同的复合造模方法,寻找建立气阴两虚证模型的优势方法。方法72只BALB/c小鼠随机分为空白组、游泳复合组、睡眠剥夺复合组,每组24只。每组再按造模3周、6周、9周随机分为3小组,每小组8只。其中游泳复合组采用饮食不节^(+)热性药灌胃^(+)负重游泳的方法,睡眠剥夺复合组采用饮食不节^(+)热性药灌胃^(+)睡眠剥夺的方法,分别造模3周、6周、9周。造模第0周,记录每只小鼠抓力与体质量。造模第3、6、9周比较各组小鼠体质量、抓力、力竭游泳时间、饮水量、唾液流率、粪便含水率、面温;进行旷场实验,记录300 s内小鼠活动的总路程、站立次数、角落时间和中心时间;计算胸腺指数、脾脏指数;检测外周血中CD4^(+)、CD8^(+)T淋巴细胞含量和血清中环磷酸腺苷(cAMP)、环磷酸鸟苷(cGMP)含量。结果试验期间,游泳复合组小鼠死亡3只,睡眠剥夺复合组死亡1只。造模第3周,与空白组比较,游泳复合组力竭游泳时间显著减少,睡眠剥夺复合组体重、力竭游泳时间显著减少,总路程、站立次数和中心时间显著增加(P<0.01);与游泳复合组同时间相比,睡眠剥夺复合组总路程、站立次数和中心时间均显著增加(P<0.05或P<0.01)。造模第6周,与空白组相比,游泳复合组角落时间、血清cAMP含量、cAMP/cGMP值显著增加,睡眠剥夺复合组cAMP/cGMP值显著增加,外周血CD4^(+)T淋巴细胞含量、唾液流率、胸腺指数显著降低,游泳复合组和睡眠剥夺复合组抓力、力竭游泳时间、脾脏指数显著减少(P<0.05或P<0.01)。造模第9周,与空白组相比,游泳复合组和睡眠剥夺复合组饮水量、角落时间、血清cAMP含量、cAMP/cGMP值显著增加,唾液流率、抓力、力竭游泳时间、总路程、站立次数、中心时间、血清cGMP含量、胸腺指数、脾脏指数及外周血CD4^(+)T淋巴细胞、CD8^(+)T淋巴细胞、CD4^(+)/CD8^(+)值显著减少,游泳复合组粪便含水率出现显著降低(P<0.05或P<0.01);与游泳复合组相比,睡眠剥夺复合组饮水量、粪便含水率、抓力显著增加(P<0.05)。造模第3、6、9周末各组小鼠面温差异均无统计学意义(P>0.05)。结论游泳复合组与睡眠剥夺复合组造模9周均能出现与临床气阴两虚相似的症状。若进行与便秘相关的实验,使用游泳复合组造模方法建立气阴两虚模型;若实验与便秘无关,则更适合使用睡眠剥夺复合组造模方法建立气阴两虚模型,以减少实验动物死亡率。

Objective To compare two different composite modeling methods and find the superior one to establish qi-yin deficiency syndrome model.Methods A total of 72 BALB/c mice were randomly divided into normal group,swimming composite model group and sleep deprivation composite model group,with 24 mice in each group.Each group was further randomly divided into 3 groups according to 3 weeks,6 weeks and 9 weeks of modeling,with 8 mice in each group.Swimming composite model group was given uncontrolled diet and drugs of heat nature by gavage plus weight-bearing swimming,while the sleep deprivation composite model group was given uncontrolled diet and drugs of heat nature by gavage plus sleep deprivation,which were modeled for 3 weeks,6 weeks and 9 weeks,respectively.At week 0 of the modelling,the grip strength and body weight of each mouse were recorded.At week 3,6,and 9,the weight,grip strength,exhaustion swimming time,water intake quantity,saliva flow rate,fecal moisture content,and surface temperature were recorded and compared between groups;open field test was used to record the total distance traveled within 300s,the standing frequency,and time spent in the center and in the corner;the thymus index and spleen index were calculated;the peripheral blood CD4^(+),CD8^(+)T lymphocytes,as well as the contents of cyclic adenosine phosphate(cAMP)and cyclic guanosine phosphate(cGMP)in serum were detected.Results During the experiment,3 mice in the swimming composite model group,and 1 mouse in the sleep deprivation composite group died.At week 3,compared to those in the normal group,the exhaustive swimming time in the swimming composite model group,as well as the weight and exhaustive swimming time in the sleep deprivation composite model group significantly decreased,while the total distance traveled,the standing frequency and the time spent in the center significantly increased in the sleep deprivation composite model group(P<0.01);the sleep deprivation composite model group had increased total distance traveled,the standing frequency and the time spent in the center than the swimming composite model group(P<0.05 or P<0.01).At week 6,compared to those in the normal group,the time spent in the corner,serum cAMP content,and cAMP/cGMP value in the swimming composite model group,as well as cAMP/cGMP value in the sleep deprivation composite model group significantly decreased,while the peripheral blood CD4^(+)T lymphocytes,salivary flow rate and thymus index in the sleep deprivation composite model group,and the grip strength,exhaustion swimming time,spleen index in both model groups significantly decreased(P<0.05 or P<0.01).At week 9,compared to those in the normal group,the water intake quantity,time spent in the corner,serum cAMP content,cAMP/cGMP value in both model groups increased significantly,while the saliva flow rate,grip strength,exhaustive swimming time,total distance traveled,standing frequency,time spent in the center,serum cGMP,thymus index,spleen index,and peripheral blood CD4^(+)T lymphocytes,CD8^(+)T lymphocytes,and CD4^(+)/CD8^(+)value decreased significantly,as well as the fecal moisture content in the swimming composite model group(P<0.05 or P<0.01);the sleep deprivation composite model group had significantly increased water intake quantity,fecal moisture content,and grip strength than the swimming composite model group(P<0.05).There was no significant difference in the surface temperature of the mice in each group measured at week 3,6 and 9(P>0.05).Conclusion Both the swimming composite model group and the sleep deprivation composite model group could have the similar symptoms with qi-yin deficiency after 9 weeks of modeling.For experiments related to constipation,the modeling method used in the swimming composite group is recommended to establish the qi-yin deficiency model;if there is nothing to do with constipation,the method used in the sleep deprivation composite model group is suitable modeling in order to reduce the mortality rate of the experiment animals.