脾虚证模型大鼠腓肠肌损伤及其机制的探查  

Exploration of gastrocnemius injury and its mechanism in rats with syndrome of spleen deficiency

作　　者：范方馨 朱昊如 钱梦 史拓 朱巳旲 谢鸣 FAN Fangxin;ZHU Haoru;QIAN Meng;SHI Tuo;ZHU Sitai;XIE Ming(School of Traditional Chinese Medicine,Beijing University of Chinese Medicine,Beijing 100029,China)

机构地区：[1]北京中医药大学中医学院,北京100029

出　　处：《北京中医药大学学报》2023年第1期77-86,共10页Journal of Beijing University of Traditional Chinese Medicine

基　　金：国家自然科学基金面上项目(No.81173193);北京中医药大学重点攻关项目(No.90020131210002)。

摘　　要：目的观察脾虚证模型大鼠腓肠肌损伤情况并对其机制进行研究。方法将雄性Wistar大鼠(SPF/VAF级)按体质量随机分为对照组和模型组,模型组采用饮食失节联合游泳劳倦法复制脾虚证模型,对照组予以正常饲养。定期观察并记录2组大鼠的外观表征、体质量、饮食量和粪便含水率。实验第2、3、4周末,测定各组大鼠的游泳力竭时间、抓握力、尿D-木糖排泄率、脾脏指数、胸腺指数及腓肠肌湿质量,HE染色观察腓肠肌病理结构,酶联免疫吸附测定法检测血清中胃动素、胃泌素、白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)和腓肠肌组织中柠檬酸合酶、Ca^(2+)-ATP酶、IL-1β、IL-6、TNF-α含量。实验第4周末,用免疫组织化学法检测腓肠肌IL-1β、IL-6、TNF-α阳性表达水平,免疫荧光法检测腓肠肌切割型天冬氨酸蛋白水解酶3(cleaved-caspase 3)阳性细胞。结果与对照组比较,第2~4周模型组大鼠体质量、饮食量、游泳力竭时间、抓握力、腓肠肌湿质量和尿D-木糖排泄率下降(P<0.05,P<0.01),粪便含水率增加(P<0.01);第3~4周模型组大鼠血清胃泌素下降(P<0.05,P<0.01);第2周模型组大鼠腓肠肌湿质量/体质量和血清IL-6增加(P<0.05,P<0.01);第4周模型组大鼠腓肠肌湿质量/体质量、脾脏指数、胸腺指数、腓肠肌柠檬酸合酶和Ca^(2+)-ATP酶水平、血清胃泌素、IL-1β和TNF-α含量下降(P<0.01),腓肠肌cleaved-caspase 3、IL-1β、IL-6、TNF-α含量增加(P<0.01)。结论饮食失节联合游泳劳倦法复制的脾虚证模型大鼠具有中医脾虚证的外观表征和实验室相关指标的变化特点,腓肠肌出现肌力减退和肌萎缩,并随造模时间延长而加重,伴有全身免疫功能减退与血清、腓肠肌炎性因子含量增加,提示脾虚证骨骼肌受损涉及线粒体损伤和细胞凋亡,并与炎性机制有关。Objective To investigate skeletal muscle injury and its mechanism in a rat model of spleen deficiency syndrome.Methods Male Wistar rats(SPF/VAF)were randomly divided into control group and model group according to body weight.The“improper diet combined with exhaustive swimming”method was used to duplicate the rat model with spleen deficiency syndrome.The control group was fed normally.We regularly recorded the appearance,body weight,dietary intake,and excrement water content of the two groups of rats.At the end of Week 2,3,and 4,the swimming exhaustion time,grip strength,urine D-xylose excretion rate,spleen index,thymus index,and gastrocnemius wet weight were measured,and the gastrocnemius was subjected to pathological examination by HE staining.Besides,at the end of Week 2,3,and 4,we used enzyme-linked immunoassay to determine the levels of motilin,gastrin,interleukin-1β(IL-1β),interleukin-6(IL-6),and tumor necrosis factor-α(TNF-α)in serum,and the levels of citrate synthase,Ca^(2+)-ATPase,IL-1β,IL-6,and TNF-αin the gastrocnemius.At the end of Week 4,immunohistochemistry method was used to measure the positive expression of IL-1β,IL-6,and TNF-α,and immunofluorescence was used to detect the cells that were positive for cysteine aspartate proteolytic enzyme 3(cleaved-caspase 3)in the gastrocnemius.Results Compared to the control group,the body weight,dietary intake,swimming exhaustion time,grip strength,gastrocnemius wet weight,and urine D-xylose excretion rate in the model group were significantly decreased from Week 2 to 4(P<0.05,P<0.01),while excrement water content increased(P<0.01).Serum gastrin in the model group was also significantly decreased at Week 3 and 4(P<0.05,P<0.01).Besides,the gastrocnemius wet weight to body weight ratio and serum IL-6 of the rats in the model group were increased at Week 2(P<0.05,P<0.01).At the end of Week 4,the gastrocnemius wet weight to body weight ratio,spleen index,thymus index,citrate synthase and Ca^(2+)-ATPase in the gastrocnemius,the levels of gastrin,IL-1β,

关 键 词：脾虚证 大鼠 腓肠肌损伤 线粒体酶 细胞凋亡 炎性机制 

分 类 号：R285.5[医药卫生—中药学]