心外膜脂肪组织源瘦素促进高脂喂食诱导的肥胖大鼠心肌成纤维细胞增殖  

Epicardial adipose tissue-derived leptin promotes cardiac fibroblast proliferation in high-fat diet induced obese rats

作　　者：李敏[1] 刘蕾[1] 宋艳[2] 田刚[1] 朱丹军[1] LI Min;LIU Lei;SONG Yan;TIAN Gang;ZHU Dan-jun(Department of Cardiology, The First Affiliated Hospital, Xian Jiaotong University, Xlan Shaanxi 710061 , China)

机构地区：[1]西安交通大学医学院第一附属医院心血管内科,陕西西安710061 [2]西安交通大学医学院第一附属医院心血管内科超声科,陕西西安710061

出　　处：《中华高血压杂志》2018年第3期233-240,共8页Chinese Journal of Hypertension

基　　金：国家自然科学基金(30871042,81600574);陕西省科学技术研究发展计划国际科技合作与交流计划项目(2012kw-40-01);陕西省科学技术研究发展计划自然科学基础研究计划项目(2014JM2-8145);心血管疾病临床技术研究[SF1416(2)]

摘　　要：目的探讨心外膜脂肪组织来源的瘦素对心肌成纤维细胞(CF)增殖的作用及其可能机制。方法 8周龄雄性Wistar大鼠20只随机分为2组,分别给予高脂饲料及标准大鼠饲料喂养,12周后筛选出7只肥胖大鼠作为肥胖组,普食组随机选取7只作为对照组,每周称重。喂养12周后颈动脉插管测得血压;酶联免疫吸附试验(ELISA)法检测大鼠血清瘦素水平;取大鼠心肌组织行HE及Masson染色,比较两组心脏结构改变;Western blot法检测两组大鼠心外膜脂肪组织瘦素和瘦素受体蛋白表达水平。利用大鼠心外膜脂肪组织制备条件培养基,ELISA法检测条件培养基中瘦素水平;分别利用条件培养基、瘦素、瘦素拮抗剂(leptin-tA)、细胞外信号调节激酶(ERK)1/2抑制剂、p38抑制剂等干预大鼠CF,四甲基偶氮唑盐比色法(MTT)检测细胞增殖情况,Western blot法检测细胞瘦素、p-ERK、t-ERK、p-p38、t-p38蛋白表达。结果与对照组相比,肥胖组大鼠血清瘦素水平升高,心肌结构紊乱,胶原沉积增加。大鼠心外膜脂肪组织瘦素及瘦素受体蛋白表达增高(均P<0.05)。体外实验发现,条件培养基(心外膜脂肪组织制成的上清液)瘦素水平明显升高[(8.03±0.75)比(4.17±0.36)μg/L,P<0.05],且高于血清瘦素水平[(8.03±0.75)比(5.39±0.26)μg/L,P<0.05];与对照组大鼠心外膜脂肪组织制备的条件培养基培养的CF相比,利用肥胖组大鼠心外膜脂肪组织制备的条件培养基培养的CF增殖显著增加,ERK1/2、p38磷酸化水平升高(均P<0.05);加入瘦素拮抗剂后,细胞ERK1/2、p38磷酸化水平降低(均P<0.05),加入瘦素拮抗剂、ERK1/2抑制剂、p38抑制剂后细胞增殖降低。结论心外膜脂肪组织来源的瘦素可以促进肥胖大鼠CF增殖,这种作用可能与ERK1/2和p38信号通路激活有关。Objective To investigate the effects of epicardial adipose tissue-derived leptin on cardiac fibroblast （CF） proliferation and its possible mechanism. Methods Twenty eight-week old male Wistar rats were randomly divided to two groups and were fed on high fat diet and standard diet respectively. After 12 weeks, 7 obese rats with high fat diet were selected as obesity group, and 7 non-obese rats with standard diet were selected as control group. Body weight was measured weekly. By the end of the experiment, blood pressure was measured by carotid artery intubation. The expression of serum leptin was detected by enzyme-linked immunosorbent assay （ELISA）. Cardiac pathological score （hematoxylin-eosin and Masson trichrome stain） were examined to compare heart structure changes of two groups. Leptin and leptin receptor level of epicardial adipose tissue were detected by Western blot. Cardiac fihrohlasts were cultured in epicardial adipose tissue-conditioned medium with [eptin,leptin triple antagonist （leptin-tA）, extracellular signal-regulated kinases （ERK） 1/2 inhibitor or p38 inhibitor. Cell proliferation was detected by methylthiazolyl tetrazolium （MTT） assay. Leptin, p-ERK1/2, t-ERK1/2, p-p38 and t-p38 level of CF were detected by Western blot. Results Compared with control group, the levels of serum leptin were significantly higher in the obese group. Leptin and leptin receptor expression in epicardial adipose tissue were significantly higher in obese rats than in control rats. In vitro, compared with control group, leptin expression in epicardial adipose tissue-conditioned medium was significantly increased in obese rats F（ 8.03 ± 0.75 ） vs （ 4.17 ±0. 36 ） μg/L, P〈0.05], and higher than the level of serum leptin [（8.03 ± 0.75） vs {5.39± 0.26） μg/L, P〈0.05]. Proliferation of cardiac fibroblasts in epicardial adipose tissue-conditioned medium was significantly increased in obese group （P〈 0.05）, and p-ERK1/2 and p-p38 expression increased significantly（all P〈0

关 键 词：肥胖 心外膜脂肪组织 瘦素 心肌成纤维细胞 

分 类 号：R54[医药卫生—心血管疾病] R589.2[医药卫生—内科学]