AMP依赖的蛋白激酶二甲双胍对老年大鼠肺气肿作用的实验研究

Experimental study of the effects of AMP-dependent protein kinase metformin on emphysema in aged rats

摘要:

目的 探讨AMP依赖的蛋白激酶(AMPK)二甲双胍在老年大鼠肺气肿中的应用效果。方法 取老年24月龄SD大鼠40只,体质量(233.00±12.00)g。按随机数表法将40只大鼠分为空白对照组、模型组、地塞米松组和联合干预组,每组10只。除对照组外,余3组大鼠采用烟熏联合气管内滴注猪胰弹性蛋白酶法制作大鼠肺气肿动物模型。模型构建完成后,对照组与模型组大鼠常规腹腔灌注5ml生理盐水,1次/d；地塞米松组大鼠腹腔灌注等体积地塞米松2mg/kg,1次/d；联合干预组大鼠在地塞米松组基础上联合AMPK二甲双胍250mg/kg腹腔灌注,联合药物体积5ml。4组大鼠均连续干预14d。HE染色观察4组大鼠肺组织损伤情况,记录并比较4组大鼠肺组织炎症因子水平[肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)和IL-8]及生化指标(白细胞计数、中性粒细胞及淋巴细胞比例)。采用SPSS 18.0软件对数据进行分析。根据数据类型,组间比较采用方差分析或两两比较。结果 对照组HE染色下未见异常；模型组HE染色下呼吸性支气管、肺泡大小不一,肺泡数量明显减少,肺泡间隔变薄,部分间隔断裂导致肺泡融合等肺气肿样改变；地塞米松组HE染色下肺泡数目略减少,肺泡间隔一般,可见少许间隔断裂,伴有肺气肿样改变；联合干预组HE染色下肺气肿样改变不明显,肺泡数量未见明显减少。对照组大鼠TNF-a、IL-6及IL-8、白细胞计数、中性粒细胞及淋巴细胞比例依次为(178.53±19.55)pg/ml、(96.58±0.06)pg/ml、(74.55±5.68)pg/ml、(3.49±0.68)×10⁸/L、(17.35±3.24)%和(12.29±2.45)%,模型组大鼠上述指标依次为(261.39±23.21)pg/ml、(753.23±43.24)pg/ml、(323.59±17.85)pg/ml、(13.29±1.46)×10⁸/L、(34.69±4.64)%和(43.53±5.77)%,地塞米松组大鼠依次为(243.66±18.68)pg/ml、(323.32±25.69)pg/ml、(132.31±10.51)pg/ml、(9.48±1.35)×10⁸/L、(25.69±5.32)%和(32.51±4.34)%,联合干预组大鼠依次为(213.69±15.32)pg/ml、(102.49±7.46)pg/ml、(89.43±6.59)pg/ml、(6.31±1.12)×10⁸/L、(21.59±4.31)%和(21.29±3.45)%。与对照组比较,模型组和地塞米松组TNF-a、IL-6及IL-8、白细胞计数、中性粒细胞及淋巴细胞比例显著升高；与模型组比较,地塞米松组和联合干预组上述指标显著降低；与地塞米松组比较,联合干预组上述指标显著降低,差异均有统计学意义(P<0.05)。结论 AMPK二甲双胍用于老年肺气肿大鼠效果理想,可降低炎症因子水平,改善肺组织的损伤。

Abstract:

Objective To investigate the effects of AMP-dependent protein kinase (AMPK) metformin on emphysema in the aged rats. Methods Forty 24-month-old SD rats with a body mass of (233.00±12.00)g were selected and were randomized into control group, model group, dexamethasone group and joint intervention group, with 10 rats in each group. Except for the control group, the other three groups were treated with smoking combined with intratracheal drip of porcine tryptoelastase to establish animal models of emphysema. Intraperitoneally, the control group and the model group were then routinely perfused with 5ml saline once a day, the dexamethasone group with an equal volume of 2mg/kg dexamethasone once a day, and the joint intervention group with an equal volume of 250mg/kg AMPK metformin plus 2mg/kg dexamethasone. All four groups were continuously intervened for 14 days. Lung damage in the four groups was observed using HE staining. They were compared in the respects of the levels of inflammatory factors[tumor necrosis factor-a (TNF-α), interleukin-6 (IL-6) and IL-8] and biochemical indices (white blood cell count, neutrophil and lymphocyte ratio). SPSS statistics 18.0 was employed for data analysis. Depending on data type, intergroup comparison was performed using analysis of variance or pairwise comparison. Results No abnormality was observed in HE staining in the control group. In the model group, the respiratory bronchi and alveoli were of different sizes, the number of alveoli decreased significantly, alveolar septum became thinner, and some alveolar septa ruptured leading to emphysema-like changes such as alveolar fusion. In the dexamethasone group, the number of alveoli decreased slightly, a few ruptures of alveolar septa were observed with emphysema-like changes. The joint intervention group showed no significant emphysema-like changes and no significant reduction in alveolar number. TNF-a, IL-6 and IL-8, white blood cell count, neutrophil and lymphocyte ratio were (178.53±19.55)pg/ml, (96.58±0.06)pg/ml,(74.55±5.68)pg/ml, (3.49±0.68)×10⁸/L, (17.35±3.24)% and (12.29±2.45)% respectively in control group; (261.39±23.21)pg/ml, (753.23±43.24)pg/ml, (323.59±17.85)pg/ml, (13.29±1.46)×10⁸/L, (34.69±4.64)% and (43.53±5.77)% in model group; (243.66±18.68)pg/ml, (323.32±25.69)pg/ml, (132.31±10.51)pg/ml, (9.48±1.35)×10⁸/L, (25.69±5.32)% and (32.51±4.34)% in the dexamethasone group; (213.69±15.32)pg/ml, (102.49±7.46)pg/ml, (89.43±6.59)pg/ml, (6.31±1.12)×10⁸/L, (21.59±4.31)% and (21.29±3.45)% in joint intervention group. Compared with the control group, TNF-a, IL-6, IL-8, white blood cell count, neutrophil and lymphocyte ratio increased significantly in the model group and dexamethasone group; compared with the model group, the above parameters decreased significantly in the dexamethasone group and the joint intervention group; compared with the dexamethasone group, the above indices decreased significantly in the joint intervention group, all the differences being statistically significant (P<0.05). Conclusion AMPK metformin is effective for emphysema in the senile rats, reducing the level of inflammatory factors and improving lung damage.