摘要
目的探讨3-磷酸肌醇依赖性蛋白激酶-1(3-phosphoinositede dependent protein kinase-1,PDK1)抑制剂对烟熏诱导的慢性阻塞性肺疾病(简称慢阻肺)模型小鼠体内前列腺素E2(prostaglandin E2,PGE2)的干预作用。方法将50只C57BL/6雄性小鼠随机分为正常对照组、熏烟组、熏烟+低剂量PDK1抑制剂组、熏烟+中剂量PDK1抑制剂组、熏烟+高剂量PDK1抑制剂组,每组10只。PDK1抑制剂选择OSU-03012。正常对照组小鼠每日雾化吸入磷酸盐缓冲液2次,持续12周;熏烟组小鼠熏烟每日2次,持续12周;其余三组小鼠每日熏烟方法与时间同熏烟组,但在熏烟前分别腹腔注射低剂量PDK1抑制剂(0.25 mg/kg)、中剂量PDK1抑制剂(0.5 mg/kg)和高剂量PDK1抑制剂(1.0 mg/kg)。熏烟结束后,测试肺功能,小鼠支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF)行细胞计数,用酶联免疫吸附试验测定小鼠血清与BALF中的PGE2,小鼠肺组织进行苏木素–伊红(hematoxylin-eosin,HE)染色,显微镜下观察病理改变。结果各熏烟组小鼠肺功能指标FEV100/FVC和FEV200/FVC与正常对照组相比均有明显下降(P<0.05);熏烟组小鼠BALF中细胞数增加,与正常对照组相比,差异有统计学意义(P<0.05)。熏烟+低剂量PDK1抑制剂组的BALF细胞总数、中性粒细胞和巨噬细胞比例与熏烟组相比,差异无统计学意义(P>0.05)。而熏烟+中剂量PDK1抑制剂组和熏烟+高剂量PDK1抑制剂组BALF细胞总数和中性粒细胞比例逐渐降低,而巨噬细胞比例逐渐升高,与熏烟组相比,差异有统计学意义(P<0.05);与正常对照组相比,熏烟组和熏烟+PDK1抑制剂组中小鼠的血清及BALF的PGE2浓度均升高,差异有统计学意义(P<0.05)。与熏烟组相比,熏烟+中剂量与高剂量的PDK1抑制剂组小鼠的血清及BALF的PGE2浓度均有所下降,差异有统计学意义(P<0.05)。肺组织HE染色可见,熏烟组呈现炎症细胞浸润、肺泡腔扩张、肺大泡形成等病理改变,均符合慢阻肺的病理改变,而熏烟+中剂量PDK1抑制剂组与熏烟+高剂量PDK1抑制剂组均可见炎性细胞浸润减少,肺泡壁变薄和肺泡扩张等情况好转,其中熏烟+高剂量PDK1抑制剂组改善情况更加明显。结论烟熏慢阻肺小鼠模型肺功能下降,气道炎症明显,同时PGE2分泌也增多,而使用PDK1抑制剂能够降低PGE2的分泌,同时减轻气道炎症及病理改变,改善肺功能,且呈现剂量相关性。
Objective To investigate the intervention effect of 3-phosphoinositede dependent protein kinase-1(PDK1)inhibitor on prostaglandin E2(PGE2)in smoking-induced chronic obstructive pulmonary disease(COPD)mice.Methods Fifty C57BL/6 male mice were randomly divided into normal control group,smoking group,smoking+low dose PDK1 inhibitor group,smoking+medium dose PDK1 inhibitor group and high dose PDK1 inhibitor group with 10 mice in each group.The mice in the normal control group inhaled phosphate-buffered saline twice a day for 12 weeks,and the mice in the smoking group were fumigated twice a day,5 days per week for 12 weeks,and the other three groups were given intraperitoneal injection of low-dose PDK1 inhibitor OSU-03012(0.25 mg/kg),medium-dose PDK1 inhibitor(0.5 mg/kg)and high-dose PDK1 inhibitor(1.0 mg/kg)respectively before smoking.After smoking,lung function was tested,the bronchoalveolar lavage fluid(BALF)of each mouse was taken for cell count,the PGE2 in serum and BALF of mice was determined by enzyme linked immunosorbent assay,and the lung tissue of mice was sectioned with paraffin and stained by hematoxylin-eosin(HE),and pathological changes were observed under microscope.Results Compared with the control group,FEV100/FVC and FEV200/FVC of the mice in each smoking group were significantly decreased(P<0.05);The number of cells in BALF of smoking group was significantly higher than that of normal control group(P<0.05).There was no significant difference in the total number of BALF cells,the proportion of neutrophils and macrophages between the smoking+low-dose PDK1 inhibitor group and the smoking group.However,the total number of BALF cells and the proportion of neutrophils in the smoking+medium dose PDK1 inhibitor group and the high dose PDK1 inhibitor group gradually decreased,while the proportion of macrophages gradually increased,compared with the normal control group,the PGE2 concentrations of serum and BALF in the smoking group and the smoking+PDK1 inhibitor group were significantly higher than those in the control group.Compared with the smoking group,the PGE2 concentrations of serum and BALF in the middle and high dose PDK1 inhibitor groups were significantly lower than those in the smoking group.HE staining of lung tissue showed that there were a large number of inflammatory cell infiltration,alveolar cavity dilatation,alveolar wall rupture and fusion,alveolar formation,significant decrease in the number of alveoli and other pathological changes in the smoking group,which were consistent with the pathological changes of COPD.The inflammatory cell infiltration,mucus obstruction and alveolar dilatation were slightly alleviated in the smoking+low-dose PDK1 inhibitor group,while the inflammatory cell infiltration,alveolar wall thinning and alveolar dilatation were improved in both the medium-dose inhibitor group and the high-dose inhibitor group,and the improvement was more obvious in the high-dose inhibitor group.Conclusion The lung function of the smoked COPD mouse decreases,the airway inflammation is obvious,and the secretion of PGE2 is also increased,while the use of PDK1 inhibitor could reduce the secretion of PGE2,reduce airway inflammation and pathological changes,and improve lung function in a dosedependent manner.
作者
吴永红
王可
刘春涛
WU Yonghong;WANG Ke;LIU Chuntao(Department of Respiratory and Critical Care Medicine,West China Hospital,Sichuan University,Chengdu,Sichuan 610041,P.R.China;Department of Critical Care Medicine,The People's Hospital of Jianyang City,Chengdu,Sichuan 641400,P.R.China)
出处
《中国呼吸与危重监护杂志》
CAS
CSCD
北大核心
2023年第5期349-355,共7页
Chinese Journal of Respiratory and Critical Care Medicine
基金
国家自然科学基金(81870034、82070019)。
