重组人CC16蛋白改善慢性阻塞性肺疾病模型小鼠肺功能及其机制

Recombinant human CC16 protein improves lung function in chronic ob⁃structive pulmonary disease mice and its mechanism

目的:探究重组人CC16蛋白(rhCC16)对慢性阻塞性肺疾病(COPD)模型小鼠肺功能的改善作用及其机制。方法:40只BALB/c雄性小鼠随机均分为对照组、COPD模型组、COPD+PBS组和COPD+rhCC16组,每组10只。采用被动吸烟6月法制备COPD模型小鼠,从熏烟16周开始,在熏烟前2 h用2.5μg/g rhCC16或等体积PBS对熏烟小鼠进行滴鼻干预,制备COPD+rhCC16组或COPD+PBS组。造模及干预结束后检测各组小鼠肺功能;HE染色观察各组小鼠肺组织形态结构;RT-qPCR法检测各组小鼠肺组织P16和P21的mRNA表达;免疫组织化学法检测各组小鼠肺组织P16、P21、衰老相关β-半乳糖苷酶(SA-β-Gal)、p-P38及磷酸化细胞外信号调节激酶1/2(p-ERK1/2)蛋白表达情况。结果:与对照组相比,COPD模型组小鼠0.3秒内用力呼气容积(FEV0.3)/用力肺活量(FVC)比值降低(P<0.05),气道阻力增加(P<0.05),肺顺应性降低(P<0.05);小鼠肺泡壁破裂塌陷,肺泡大小不均匀,多呈不规则扩大,管腔狭窄,管壁增厚;小鼠肺组织P16和P21的mRNA表达水平显著升高(P<0.05),小鼠肺组织P16、P21、SA-β-Gal、p-P38和p-ERK1/2蛋白表达量升高(P<0.05);与COPD模型组相比,rhCC16干预组小鼠各项肺功能指标均有缓解(P<0.05),肺组织病理损伤减轻,小鼠肺组织P16和P21 mRNA表达水平降低(P<0.05),小鼠肺组织P16、P21、SA-β-Gal、p-P38和p-ERK1/2蛋白表达量降低(P<0.05);PBS组各项指标与COPD组相似。结论:rhCC16可能通过P38 MAPK/ERK1/2信号通路减轻COPD小鼠肺组织衰老,进而改善COPD模型小鼠肺功能。

AIM:To investigate the effect of recombinant human CC16 protein(rhCC16)on lung function of chronic obstructive pulmonary disease(COPD)mice,and to reveal its underlying mechanism.METHODS:Forty male BALB/c mice were randomly divided into control group,COPD group,COPD+rhCC16 group and COPD+PBS group,with 10 mice in each group.The COPD mice were induced using passive smoking for 6 months.From the beginning of 16 weeks of smoking,the mice were intranasally administered with rhCC16(2.5µg/g)or equal volume(20µL)of PBS at 2 h before smoking in COPD+rhCC16 group or COPD+PBS group.The lung function indexes were examined and the morpho‐logical changes of lung tissues were observed by HE staining.The mRNA levels of P16 and P21 in lung tissues were tested by RT-qPCR,and the protein levels of P16,P21,senescence-associatedβ-galactosidase(SA-β-Gal),p-P38 and phos‐phorylated extracellular signal-regulated kinase 1/2(p-ERK1/2)were detected by immunohistochemical staining.RE-SULTS:Compared with control group,the forced expiratory volume in 0.3 s(FEV0.3)/forced vital capacity(FVC)ratio and the lung compliance were declined in COPD group(P<0.05),the airway resistance was enhanced(P<0.05).Com‐pared with COPD group,the FEV0.3/FVC ratio and lung compliance were elevated in COPD+rhCC16 group(P<0.05),and the airway resistance was reduced(P<0.05).Compared with the control mice,the alveolar walls were ruptured and collapsed in COPD mice,and there were uneven alveolar size,irregular enlargement,luminal narrowing and thickened walls.Nevertheless,rhCC16 treatment mitigated the pathological damages of lung tissues compared with COPD mice.Compared with control group,the mRNA levels of P16 and P21 in lung tissues were significantly increased in COPD group(P<0.05),and the protein levels of P16,P21,SA-β-Gal,p-P38 and p-ERK1/2 in lung tissues of COPD mice were also significantly augmented(P<0.05).Compared with COPD group,the mRNA levels of P16 and P21 in lung tissues were significantly decreased in COPD group(P<0.05),and the protein levels of P16,P21,SA-β-Gal,p-P38 and p-ERK1/2 in lung tissues were also significantly inhibited(P<0.05).The lung function indexes,pathological changes,mRNA or protein of P16,P21,SA-β-Gal,p-P38 and p-ERK1/2 showed no significant difference between PBS group and COPD group.CONCLUSION:The rhCC16 can mitigate the senescence of lung tissues through P38 MAPK/ERK1/2 signaling pathway,thus improving the lung function of COPD mice.