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目的探讨慢性哮喘病理生理过程中肺组织组蛋白去乙酰化酶(HDAC)活性变化趋势及其对气道平滑肌细胞表型及生物学性状的影响。方法构建慢性哮喘小鼠模型,检测肺组织HDAC活性。以HDAC抑制剂曲古霉素A(TSA)刺激体外培养的大鼠气管平滑肌及人原代支气管平滑肌细胞,采用蛋白质印迹、MTT、Transwell及三维凝胶收缩等方法检测抑制HDAC活性对气道平滑肌细胞表型转变的影响及其相应的细胞增殖、迁移、收缩能力的变化。结果慢性哮喘组小鼠肺组织HDAC活性为(0.371±0.054)μmol/(L·μg),较正常对照组的(0.603±0.034)μmol/(L·μg)显著降低,差异有统计学意义(P<0.01)。TSA(0.5μmol/L)作用后12～24h,体外培养的大鼠气管平滑肌环及人支气管平滑肌细胞的α-sm-actin、SM22-α表达明显增加,作用后24h及48h支气管平滑肌细胞数目分别为(1.719±0.044)×104及(1.808±0.009)×104个,明显低于空白对照组的(1.911±0.048)×104及(2.537±0.01)×104个(P<0.05)。血小板源性生长因子(PDGF)诱导的支气管平滑肌细胞迁移数也在TSA作用后显著降低(88±7.632vs52±7.5,P<0.05)。此外,TSA作用24h后的三维凝胶收缩率[(9.885±7.084)%]较空白对照组[(44.844±3.808)%]及PDGF组[(41.315±7.943)%]明显降低(P<0.05)。结论慢性哮喘小鼠肺部HDAC的活性下降,可能与气道平滑肌细胞向“收缩型”转变有关,但HDAC活性下降并不增加细胞的收缩能力。
Objective To investigate the change trend of histone deacetylase (HDAC) activity in lung tissue and its effect on the phenotype and biological characteristics of airway smooth muscle cells in the pathophysiology of chronic asthma. Methods The chronic asthmatic mice model was constructed and HDAC activity in lung tissue was detected. The rat tracheal smooth muscle cells and human primary bronchial smooth muscle cells cultured in vitro were stimulated with HDAC inhibitor of trichostatin A (TSA). The inhibitory effect of HDAC activity on airway smooth muscle cells was evaluated by Western blotting, MTT, Transwell and three-dimensional gel contraction. Cell phenotype changes and the corresponding cell proliferation, migration, changes in contractility. Results The HDAC activity in lung tissue of mice with chronic asthma was (0.371 ± 0.054) μmol / (L · μg), which was significantly lower than that of the normal control group (0.603 ± 0.034) μmol / (L · μg) P <0.01). After TSA (0.5μmol / L) for 12-24h, the expression of α-sm-actin and SM22-α in tracheal smooth muscle and bronchial smooth muscle cells were significantly increased in vitro. The number of bronchial smooth muscle cells at 24h and 48h (1.719 ± 0.044) × 104 and (1.808 ± 0.009) × 104, which were significantly lower than those in the blank control group (1.911 ± 0.048) × 104 and (2.537 ± 0.01) × 104 (P <0.05). The migration of bronchial smooth muscle cells induced by platelet derived growth factor (PDGF) was also significantly decreased after TSA (88 ± 7.632 vs. 52 ± 7.5, P <0.05). In addition, the three-dimensional gel shrinkage rate after TSA treatment for 24h was significantly lower than that of the blank control group [(44.844 ± 3.808)%] and (41.315 ± 7.943%) in the PDGF group [(9.885 ± 7.084)%] . Conclusions The decrease of HDAC activity in the lungs of chronic asthmatic mice may be related to the change of airway smooth muscle cells toward “contractile ”, but the decrease of HDAC activity does not increase the contractile activity of the cells.