TAL对慢支模型大鼠肺泡巨噬细胞凋亡的影响及部分机制研究

Effect of TAL and MAPK signal transduction on alveolar macrophage apoptosis and activity of chronic bronchitis rats

目的探讨枇杷叶三萜酸(TAL)对慢支模型(CB)大鼠肺泡巨噬细胞(AM)活性及凋亡的影响及其可能的作用机制。方法应用BCG联用LPS的方法复制大鼠CB模型,采用体内外给药的方法,观察TAL及MAPK3条通路对CB大鼠AM凋亡及活性的影响。MTT法检测细胞活性,电镜观察AM凋亡,流式细胞技术检测AM凋亡率及AM中凋亡相关基因Bcl-2、Bax的表达。Western blot法检测AM中ERK蛋白磷酸化水平。结果①与正常对照组比较,模型大鼠支气管肺泡灌洗液中AM数增加(P<0.01),而TAL灌胃给药可抑制模型大鼠异常增多的AM数量(P<0.01)。②慢支模型组AM活性较正常组相比明显增强(P<0.01),TAL可抑制CB模型组大鼠AM的活性(P<0.05)。③与正常组比较,模型大鼠AM凋亡率明显降低(P<0.01);TAL给药组及ERK通路抑制剂PD98059组AM凋亡率较模型组比较升高(P<0.05,P<0.01)。而JNK通路抑制剂Curcumin组AM凋亡率降低(P<0.05)。④模型大鼠AMERK磷酸化水平明显升高(P<0.01);TAL(5mg.L-1)体外给药可抑制慢支大鼠AM内ERKMAPK通路的磷酸化(P<0.05)。⑤模型大鼠AMBax表达明显降低而Bcl-2表达升高(P<0.01),TAL给药组可降低慢支大鼠AMBcl-2表达,升高Bax表达。结论慢支模型大鼠AM活性增强、凋亡减少,TAL可诱导AM凋亡,抑制AM活性,使AM内Bcl-2/Bax的表达趋于平衡。ERK、JNKMAPK信号传导通路参与慢支大鼠AM凋亡的调节,JNKMAPK信号通路促进AM凋亡而ERKMAPK信号通路抑制AM凋亡。TAL的促AM凋亡作用可能与其抑制慢支大鼠AMERKMAPK信号通路的磷酸化活化、调节Bcl-2/Bax的表达有关。

Aim To investigate the effect of TAL and MAPK signal transduction on alveolar macrophage （AM） apoptosis and activity of chronic bronchitis （CB） rats.Methods CB model was established by BCG＋LPS method and the in vitro and in vivo experiments were used.MTT method was used to detect the AM activity,and the apoptosis of AM was observed by electron microscope.Results The number of AM in BALF of CB rats was increased than that of normal group （P〈0.01）.The activity of AM was increased in model group than in control group.The apoptotic rate of AM in CB group was much lower than that in the control group [（13.93±3.34）% vs （5.37±1.38）%] （P〈0.01）.ERK inhibitor PD98059 induced the apoptosis of cultured AM while JNK inhibitor Curcumin reduced the apoptosis.TAL could inhibit ERK MAPK phosphorylation in AM of CB rats.Further investigation showed that Bcl-2 protein expression was significantly increased while Bax evidently decreased in AM of CB rats.TAL could significantly decrease Bcl-2 expression and increase Bax protein expression,which might be the mechanism of its effect.Conclusions There is an increased activity and decreased apoptosis of AM in CB rats compared with normal rats.TAL can inhibit AM activity and increase apoptosis of AM in CB rats which may be related to the therapeutic effect of CB.ERK and JNK MAPK signal transduction participates in the apoptosis of AM.Regulation of Bcl-2/Bax imbalance and MAPK phosphorylation in AM of CB rats might be the mechanism of its effect.