PDGF对哮喘大鼠气道平滑肌细胞中ERK通路的影响研究

PDGF induces proliferation of airway smooth muscle cells of asthmatic rats through activation of ERK signal pathway

目的研究血小板源性生长因子(PDGF)对哮喘大鼠气道平滑肌细胞(ASMC)中ERK信号通路的调控作用。方法建立哮喘大鼠模型,原代分离培养ASMC,设未干预组(A组)、ERK阻断剂组(B组)、PDGF组(C组)和PDGF+ERK阻断剂组(D组)。A组不加干预;B组又分为B1、B2、B3、B4组,分别加入浓度为0.1、1、5、10μmol/L的ERK阻断剂U0126;C组又分为C1、C2、C3、C4组,分别加入浓度为1、10、25、50μg/L的PDGF同二聚体PDGF-BB;D组加入10μmol/L的U0126和50μg/L的PDGF-BB。以RT-PCR法检测各组ERK1和TGF-β1的mRNA表达,免疫组化法检测A、B4、C4和D组中以上蛋白的表达。结果 RT-PCR提示B1、B2、B3、B4组ERK1 mRNA表达均显著低于A组(均P<0.01),U0126以浓度依赖的方式抑制PDGF诱导的ASMC中ERK的活化,C1、C2、C3、C4组ERK1 mRNA表达均显著高于A组(均P<0.01),ERK1 mRNA的表达与PDGF-BB存在明显的浓度依赖关系,D组与A组比较无统计学差异(P>0.05)。免疫组化提示B4组ASMC中ERK1蛋白表达显著低于A组,C4组显著高于A组(均P<0.01),D组与A组相比无统计学差异(P>0.05);B4组TGF-β1蛋白表达显著低于A组,C4组显著高于A组,同时C4组显著高于B4组(均P<0.01),D组与A组相比无统计学差异(P>0.05)。结论 PDGF可剂量依赖性激活哮喘大鼠ASMC内的ERK通路,同时伴随TGF-β1信号通路的活化。ERK通路参与了ASMC增殖的细胞内信号转导过程,PDGF可能经ERK环节促进TGF-β1通路活化。

Objective To investigate the effect of platelet- derived growth factor （PDGF）on proliferation of airway smooth muscle cells （ASMC） in asthmatic rats and its relation to external signal regulated kinase （ERK） signal pathway. Methods Asth-ma model was established in Sprague- Dawley rats. ASMCs separated from asthmatic rats were divided into four groups：group A without any intervention served as control, group B was treated with ERK blocking agent U0126 0.1μmol/L（B1）, 1μmol/L （B2）, 5μmol/L （B3） or 10μmol/L （B4）, group C was treated with recombinant rat PDGF- BB 1μg/L （C1）, 10μg/L （C2）, 25μg/L （C3） or 50μg/L （C4）, group D was treated with 10μmol/L U0126 and 50μg/L PDGF. The mRNA expressions of ERK1 and TGF- β1 in ASMCs of asthmatic rats were detected by RT- PCR. The protein expressions of ERK1 and TGF- β1 were detected by immuno-histochemistry （group A, B4, C4 and D only）. Results Compared to group A, the expressions of ERK1 mRNA in ASMCs of groups B1, B2, B3 and B4 were significantly decreased（P〈0.01） in a concentration- dependent manner;while the expressions of ERK1 mRNA in groups C1, C2, C3 and C4 were significantly increased （P〈0.01） in a concentration- dependent manner. There was no significant difference in expressions of ERK1 mRNA between group D and group A （P〉0.05）. Immunohistochemistry showed that the expression of P- ERK1 in ASMCs of group B4 was significantly lower and that of group C4 was higher than that in group A （both P〈0.01）;while there was no significant difference between group D and group A （P〉0.05）. Conclusion PDGF can induce the proliferation of airway smooth muscle cells of asthmatic rats, in which the activation of ERK signal pathway is in-volved.