肺纤方提取物对肺间质纤维化大鼠肺微血管VEGF、VEGFR2、PAI-1、VCAM-1 mRNA表达的影响

Effect of Feixian Fang extract on mRNA expression of VEGF,VEGFR2,PAI-1 and VCAM-1 in microvascular endothelial cells of rats with pulmonary fibrosis: an in vitro study

目的通过观察肺纤方提取物干预体外培养肺微血管的血管内皮细胞生长因子(VEGF)、血管内皮细胞生长因子受体2(VEGFR 2)、纤溶酶原激活物抑制物1(PAI-1)、血管细胞黏附因子1(VCAM-1)基因表达的影响,探讨肺纤方干预肺纤维化微血管生成的机制。方法从肺纤维化模型大鼠分离培养肺微血管内皮细胞,将其等量分为模型组,氯沙坦组,泼尼松组,肺纤方大、中、小剂量组。模型组加入含20%胎牛血清的DMEM培养基,氯沙坦组加入含20%胎牛血清及10 mg/L氯沙坦的DMEM培养基,泼尼松组加入含20%胎牛血清及5 mg/L泼尼松的DMEM培养基,肺纤方大、中、小剂量组分别加入含20%胎牛血清及100、60、20 mg/L肺纤方提取物的DMEM培养基,分别作用24 h。双链嵌合荧光染料SYBR GreenⅠ实时荧光定量PCR法检测VEGF、VEGFR 2、PAI-1与VCAM-1的mRNA转录水平。结果与模型组比较,肺纤方大、中、小剂量组VEGF基因表达水平均显著降低,而VEGFR2无显著差异;肺纤方大剂量组PAI-1 mRNA表达显著降低;肺纤方大、中剂量组VCAM-1 mRNA表达显著降低(P<0.05)。结论肺纤方可通过抑制肺微血管VEGF、PAI-1及VCAM-1的表达改善血凝及纤溶,抑制血管形成而发挥抗肺纤维化作用。

Objective To investigate the effect of Feixian Fang extract （FXF） on the expression of vascular endothelial cell growth factor（VEGF） , vascular endothelial cell growth factor receptor 2 （ VEGFR 2） , vascular cell adhesion molecule-1 （PAI-1） andplasminogen activator inhibitor-1 （VCAM-1） in pul- monary microvascular endothelial ceils in vitro, so as to explore the mechanism of intervention of FXF on microvascular angiogenesis of pulmonary fibrosis. Methods The lung microvascular endothelial cells which were isolated from pulmonary fibrosis model rats were equally divided into model group, losartan group, prednisone group, FXF high-, mid-, and low- dose group. The cells of model group were cultured in DMEM containing 20% fetal bovine serum, those of losartan group were cultured in DMEM containing 20% fetal bovine serum with 10 g/L losartan, those of prednisone group were cultured in DMEM contai- ning 20% fetal bovine serum with 5 mg/L prednisone, and those of FXF groups were cultured in DMEM containing 20% fetal bovine serum with 100, 60, or 20 mg/L Feixian Fang extract. All the cells were cultured for 24 h. Then the mRNA expression of VEGF, VEGFR2, PAI-1 and VCAM-1 were detected by using method of double chain chimeric fluorescent dye SYBR Green real-time fluorescence quantitative detection. Results Campared with the model group, VEGF mRNA expression decreased significantly in all FXF group, while there was no difference of VEGFR2 ; PAI-1 mRNA was downregulated in FXF high- close group; VCAM-1 mRNA expression decreased significantly in both FXF high-dose group and FXF low-dose group （ P 〈 O. 05 ）. Conclusion The effect of FXF against pulmonary fibrosis via the way of improving the blood coagulation and fibrinolysis leading to inhabitation of angiogenesis, due to downregulat- ing the mRNA expression of VEGF, PAI-1 and VCAM-1 in the lung mierovascular endothelial cells.