血小板源生长因子BB通过JNK调控大鼠主动脉平滑肌细胞β-catenin核内外分布
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JNK Phosphorylation Is Required for Platelet Derived Growth Factor-BB-Induced β-catenin Nuclear Accumulation in Rat Aortic Smooth Muscle Cell
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    目的探讨阻断JNK后对血小板源生长因子BB引起的细胞增殖的影响以及JNK对血小板源生长因子BB(50μg/L)刺激导致的β-catenin核内外分布的影响。方法应用CCK8法测定不同浓度JNK抑制剂SP600125(10、20和40μg/L)对血小板源生长因子BB诱导的大鼠主动脉平滑肌细胞增殖的影响。应用WesternBlotting法检测血小板源生长因子BB刺激后不同时间点p-JNK、JNK及胞浆和胞核β-catenin的表达变化,以及在应用不同浓度SP600125(10、20和40μg/L)后对p-JNK以及胞核β-catenin表达的影响。免疫荧光法检测血小板源生长因子BB刺激后和给予JNK抑制剂后β-catenin的核内外分布变化。结果血小板源生长因子BB(50μg/L)显著促进大鼠主动脉平滑肌细胞的增殖(0.876±0.041比0.370±0.082,P><0.01),在给予不同浓度(10、20和40μg/L)JNK抑制剂SP600125后平滑肌细胞增殖成浓度依赖性下降(0.635±0.063、0.470±0.044和0.381±0.054比0.876±0.041,P><0.01)。血小板源生长因子BB刺激15 min后p-JNK及60 min后细胞核内β-catenin的表达达峰值,在给予不同浓度SP600125后p-JNK和细胞核内β-catenin的表达成浓度依赖性下降。免疫荧光检测提示血小板源生长因子BB刺激60 min后β-catenin核内聚集明显,而在给予SP600125(40μg/L)后β-catenin核内聚集受到了抑制。结论JNK磷酸化在血小板源生长因子BB刺激引起的β-catenin核内聚集中起关键调节作用。

    Abstract:

    Aim To investigate the effect of JNK in the proliferation and β-catenin nuclear accumulation stimulated by platelet derived growth factor-BB(PDGF-BB). Methods The inhibitory effect of JNK inhibitor-SP600125(10,20,40 μg/L) in the PDGF-BB stimulating vascular smooth muscle cell proliferation was detected by CCK8 assay.Expression of JNK,p-JNK,nuclear and cytosolic β-catenin stimulated by PDGF-BB at different time point was examined.The effect of SP600125 on the nuclear accumulation of β-catenin was detected by Western Blotting and immunofluorescence. Results CCK8 OD value was greatly enhanced after stimulated by PDGF-BB(50 μg/L)(0.876±0.041 vs 0.370±0.082,P><0.01),it was significantly inhibited by JNK inhibitor-SP600125(10,20,40 μg/L)(0.635±0.063,0.470±0.044,0.381±0.054 vs 0.876±0.041,P><0.01) in a concentration-dependent manner.After stimulated by PDGF-BB,expression of p-JNK and nuclear β-catenin increased over time,and p-JNK at 15 minute and nuclear β-catenin at 60 minute reached its peak.The expression of p-JNK at 15 minute and nuclear β-catenin at 60 minute was inhibited by using SP600125(10,20,40 μg/L)in a concentration-dependent manner.The immunofluorescence test also showed that nuclear accumulation of β-catenin in vascular smooth muscle cell was significantly inhibited by SP600125. Conclusion The phosphorylation of JNK was a key step in the nuclear accumulation of β-catenin induced by PDGF-BB.

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刘勇,何延政,李雯,李梓伦,王冕,王深明.血小板源生长因子BB通过JNK调控大鼠主动脉平滑肌细胞β-catenin核内外分布[J].中国动脉硬化杂志,2010,18(5):375~379.

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  • 收稿日期:2010-03-26
  • 最后修改日期:2010-05-10
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