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目的 探讨转化生长因子 -β1(TGF -β1)调节基质金属蛋白酶- 9(MMP -9)生成的分子机理。方法 选用小鼠肾小球足突细胞系,以细胞因子TGF -β1为刺激物,建立炎症细胞模型,分别应用明胶酶谱法和逆转录 PCR方法观察TGF -β1刺激细胞后培养上清MMP- 9活性及细胞MMP- 9mRNA变化;应用Western蛋白印迹检测TGF β1调节MMP-9中对ERK信号途径和转录因子Ets 1蛋白的影响。结果 对照组细胞上清有微弱MMP- 9活性, TGF β1刺激细胞 24h后培养上清MMP 9活性较对照组明显升高 (P<0 01),并呈TGF -β1刺激剂量依赖趋势; TGF- β1刺激 6h细胞MMP 9mRNA较对照组明显增加 (P<0 01),并且维持高水平至 24h;TGF- β1刺激细胞 4h转录因子Ets- 1蛋白增加(P<0 01),持续高水平至 24h。TGF- β1可以刺激细胞ERK-1 /2活化; ERK-1 /2活化阻断剂PD98059预处理细胞,可以阻断TGF β1刺激引起的Ets 1蛋白、MMP -9活性、MMP -9mRNA增加的效应。结论 以上结果提示TGF β1是通过活化细胞内ERK信号途径和上调转录因子Ets -1蛋白刺激足突细胞MMP- 9mRNA表达,继而蛋白活性增加。
Objective To investigate the molecular mechanism of transforming growth factor-β1 (TGF-β1) regulating the production of matrix metalloproteinase-9 (MMP-9). Methods Mouse glomerular foot process cell line was selected and the inflammatory cell model was established by stimulating with TGF-β1. Gelatin zymography and RT-PCR were used to observe the expression of MMP- 9 activity and MMP-9 mRNA level. Western blot was used to detect the effect of TGFβ1 on the ERK signaling pathway and the transcription factor Ets 1 in MMP-9. Results The supernatant of control group had weak MMP-9 activity. The activity of MMP-9 in supernatant of TGF-β1-stimulated cells was significantly higher than that of control group (P <0.01) (P <0.01), and maintain a high level of 24h; TGF-β1 stimulated cells 4h transcription factor Ets-1 protein increased (P <0.01), sustained high level to 24h . TGF-β1 can stimulate cell ERK-1/2 activation; ERK-1/2 activation blocker PD98059 pretreatment cells, can block TGFβ1 stimulated Ets 1 protein, MMP-9 activity, MMP -9 mRNA increased effect . Conclusion The above results suggest that TGFβ1 stimulates MMP-9 mRNA expression in footprotected cells by activating ERK signaling pathway and up-regulating the transcription factor Ets-1 protein, which in turn leads to increased protein activity.