转化生长因子TGF-β诱导成骨细胞自噬活性的实验研究
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海南省自然科学基金(814378,20168313);海南省重点科技计划项目(ZDXM20120054)


Transforming growth factor-β regulates autophagic activity on osteoblasts
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    摘要:

    目的:探讨转化生长因子-β (transforming growth factor-β,TGF-β)对成骨细胞自噬活性的调控作用。方法:应用TGF-β 处理成骨细胞系hFOB1.19细胞,实时荧光定量PCR(real-time PCR,RT-PCR)及蛋白质免疫印迹(Western blot)分别检测hFOB1.19细胞自噬相关基因Beclin 1及微管相关蛋白轻链3(microtubule-associated protein light chain 3,LC3)在mRNA及蛋白水平的表达变化;透射电镜(transmission electron microscope,TEM)观察TGF-β 处理后成骨细胞自噬小体的形成;SPSS19.0统计软件进行数据分析。结果:与对照组相比,TGF-β处理hFOB1.19细胞3 h后,Beclin 1 mRNA的表达开始下调(P < 0.05),同时LC3 mRNA开始升高,在6 h达到最高(P < 0.05),随后依作用时间的延长不断下降;在TGF-β 作用后3 h,Beclin 1蛋白的表达水平升高,随后6-12-24 h表达量下降(P < 0.05),而LC3-Ⅱ的蛋白表达自TGF-β作用后6 h开始升高,12 h蛋白表达达到最高(P < 0.01);TGF-β 作用后12 h,TEM可观察到细胞中自噬小体形成。结论:TGF-β 能够上调成骨细胞hFOB1.19的自噬活性。

    Abstract:

    Objective:To investigate the effect of ransforming growth factor-β (TGF-β) on regulating autophagic activity in osteoblasts cell line. Methods:Osteoblasts cell line (hFOB1.19) was treated with TGF-β to induce autophagy. Then,RT-PCR and Western blotting assay were performed to examine the expression of mRNA and protein of Beclin1 and microtubule-associated protein light chain 3(LC3),respectively. Transmission electron microscope(TEM) was performed to identify autophagosomes. Data was analyzed by SPSS 19.0 statistical software. Results:Compared with the control group,after 3 h of TGF-β treatment in hFOB1.19 cells,the mRNA expression of Beclin 1 was initially decreased (P < 0.05). Meanwhile,the mRNA expression of LC3 was increased and reached the highest level at 6 h (P < 0.05),and then decreased with time. At the same time,Beclin 1 was initially increased at protein level after 3 h,and then decreased at 6 h,12 h,and 24 h(P < 0.05),and LC3-Ⅱ was increased after 6 h,reached its peak at 12 h(P < 0.01). Autophagosomes were observed in hFOB1.19 cell line by transmission electron microscope TEM at 12 h. Conclusion:TGF-β can induce autophagic activity in osteoblasts cell line hFOB1.19.

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王雅雯,王彬娉,程亚楠,毛秋华,刘晓晶,刘双喜,徐 普.转化生长因子TGF-β诱导成骨细胞自噬活性的实验研究[J].南京医科大学学报(自然科学版),2016,(9):1041-1045

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  • 收稿日期:2016-05-07
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  • 在线发布日期: 2016-09-18
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