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[目的]研究miR-29a对PC12细胞向神经细胞分化的影响,为进一步探究miR-29a生物学作用奠定基础。[方法]以PC12高分化(PC12D)细胞和PC12低分化(PC12poor D)细胞为研究对象,建立高表达miR-29a细胞模型(PC12-p LKO-miR-29a),以高表达p LKO.1-puro-vector空载的细胞(PC12-p LKO-vector)作为对照,使用嘌呤霉素筛选稳定表达的细胞株,通过RT-PCR、细胞免疫荧光、Western blot观察:1miR-29a对PTEN干扰效率;2PTEN干扰后PC12 poor D细胞形态学改变;3PC12-p LKO-miR-29a细胞株中神经生长相关蛋白GAP-43变化。[结果]PC12D和PC12poor D细胞内均有PTEN的表达;与PC12poor D细胞相比,PC12D细胞转录的PTEN水平较低,轴突较长、GAP-43表达较高。过表达miR-29a可以降低PTEN表达,增加PC12poor D细胞的GAP-43表达水平,促使PC12poor D细胞突起增加、轴突伸长。[结论]miR-29a能够促进PC12细胞分化,轴突伸长,该过程同miR-29a抑制PTEN表达有关,能够为miR-29a治疗神经损伤提供研究基础。
[Objective] To investigate the effect of miR-29a on the differentiation of PC12 cells into neurons and lay the foundation for further study on the biological role of miR-29a. [Method] With PC12D cells and PC12poor D cells as the research object, a high expression of miR-29a cell model (PC12-p LKO-miR-29a) was established to express pLKO.1 (PC12-p LKO-vector) was used as a control and puromycin was used to screen the stable cell lines. RT-PCR, immunofluorescence and Western blot were used to observe the effect of 1miR-29a on PTEN ; Morphological changes of PC12 poor D cells after 2PTEN interference; and changes of GAP-43 protein in 3PC12-p LKO-miR-29a cell line. [Results] The expression of PTEN was found in both PC12D and PC12poor D cells. Compared with PC12po D cells, PC12D cells had lower PTEN, longer axons and higher GAP-43 expression. Overexpression of miR-29a can reduce the expression of PTEN, increase the expression of GAP-43 in PC12po D cells, and promote the proliferation of PC12po D cells and the elongation of axons. [Conclusion] miR-29a can promote the differentiation and axonal elongation of PC12 cells. This process is related to the inhibition of PTEN expression by miR-29a and provides the basis for the study of miR-29a treatment of nerve injury.