PTEN调节神经样细胞微管相关蛋白tau的聚集及意义

Aggregation of microtubule-associated protein tau modulated by PTEN during the differentiation of bone marrow stem cell-derived nerve cells

目的观察体外诱导成人骨髓基质干细胞(BMSC)向神经细胞分化过程中,在第10号染色体缺失的磷酸酶及张力蛋白同源物基因(PTEN)调节细胞骨架中轴突标志分子微管相关蛋白tau(MAPT)分布及聚集的特征,并探讨其意义。方法从成人骨髓中分离基质干细胞,在体外经细胞因子诱导分化2周后成为神经样细胞,将未分化的BMSC作为对照组;应用半定量逆转录-聚合酶链反应(RT-PCR)和Western blot方法研究MAPT的表达变化;经PTEN抑制剂BPV作用于该细胞后,用结合有荧光剂的鬼笔环肽直接染色细胞微丝肌动蛋白,并联合用免疫荧光细胞化学方法染色MAPT,在激光共聚焦显微镜下观察MAPT和肌动蛋白的定位及关联性。结果成人骨髓来源的基质干细胞经诱导分化后,MAPT的m RNA在分化1周和2周时相对表达水平为0.24±0.04和0.52±0.04,高于对照组BMSC的0.04±0.02(P<0.05)。MAPT蛋白表达水平分别为0.18±0.03和0.44±0.05,高于对照组0.06±0.04(P<0.05)。细胞染色后,经BPV作用后MAPT由弥散状态变为聚集状态,并分布于细胞一侧,神经样细胞开始出现极性。结论 BMSC来源的神经细胞分化成熟时,PTEN参与调节细胞骨架中轴突标志分子MAPT的转运和聚集,为神经细胞的轴突进一步生长提供物质支持。

Objective To observe the diffusion and aggregation of the microtubule associated protein tau（MAPT）modu？lated by phosphatase and tensin homolog deleted on chromosome ten（PTEN）during the nerve cell differentiation by human bone marrow stem cells（BMSC）in vitro, and to analyse the signification. Methods Adult bone marrow stem cells were iso？lated and induced into nerve-like cells by some cytokines in vitro. The mRNA expression of MAPT was detected by semi-quantitative RT-PCR and Western blot assay. The patterns of diffusion and aggregation of the MAPT association of the actin were indicated by Phalloidin-fluoresceineisothioeyanate （FITC） and immunofluorescence （IF） cyto-chemistry, and observed by the laser-confocal microscopy. Results The MAPT mRNA levels were 0.24 ± 0.04 and 0.52 ± 0.04 at 1 week and 2 weeks after the induction,which were significantly higher compared with those of BMSC （0.04 ± 0.02） after the induction （P〈0.05）. The MAPT protein levels were 0.18 ± 0.03 and 0.44 ± 0.05 at 1 week and 2 weeks after the induction, which were significantly higher compared those of BMSC （0.06 ± 0.04, P〈0.05）. The distribution patterns of MAPT were changed from the diffusion to the aggregation in cells after treatment by BPV. The nerve-like cells appeared the characteristic of polarization. Conclusion When the nerve cells derived from bone marrow stem cells obtain the mature differentiation, PTEN may possess the ability of modulating the diffusion and aggregation of MAPT in vitro, also may provide a kind of material ba？sis for the growth of the nerve axon.