The effects of SOCS3 on neural stem cell fate and feasibility of the SOCS3-overexpressingneural stem cells for stake therapy

为了研究细胞因子信号转导抑制因子3(SOCS3)对神经干细胞命运的影响，用表达SOCS3的腺病毒载体感染干细胞。3d后，Western印迹分析和免疫细胞化学分析显示，SOCS3转染组MAP2蛋白水平和阳性细胞数显著增加，而GFAP蛋白水平和阳性细胞数显著减少。此外，启动子分析显示，信号转导和转录激活因子3(STAT3)的转录水平在转基因细胞中显著降低。此外，在SOCS3过表达后的第1天，Notch家族成员(Notch1)和抑制bHLH因子(ess5和id3)的mRNA水平显著上调。转染后3d，HES5基因的表达水平明显下降，而Noch1基因的表达水平仍然上调。SOCS3阳性细胞均表达Nestin蛋白，但不表达MAP2和GFAP蛋白。这些数据表明，SOCS3的过表达诱导了神经干细胞的神经发生，并抑制了星形胶质细胞的形成。我们的数据还表明，SOCS3促进了神经干细胞的维持。此外，我们还评估了SOCS3高表达神经干细胞用于卒中治疗的可行性。大鼠局灶性脑缺血60min。再灌注后，将高表达SOCS3的神经干细胞(1.0×105/5ul)注入颈总动脉。1天后，用TTC染色评价心肌梗死面积。然而，SOCS3治疗组和赋形剂治疗组之间的脑梗塞体积没有差异。我们现在正试图评估将神经干细胞移植到大脑中的其他方法，尽管这一次我们未能证明SOCS3过表达的神经干细胞用于中风治疗的可行性。

To investigate the effects of suppressors of cytokine signaling 3 (SOCS3) on neural stem cell fate, stem cells were infected with an adenoviral vector expressing SOCS3. Three days later, western blot analysis and immunocytochemical analysis revealed that the protein level of MAP2 and the number of MAP2-positive cells were significantly increased in SOCS3-transfected cells, while the protein level of GFAP and the number of GFAP-positive cells were significantly decreased. Furthermore, promoter assay revealed a significant reduction in the transcriptional level of signal transducer and activator of transcription 3 (Stat3) in the transfected cells. In addition, the mRNA levels of Notch family member (notchl) and inhibitory bHLH factors (hes5 and id3) were significantly up-regulated at one day after overexpression of SOCS3. At three days after transfection, the mRNA level of hes5 was significantly decreased, while that of notchl was still up-regulated. Moreover, all of SOCS3-positive cells expressed Nestin protein but did not express both MAP2 and GFAP proteins. These data indicate that overexpression of SOCS3 induced neurogenesis and inhibited astrogliogenesis in neural stem cells. Our data also show that SOCS3 promoted maintenance of neural stem cells. Furthermore, we evaluated the feasibility of the SOCS3-overexpressing neural stem cells for stroke therapy. Rats were subjected to focal cerebral ischemia for 60 min. Following reperfusion, neural stem cells over-expressing SOCS3 (1.0 x 10^5/5ul) were injected into common carotid artery. One day later, we evaluated the infarct size by TTC staining. However, there was no difference in infarct volume between the SOCS3-treated group and vehicle-treated group. We are now trying to evaluate other methods to transplant NSC into the brain, although we failed to demonstrate the feasibility SOCS3-overexpressing neural stem cells for stroke therapy this time.