Morizane Asuka, Takahashi Jun, Takagi Yasushi, Sasai Yoshiki, Hashimoto Nobuo
Department of Neurosurgery, Clinical Neuroscience, Kyoto University Graduate School of Medicine, Kyoto, Japan.
J Neurosci Res. 2002 Sep 15;69(6):934-9. doi: 10.1002/jnr.10363.
A method of inducing dopamine (DA) neurons from mouse embryonic stem (ES) cells by stromal cell-derived inducing activity (SDIA) was previously reported. When transplanted, SDIA-induced DA neurons integrate into the mouse striatum and remain positive for tyrosine hydroxylase (TH) expression. In the present study, to optimize the transplantation efficiency, we treated mouse ES cells with SDIA for various numbers of days (8-14 days). SDIA-treated ES cell colonies were isolated by papain treatment and then grafted into the 6-hydroxydopamine (6-OHDA)-lesioned mouse striatum. The ratio of the number of surviving TH-positive cells to the total number of grafted cells was highest when ES cells were treated with SDIA for 12 days before transplantation. This ratio revealed that grafting cell colonies was more efficient for obtaining TH-positive cells in vivo than grafting cell suspensions. When we grafted a cell suspension of 2 x 10(5), 2 x 10(4), or 2 x 10(3) cells into the 6-OHDA-lesioned mouse striatum, we observed only a few surviving TH-positive cells. In conclusion, inducing DA neurons from mouse ES cells by SDIA for 12 days and grafting cell colonies into mouse striatum was the most effective method for the survival of TH-positive neurons in vivo.
此前曾报道过一种通过基质细胞衍生诱导活性（SDIA）从小鼠胚胎干细胞诱导生成多巴胺（DA）神经元的方法。移植后，SDIA诱导的DA神经元可整合到小鼠纹状体中，并保持酪氨酸羟化酶（TH）表达阳性。在本研究中，为优化移植效率，我们用SDIA处理小鼠胚胎干细胞不同天数（8 - 14天）。经木瓜蛋白酶处理分离出经SDIA处理的胚胎干细胞集落，然后将其移植到6 - 羟基多巴胺（6 - OHDA）损伤的小鼠纹状体中。当在移植前用SDIA处理胚胎干细胞12天时，存活的TH阳性细胞数与移植细胞总数的比例最高。该比例表明，在体内获取TH阳性细胞时，移植细胞集落比移植细胞悬液更有效。当我们将2×10⁵、2×10⁴或2×10³个细胞的细胞悬液移植到6 - OHDA损伤的小鼠纹状体中时，仅观察到少数存活的TH阳性细胞。总之，用SDIA诱导小鼠胚胎干细胞12天并将细胞集落移植到小鼠纹状体中是使TH阳性神经元在体内存活的最有效方法。
