Laboratory of Molecular Neurosurgery, Department of Stereotactic and Functional Neurosurgery, Neurocentre, University Hospital Freiburg, Freiburg, Germany.
Cell Transplant. 2012;21(9):1837-51. doi: 10.3727/096368911X627426. Epub 2012 Mar 8.
Stem cell transplantation is a promising therapeutic approach in neurodegenerative diseases. Studying graft survival and development has important implications for the further development of experimental and clinical transplantation protocols. Cellular elements in neural transplants are sometimes difficult to identify. The existing labeling methods cannot reliably provide stably labeled cells that can be detected in long-term experiments. Transgenic (tg) Lewis rats ubiquitously expressing green fluorescent protein (GFP) provide an ideal donor source. The aim of this project was to investigate the potential of GFP-tg Lewis rats to serve as donor tissue for neural stem cell transplantation. Ventral mesencephalon (VM) GFP-tg E14.5-derived cells were compared to wild-type (wt) in vitro and in vivo. Firstly, cells from GFP and non-GFP VM tissue were compared with regard to their proliferation and response towards 6-OHDA-toxicity in culture. Secondly, 6-OHDA-lesioned hemiparkinsonian Sprague-Dawley/Crl:CD(SD) rats received intrastriatal grafts derived from VM of E14.5 GFP-tg rats. Due to the fact that donor and recipient belong to two different rat strains, we focused on graft survival in correlation with immunosuppression and graft GFP and tyrosine hydroxylase (TH) expression. In summary, in vitro tg cells exhibited 98% GFP expression and did not differ from wt cells in any of the measured parameters. In vivo, all experimental groups showed a significant compensation in rotation behavior after transplantation. Furthermore, there was no difference on rotation behavior or graft morphology and survival pattern as well as GFP expression between immunosuppressed and nonimmunosuppressed animals. The GFP-positive population of the graft was composed of 13.3% GFAP-positive, 56.1% NeuN-positive, and 1.9% TH-positive cells. Analysis of graft subpopulations manifested that 70.6% of GFAP-positive, 86.9% of NeuN-positive, and 80.1% of TH-positive cells coexpressed GFP. In conclusion, our data show that the Lewis GFP-tg rats serve as an excellent cell source for studying primary neural precursor cells in the transplantation paradigm.
干细胞移植是神经退行性疾病有前途的治疗方法。研究移植物的存活和发育对进一步发展实验和临床移植方案具有重要意义。神经移植中的细胞成分有时难以识别。现有的标记方法不能可靠地提供可在长期实验中检测到的稳定标记细胞。泛表达绿色荧光蛋白(GFP)的转基因(tg)Lewis 大鼠提供了理想的供体来源。本项目旨在研究 GFP-tgLewis 大鼠作为神经干细胞移植供体组织的潜力。比较了腹侧中脑(VM)E14.5 衍生的 GFP-tg 细胞与野生型(wt)细胞在体外和体内的情况。首先,比较 GFP 和非 GFP VM 组织的细胞在培养中增殖和对 6-OHDA 毒性的反应。其次,6-OHDA 致半帕金森病 Sprague-Dawley/Crl:CD(SD)大鼠接受源自 E14.5 GFP-tg 大鼠 VM 的纹状体移植。由于供体和受体属于两个不同的大鼠品系,我们重点研究了与免疫抑制和移植 GFP 及酪氨酸羟化酶(TH)表达相关的移植物存活情况。总之,体外 tg 细胞的 GFP 表达率为 98%,并且在任何测量参数上都与 wt 细胞没有差异。在体内,所有实验组在移植后旋转行为均有明显改善。此外,在免疫抑制和非免疫抑制动物之间,在旋转行为或移植物形态和存活模式以及 GFP 表达方面没有差异。移植物中 GFP 阳性细胞群由 13.3%GFAP 阳性、56.1%NeuN 阳性和 1.9%TH 阳性细胞组成。移植物亚群分析表明,70.6%GFAP 阳性、86.9%NeuN 阳性和 80.1%TH 阳性细胞共表达 GFP。总之,我们的数据表明,Lewis GFP-tg 大鼠是研究移植模型中初级神经前体细胞的优秀细胞来源。