Medical Neuroscience Dalhousie University Halifax Nova Scotia Canada; Department of Surgery (Neurosurgery) Dalhousie University Halifax Nova Scotia Canada B3H 4R2.
Medical Neuroscience Dalhousie University Halifax Nova Scotia Canada; Department of Surgery (Neurosurgery) Dalhousie University Halifax Nova Scotia Canada B3H 4R2; Sobell Department of Motor Neuroscience and Movement Disorders Institute of Neurology University College London London WC1N 3BG United Kingdom.
Ann Clin Transl Neurol. 2016 Jul 1;3(8):637-49. doi: 10.1002/acn3.327. eCollection 2016 Aug.
Nerve injuries resulting in prolonged periods of denervation result in poor recovery of motor function. We have previously shown that embryonic stem cell-derived motoneurons transplanted at the time of transection into a peripheral nerve can functionally reinnervate muscle. For clinical relevance, we now focused on delaying transplantation to assess reinnervation after prolonged denervation.
Embryonic stem cell-derived motoneurons were transplanted into the distal segments of transected tibial nerves in adult mice after prolonged denervation of 1-8 weeks. Twitch and tetanic forces were measured ex vivo 3 months posttransplantation. Tissue was harvested from the transplants for culture and immunohistochemical analysis.
In this delayed reinnervation model, teratocarcinomas developed in about one half of transplants. A residual multipotent cell population (~ 6% of cells) was found despite neural differentiation. Exposure to the alkylating drug mitomycin C eliminated this multipotent population in vitro while preserving motoneurons. Treating neural differentiated stem cells prior to delayed transplantation prevented tumor formation and resulted in twitch and tetanic forces similar to those in animals transplanted acutely after denervation.
Despite a neural differentiation protocol, embryonic stem cell-derived motoneurons still carry a risk of tumorigenicity. Pretreating with an antimitotic agent leads to survival and functional muscle reinnervation if performed within 4 weeks of denervation in the mouse.
导致去神经支配期延长的神经损伤会导致运动功能恢复不良。我们之前已经表明，在周围神经横断时移植的胚胎干细胞衍生的运动神经元可以功能性地再支配肌肉。为了达到临床相关性，我们现在专注于延迟移植，以评估延长去神经支配后的再神经支配。
在成年小鼠的胫神经横断后延长去神经支配 1-8 周后，将胚胎干细胞衍生的运动神经元移植到横断的远端段。移植后 3 个月，进行体外抽搐和强直力测量。从移植组织中进行培养和免疫组织化学分析。
在这个延迟再神经支配模型中，大约一半的移植物中出现了畸胎瘤。尽管发生了神经分化，但仍存在残余的多能细胞群（约 6%的细胞）。暴露于烷化剂丝裂霉素 C 可在体外消除这种多能细胞群，同时保留运动神经元。在延迟移植前对神经分化的干细胞进行预处理可防止肿瘤形成，并导致抽搐和强直力与去神经支配后急性移植的动物相似。
尽管采用了神经分化方案，但胚胎干细胞衍生的运动神经元仍存在致瘤风险。在小鼠去神经支配后 4 周内，如果进行抗有丝分裂药物预处理，可导致存活和功能性肌肉再神经支配。
