Human neural stem cells (hNSCs) have potential as a cell therapy after traumatic brain injury (TBI). While various studies have demonstrated the efficacy of NSCs from ongoing culture, there is a significant gap in our understanding of freshly thawed cells from cryobanked stocks—a more clinically relevant source. To address these shortfalls, the therapeutic potential of our previously validated Shef-6.0 human embryonic stem cell (hESC)-derived hNSC line was tested after long-term cryostorage and thawing before transplant. Immunodeficient athymic nude rats received a moderate unilateral controlled cortical impact (CCI) injury. At four weeks post-injury, 6 × 10⁵ freshly thawed hNSCs were transplanted into six injection sites (two ipsi- and four contra-lateral) with 53.4% of cells surviving three months post-transplant. Interestingly, most hNSCs were engrafted in the meninges and the lining of lateral ventricles, associated with high CXCR4 expression and a chemotactic response to SDF1alpha (CXCL12). While some expressed markers of neuron, astrocyte, and oligodendrocyte lineages, the majority remained progenitors, identified through doublecortin expression (78.1%). Importantly, transplantation resulted in improved spatial learning and memory in Morris water maze navigation and reduced risk taking in an elevated plus maze. Investigating potential mechanisms of action, we identified an increase in ipsilateral host hippocampus cornu ammonis (CA) neuron survival, contralateral dentate gyrus (DG) volume, and DG neural progenitor morphology as well as a reduction in neuroinflammation. Together, these findings validate the potential of hNSCs to improve function after TBI and demonstrate that long-term biobanking of cells and thawing aliquots before use may be suitable for clinical deployment.

中文翻译：

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新鲜解冻的冷冻库人类神经干细胞移植到内源性神经源性微环境中并在慢性创伤性脑损伤后恢复认知功能

人类神经干细胞（hNSC）具有作为创伤性脑损伤（TBI）后细胞疗法的潜力。虽然各种研究已经证明了持续培养的 NSC 的功效，但我们对冷冻库储存的新鲜解冻细胞（一种与临床更相关的来源）的理解存在显着差距。为了解决这些不足，我们之前验证的 Shef-6.0 人胚胎干细胞 (hESC) 衍生的 hNSC 系在移植前长期冷冻保存和解冻后的治疗潜力进行了测试。免疫缺陷的无胸腺裸鼠受到中度单侧控制性皮质撞击（CCI）损伤。损伤后 4 周，将 6  ×  10 ^{5 个}新鲜解冻的 hNSC 移植到六个注射部位（两个同侧，四个对侧），53.4% 的细胞在移植后三个月存活。有趣的是，大多数 hNSC 被植入脑膜和侧脑室内壁，与 CXCR4 的高表达和对 SDF1α (CXCL12) 的趋化反应相关。虽然一些表达神经元、星形胶质细胞和少突胶质细胞谱系的标记物，但大多数仍然是祖细胞，通过双皮质素表达来鉴定（78.1%）。重要的是，移植改善了莫里斯水迷宫导航的空间学习和记忆，并减少了高架十字迷宫中的风险。通过研究潜在的作用机制，我们发现同侧宿主海马角（CA）神经元存活率、对侧齿状回（DG）体积和DG神经祖细胞形态有所增加，并且神经炎症有所减少。总之，这些发现验证了 hNSC 在 TBI 后改善功能的潜力，并证明细胞的长期生物库和使用前解冻等分试样可能适合临床部署。