Wang Le, Wei Fu-Xin, Cen Jing-Sheng, Ping Su-Ning, Li Zi-Qing, Chen Ning-Ning, Cui Shang-Bin, Wan Yong, Liu Shao-Yu
Department of Spine Surgery, The First Affiliated Hospital, Sun Yat-sen University, 58 Zhongshan Rd, Guangzhou, China.
Department of Histology and Embryology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, China.
Brain Res. 2014 Aug 5;1575:87-100. doi: 10.1016/j.brainres.2014.05.038. Epub 2014 Jun 2.
Neural stem cell (NSC) transplantation has been reported to be a leading strategy to stimulate neuroplasticity, repair neuronal loss and promote the morphologic and functional recovery of spinal cord injury (SCI). However, massive death of transplanted NSCs is still a problem, which is considered to be related to a series of pro-inflammatory cytokines that induce apoptosis, extensive demyelination and axonal destruction. Tumor necrosis factor alpha (TNF-α), as one of the major inflammation initiators, contributes to secondary neural cell death. We previously found that the administration of the TNF-α antagonist etanercept during the acute phase of SCI can reduce the apoptosis of neurons and oligodendrocytes. To investigate whether etanercept can suppress transplanted NSC apoptosis and promote NSC survival, axon myelination and functional recovery, we tested the combination strategy of the early administration of etanercept and NSC transplantation. First we observed that etanercept suppressed the TNF-α expression and apoptosis of transplanted NSCs by Western blot, TUNEL and immunofluorescence staining. The Basso, Beattle and Bresnahan scale and motor-evoked potential were used to evaluate functional recovery. The results suggest significantly better recovery after combination therapy. Further, histopathological alterations were evaluated by hematoxylin and eosin staining and Nissl staining. These procedures showed that the early administration of etanercept improved survival of neurons in the ventral horn, restored neural morphology and produced a smaller cavity area. We observed most abundant NF-positive fibers after the combination treatment, indicating that combination therapy retained and promoted neural regeneration. Finally, the early suppression of TNF-α reduced the occurrence of demyelination, and the combination therapy led to more myelinated axons, as shown by electron microscopy. These data suggest that this strategy significantly protected transplanted NSCs via the anti-inflammation and anti-apoptosis effects of etanercept, promoting re-myelination, neural regeneration and locomotor function.
据报道，神经干细胞（NSC）移植是刺激神经可塑性、修复神经元损失以及促进脊髓损伤（SCI）形态和功能恢复的主要策略。然而，移植的神经干细胞大量死亡仍是一个问题，这被认为与一系列诱导细胞凋亡、广泛脱髓鞘和轴突破坏的促炎细胞因子有关。肿瘤坏死因子α（TNF-α）作为主要的炎症引发剂之一，会导致继发性神经细胞死亡。我们之前发现，在脊髓损伤急性期给予TNF-α拮抗剂依那西普可减少神经元和少突胶质细胞的凋亡。为了研究依那西普是否能抑制移植神经干细胞的凋亡并促进其存活、轴突髓鞘形成和功能恢复，我们测试了依那西普早期给药与神经干细胞移植的联合策略。首先，我们通过蛋白质免疫印迹、TUNEL和免疫荧光染色观察到依那西普抑制了移植神经干细胞的TNF-α表达和凋亡。使用Basso、Beattle和Bresnahan评分量表以及运动诱发电位来评估功能恢复情况。结果表明联合治疗后的恢复情况明显更好。此外，通过苏木精-伊红染色和尼氏染色评估组织病理学改变。这些操作表明，依那西普的早期给药改善了腹角神经元的存活，恢复了神经形态，并产生了较小的空洞面积。联合治疗后，我们观察到NF阳性纤维最为丰富，表明联合治疗保留并促进了神经再生。最后，如电子显微镜所示，早期抑制TNF-α减少了脱髓鞘的发生，联合治疗导致更多有髓轴突。这些数据表明，该策略通过依那西普的抗炎和抗凋亡作用显著保护了移植的神经干细胞，促进了再髓鞘形成、神经再生和运动功能。
