| 李长明.3D打印支架联合IPSCs-NSCs对大鼠脊髓损伤后神经细胞凋亡及运动功能的变化[J].浙江中西医结合杂志,2021,31(6): |
| 3D打印支架联合IPSCs-NSCs对大鼠脊髓损伤后神经细胞凋亡及运动功能的变化 |
| The effect of three-dimensional scaffolds seeded with IPSCs-NSCs apotosis and locomotor recovery in a rat model of spinal cord injury |
| 投稿时间：2020-10-23 修订日期：2021-04-10 |
| DOI： |
| 中文关键词: 脊髓损伤 神经干细胞 仿生支架 尿 神经元细胞凋亡 |
| 英文关键词:Spinal cord injuries Neural stem cells Bionic scaffold Urine Neuronal apoptosis |
| 基金项目:浙江省医药卫生科技计划项目（2018248755） |
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| 中文摘要: |
| 目的：3D打印载重编程IPSCs-NSCs移植对大鼠脊髓损伤(spinal cord injurys,SCI)的作用及其相关机制。方法：制备3D脊髓支架，将UC细胞来源iPS细胞分化为神经干细胞（NSCs），选择成年雄性SD大鼠42只，按随机数字表法分为假手术组、模型组、IPSCs-NSCs组，每组各14只。采用改良 Allen’s 重物打击法制作SCI大鼠模型，1周后假手术组不做处理，模型组植入DMEM 培养液0.2ml，iPSCs-NSCs 组植入载iPSCs-NSCs的2~3mm支架预处理。于实验干预后1天、3天、7天、14天、28天采用 BBB 评分评估大鼠运动能力，28天后处死大鼠取伤段脊髓组织做病理切片，HE染色观察脊髓组织改变；免疫组化与Western-blot检测脊髓组织中Bax、Bcl-2、Caspase3的表达。结果：①BBB评分：术后 7、14、28天iPSCs-NSCs组与模型组下肢功能均有恢复（P<0.05），且iPSCs-NSCs组较模型组恢复更明显［（3.76±0.65）分比（2.22±0.46）分，
（7.02±0.68）分比（4.41±0.42）分，（11.72±0.81）分比（7.52±0.53）分，P<0.05］。②HE染色：假手术组脊神经生长良好；模型组脊神经受到损伤，组织水肿，细胞稀疏，细胞之间的间隙增大；IPSCs-NSCs组细胞生长较模型组紧密，各组织生长间隙缩小，空泡减小，组织水肿消失。③免疫组化检测：IPSCs-NSCs组Bcl-2阳性细胞的表达较模型组增加［（0.32±0.02）个比（0.14±0.01）个，P<0.05］；IPSCs-NSCs组Bax阳性细胞的表达较模型组少［（0.08±0.00）个比（0.23±0.02）个，P<0.05］；④RT-PCR检测：IPSCs-NSCs组Caspase-3的mRNA含量较模型组减少［（0.96±0.07）ng/L比（1.33±0.03）ng/L，P<0.05］；结论：3D打印支架联合IPSCs-NSCs移植治疗SCI可以显著促进Bcl-2的表达，降低Bax、Caspase3 的表达，改善SCI引起的运动功能下降，其机制可能与上调Bcl-2的表达及下调Bax、Caspase3表达，降低脊髓神经元细胞凋亡相关。 |
| 英文摘要: |
| Objective To investigate the role of three-dimensional scaffolds seeded with IPSCs-NSCs for repair of spinal cord injury in rats and investigate the underlying mechanism. Methods A new three-dimensional bio-printer was used to make bionic spinal cord scaffolds.IPS cells derived from UC cells were differentiated into neural stem cells (NSCs).Forty-two healthy male SD rats were divided into sham group, control group,IPSCs-NSCs group with 14 rats per group,according to the random number table,14 in each group.Spinal cord injury in rats was induced using the electric controlled cortical impactor a week later.Rat neurological motor function was evaluated with BBB score was evaluated with Tarlov and Rivlin scores at postoperative 1-28 days.At 28 d,all rats were killed, He staining was used to observe the changes of the injured spinal cord.Immunofluorescence staining and RT-PCR were used to detect the expression of apoptotic related proteins expression
(Bcl-2, Bax, Caspase 3).Results: Both of the IPSCs-NSCs group and model group had locomotor function recovery on postoperative day 7 ,14 and 28(P<0.05)，but the IPSCs-NSCs group had significantly better locomotor function recov- ery than model group did （D7：3.76±0.65 vs 2.22±0.46; D14:7.02±0.68 vs 4.41±0.42; D28:11.72±0.81 vs 7.52±0.53, all P<0.05）.HE staining showed that the cell growth in iPSCs-NSCs group was more compact than that in model group, and the growth gap and vacuoles of each tissue were reduced, and tissue edema disappeared; Compared with model group，the iPSCs-NSCs group had reduced expression of Bax（0.08±0.00 vs 0.23±0.02,P<0.05）,increased expression of Bcl-2（0.32±0.02 vs 0.14±0.01,P<0.05）.Compared with model group，the iPSCs-NSCs group had reduced mRNA expression of caspase-3（0.96±0.07 vs 1.33±0.03,P<0.05）Condusion ①Three-dimensional scaffolds seeded with IPSCs-NSCs for repair of spinal cord injury in rats can significantly promote the expression of Bcl-2 and decrease the expression of Bax and caspase-3.②Three-dimensional scaffolds seeded with IPSCs-NSCs for repair of spinal cord injury in rats can improve the motor function,The mechanism may be related to up regulating the expression of Bcl-2, down regulating the expression of Bax and caspase 3, and reducing the apoptosis of spinal cord neurons. |
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