Key Laboratory of Health Ministry for Congenital Malformation, Shengjing Hospital, China Medical University, No. 36, Sanhao Street, Heping District, Shenyang, PR China.
Department of Pediatrics, The First Affiliated Hospital of China Medical University, Shenyang, PR China.
Gene Ther. 2020 Dec;27(12):567-578. doi: 10.1038/s41434-020-0131-2. Epub 2020 Feb 24.
Spina bifida aperta is a type of neural tube defect (NTD). Although prenatal fetal surgery has been an available and effective treatment for it, the neurological functional recovery is still need to be enhanced. Our previous results revealed that deficiencies of sensory, motor, and parasympathetic neurons were primary anomalies that occurred with the spinal malformation. Therefore, we emphasized that nerve regeneration is critical for NTD therapy. We delivered an adenoviral construct containing genes inserted for green fluorescent protein and brain-derived neurotrophic factor (Ad-GFP-BDNF) into the amniotic fluid to investigate its prenatal therapeutic potential for rat fetuses with spina bifida aperta. Using immunofluorescence, TdT-mediated dUTP nick-end labeling staining, and real-time polymerase chain reaction analysis, we assessed cell apoptosis in the defective spinal cord and Brn3a positive neuron survival in the dorsal root ganglion (DRG); a protein array was used to investigate the microenvironmental changes of the amniotic fluid. We found that most of the overexpressed BDNF was present on the lesions of the spina bifida fetuses, the number of apoptosis cells in Ad-GFP-BDNF-transfected spinal cords were reduced, mRNA levels of Bcl2/Bax were upregulated and Casp3 were downregulated compared with the controls, the proportion of Brn3a positive neurons in DRG were increased by activating the BDNF/TrkB/Akt signaling pathway, and most of the significant changes in cytokines in the amniotic fluid were related to the biological processes of regulation of apoptotic process and generation of neurons. These results suggest that intra-amniotic Ad-GFP-BDNF gene delivery might have potential as a supplementary approach to treat congenital malformations of neural tubes.
开放性脊柱裂是神经管缺陷（NTD）的一种类型。虽然产前胎儿手术已成为一种可行且有效的治疗方法，但神经功能的恢复仍有待提高。我们之前的结果表明，感觉神经元、运动神经元和副交感神经元的缺失是与脊柱畸形同时发生的主要异常。因此，我们强调神经再生对于 NTD 治疗至关重要。我们将含有插入绿色荧光蛋白和脑源性神经营养因子基因的腺病毒构建体（Ad-GFP-BDNF）递送到羊水中，以研究其对开放性脊柱裂胎鼠的产前治疗潜力。通过免疫荧光、TdT 介导的 dUTP 缺口末端标记染色和实时聚合酶链反应分析，我们评估了缺陷脊髓中的细胞凋亡和背根神经节（DRG）中 Brn3a 阳性神经元的存活情况；使用蛋白质阵列来研究羊水的微环境变化。我们发现，大多数过表达的 BDNF 存在于脊柱裂胎儿的病变部位，与对照组相比，在 Ad-GFP-BDNF 转染的脊髓中，凋亡细胞的数量减少，Bcl2/Bax 的 mRNA 水平上调，Casp3 下调，DRG 中 Brn3a 阳性神经元的比例增加，通过激活 BDNF/TrkB/Akt 信号通路，羊水细胞因子的大多数显著变化与调节凋亡过程和神经元生成的生物学过程有关。这些结果表明，羊水中的 Ad-GFP-BDNF 基因传递可能具有作为治疗神经管先天性畸形的补充方法的潜力。
