Establishment of a system for regulating the gene expression of embryonic mouse cerebral cortex neural stem cells by in utero electroporation

OU Wei-Ming; HE Long-Kai; WANG Xiao-Yu; YANG Xue-Song; WANG Guang; LI Bing-Xiao; JIN Ya; HAN Sha-Sha; LIU Guo-Sheng

doi:10.7499/j.issn.1008-8830.2204096

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Chinese Journal of Contemporary Pediatrics ›› 2022, Vol. 24 ›› Issue (9) : 1061-1067. DOI: 10.7499/j.issn.1008-8830.2204096

EXPERIMENTAL RESEARCH

Establishment of a system for regulating the gene expression of embryonic mouse cerebral cortex neural stem cells by in utero electroporation

OU Wei-Ming, HE Long-Kai, WANG Xiao-Yu, YANG Xue-Song, WANG Guang, LI Bing-Xiao, JIN Ya, HAN Sha-Sha, LIU Guo-Sheng

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Abstract

Objective To establish a system for regulating the gene expression of embryonic mouse cerebral cortex neural stem cells (NSCs) using in utero electroporation (IUE). Methods At embryonic day 14.5, the mouse cerebral cortex NSCs were electro-transfected with the pCIG plasmid injected into the ventricle of the mouse embryo. At embryonic day 16.5 or day 17.5, embryonic mouse brain tissues were collected to prepare frozen sections. Immunofluorescence staining was used to observe the proliferation, apoptosis, division, directional differentiation, migration, and maturation of NSCs. Results The differentiation of NSCs into intermediate progenitors, the proliferation and apoptosis of NSCs, and the morphological development of radial axis of radial glial cells were observed at embryonic day 16.5. The differentiation of NSCs into neurons in layers V-VI of the cerebral cortex, the migration of NSCs to the lateral cerebral cortex, the development of dendrites of migrating neurons, and the maturation of neurons were observed at embryonic day 17.5. Conclusions The system for regulating the gene expression of embryonic mouse cerebral cortex NSCs can be established using IUE, which is useful for the study of neural development related to the proliferation, apoptosis, division, directional differentiation, migration and maturation of NSCs in the cerebral cortex. Citation:Chinese Journal of Contemporary Pediatrics, 2022, 24(9): 1061-1067

Key words

In utero electroporation / Cerebral cortex / Neural stem cell / Neurogenesis / Regulation of gene expression / Mouse

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OU Wei-Ming, HE Long-Kai, WANG Xiao-Yu, YANG Xue-Song, WANG Guang, LI Bing-Xiao, JIN Ya, HAN Sha-Sha, LIU Guo-Sheng. Establishment of a system for regulating the gene expression of embryonic mouse cerebral cortex neural stem cells by in utero electroporation[J]. Chinese Journal of Contemporary Pediatrics. 2022, 24(9): 1061-1067 https://doi.org/10.7499/j.issn.1008-8830.2204096

References

1 Lindsey BW, Hall ZJ, Heuzé A, et al. The role of neuro-epithelial-like and radial-glial stem and progenitor cells in development, plasticity, and repair[J]. Prog Neurobiol, 2018, 170: 99-114. PMID: 29902500. DOI: 10.1016/j.pneurobio.2018.06.004.
2 Hoerder-Suabedissen A, Molnár Z. Development, evolution and pathology of neocortical subplate neurons[J]. Nat Rev Neurosci, 2015, 16(3): 133-146. PMID: 25697157. DOI: 10.1038/nrn3915.
3 Oliveira NCM, Lins éM, Massirer KB, et al. Translational control during mammalian neocortex development and postembryonic neuronal function[J]. Semin Cell Dev Biol, 2021, 114: 36-46. PMID: 33020045. DOI: 10.1016/j.semcdb.2020.09.006.
4 Dhaya I, Griton M, Konsman JP. Magnetic resonance imaging under isoflurane anesthesia alters cortical cyclooxygenase-2 expression and glial cell morphology during sepsis-associated neurological dysfunction in rats[J]. Animal Model Exp Med, 2021, 4(3): 249-260. PMID: 34557651. PMCID: PMC8446714. DOI: 10.1002/ame2.12167.
5 徐瑄培, 黄凌依, 赵凤艳, 等. 鸢尾素对新生大鼠缺氧缺血性脑损伤的作用及机制[J]. 中国当代儿科杂志, 2020, 22(1): 58-64. PMID: 31948526. PMCID: PMC7389714. DOI: 10.7499/j.issn.1008-8830.2020.01.012.
6 Sato M, Takabayashi S, Akasaka E, et al. Recent advances and future perspectives of in vivo targeted delivery of genome-editing reagents to germ cells, embryos, and fetuses in mice[J]. Cells, 2020, 9(4): 799. PMID: 32225003. PMCID: PMC7226049. DOI: 10.3390/cells9040799.
7 dal Maschio M, Ghezzi D, Bony G, et al. High-performance and site-directed in utero electroporation by a triple-electrode probe[J]. Nat Commun, 2012, 3: 960. PMID: 22805567. PMCID: PMC5972006. DOI: 10.1038/ncomms1961.
8 Zhang L, Getz SA, Bordey A. Dual in utero electroporation in mice to manipulate two specific neuronal populations in the developing cortex[J]. Front Bioeng Biotechnol, 2021, 9: 814638. PMID: 35096799. PMCID: PMC8790278. DOI: 10.3389/fbioe.2021.814638.
9 Saito T. In vivo electroporation in the embryonic mouse central nervous system[J]. Nat Protoc, 2006, 1(3): 1552-1558. PMID: 17406448. DOI: 10.1038/nprot.2006.276.
10 Taniguchi Y, Young-Pearse T, Sawa A, et al. In utero electroporation as a tool for genetic manipulation in vivo to study psychiatric disorders: from genes to circuits and behaviors[J]. Neuroscientist, 2012, 18(2): 169-179. PMID: 21551077. PMCID: PMC3530425. DOI: 10.1177/1073858411399925.
11 Moon HM, Hippenmeyer S, Luo L, et al. LIS1 determines cleavage plane positioning by regulating actomyosin-mediated cell membrane contractility[J]. Elife, 2020, 9: e51512. PMID: 32159512. PMCID: PMC7112955. DOI: 10.7554/eLife.51512.
12 Pearson CA, Moore DM, Tucker HO, et al. Foxp1 regulates neural stem cell self-renewal and bias toward deep layer cortical fates[J]. Cell Rep, 2020, 30(6): 1964-1981.e3. PMID: 32049024. PMCID: PMC8397815. DOI: 10.1016/j.celrep.2020.01.034.
13 Baumgart J, Grebe N. C57BL/6-specific conditions for efficient in utero electroporation of the central nervous system[J]. J Neurosci Methods, 2015, 240: 116-124. PMID: 25445056. DOI: 10.1016/j.jneumeth.2014.11.004.
14 Comer AL, Sriram B, Yen WW, et al. A pipeline using bilateral in utero electroporation to interrogate genetic influences on rodent behavior[J]. J Vis Exp, 2020(159): e61350. PMID: 32510510. DOI: 10.3791/61350.
15 Farhy-Tselnicker I, Allen NJ. Astrocytes, neurons, synapses: a tripartite view on cortical circuit development[J]. Neural Dev, 2018, 13(1): 7. PMID: 29712572. PMCID: PMC5928581. DOI: 10.1186/s13064-018-0104-y.
16 Nishimura YV, Shinoda T, Inaguma Y, et al. Application of in utero electroporation and live imaging in the analyses of neuronal migration during mouse brain development[J]. Med Mol Morphol, 2012, 45(1): 1-6. PMID: 22431177. DOI: 10.1007/s00795-011-0557-0.
17 瞿航, 陈爱国, 陈锐泓, 等. 聋哑儿童大脑灰质皮层结构及白质纤维的改变[J]. 中华行为医学与脑科学杂志, 2020, 29(11): 978-982. DOI: 10.3760/cma.j.cn371468-20191112-00832.
18 陈娇阳, 杨莹, 牛雪阳, 等. 新生儿期起病的遗传性癫痫141例致病基因和表型特点[J]. 中华儿科杂志, 2021, 59(9): 767-771. PMID: 34645217. DOI: 10.3760/cma.j.cn112140-20210206-00113.
19 László ZI, Lele Z, Z?ldi M, et al. ABHD4-dependent developmental anoikis safeguards the embryonic brain[J]. Nat Commun, 2020, 11(1): 4363. PMID: 32868797. PMCID: PMC7459116. DOI: 10.1038/s41467-020-18175-4.
20 Da Silva F, Zhang K, Pinson A, et al. Mitotic WNT signalling orchestrates neurogenesis in the developing neocortex[J]. EMBO J, 2021, 40(19): e108041. PMID: 34431536. PMCID: PMC8488556. DOI: 10.15252/embj.2021108041.
21 Geng A, Qiu R, Murai K, et al. KIF20A/MKLP2 regulates the division modes of neural progenitor cells during cortical development[J]. Nat Commun, 2018, 9(1): 2707. PMID: 30006548. PMCID: PMC6045631. DOI: 10.1038/s41467-018-05152-1.
22 Nakagawa N, Plestant C, Yabuno-Nakagawa K, et al. Memo1-mediated tiling of radial glial cells facilitates cerebral cortical development[J]. Neuron, 2019, 103(5): 836-852.e5. PMID: 31277925. PMCID: PMC6728225. DOI: 10.1016/j.neuron.2019.05.049.
23 Zhong S, Zhao Z, Xie W, et al. GABAergic interneuron and neurotransmission are mTOR-dependently disturbed in experimental focal cortical dysplasia[J]. Mol Neurobiol, 2021, 58(1): 156-169. PMID: 32909150. DOI: 10.1007/s12035-020-02086-y.
24 俞华莉, 巨兴达, 王俊杰, 等. 鼠胚大脑皮层子宫内电穿孔法基因转移体系的建立[J]. 神经解剖学杂志, 2009, 25(1): 25-29.
25 邹明明, 倪莉, 周立宇, 等. 子宫内电穿孔转染胎鼠脑室管膜下区神经干细胞的条件优化[J]. 中国组织工程研究, 2022, 26(30): 4879-4883. DOI: 10.12307/2022.769.