Improving neuronal transfection by electroporation of dorsal
root ganglion

doi:10.3969/j.issn.2095-4344.2072

Abstract

Abstract:

BACKGROUND: Electroporation of dorsal root ganglion is a high-efficiency gene transfection method to study nerve regeneration. In the past, the voltage condition of dorsal root ganglion electroporation resulted in a reduction in the number of labeled neurons and axons, with a high statistical error.

OBJECTIVE: To improve the marker rate of neurons and their axons, and to provide theoretical basis for the study of peripheral nerve regeneration.

METHODS: The enhanced green fluorescent protein was as an outcome measure to optimize dorsal root ganglion electroporation in axonal regeneration. ICR mice were randomly divided into two groups, respectively, and underwent dorsal root ganglion electroperforation surgery to detect the labeling rates of neurons and their axons under the intervention of 35 and 60 V voltages.

RESULTS AND CONCLUSION: Voltages at 35 and 60 V did not cause significant neuronal death. Compared with 35 V voltage, 60 V voltage significantly increased the labeling rate of neurons and their axons as well as the number of axons passing through the injured site (P < 0.05). The 60 V voltage did not damage the behavioral function of the experimental animals. These results suggest that 60 V voltage can increase the labeling rate of neurons and their axons, providing a basis for the study of axonal regeneration in peripheral nerves.

Key words: dorsal root ganglion, axon, electroporation, voltage, peripheral nerve, labeling rate

CLC Number:

Qi Shibin, Ma Yanxia, Zhang Haonan, Ma Jinjin, Xu Jinhui, Saijilafu. Improving neuronal transfection by electroporation of dorsal root ganglion[J]. Chinese Journal of Tissue Engineering Research, 2020, 24(19): 3042-3047.

Figures/Tables 5

References

[1] NOBLE J, MUNRO CA, PRASAD VS, et al. Analysis of upper and lower extremity peripheral nerve injuries in a population of patients with multiple injuries. J Trauma. 1998;45(1): 116-122.
[2] 陈明伟,袁晓华,潘敏,等.从道路交通事故统计分析对比谈预防措施[J].中国安全科学学报, 2004,14(8):59-63.
[3] 韦正超,蔡道章,徐柜如,等.医源性外周神经损伤临床治疗分析[J].中华显微外科杂志,2002,25(2):152-153.
[4] 李志杰,高伟阳,廖孔荣,等.医源性周围神经损伤及显微外科治疗[J].温州医学院学报,2002,32(2): 115-116.
[5] 张国亮,蔡启卿,冯明录.医源性周围神经损伤的治疗[J].中国矫形外科杂志,2000,7(5):503.
[6] MOKARRAM N, BELLAMKONDA RV. Overcoming endogenous constraints on neuronal regeneration. IEEE Trans Biomed Eng. 2011;58(7):1900-1906.
[7] JACOB HE, FÖRSTER W, BERG H. Microbiological implications of electric field effects. II. Inactivation of yeast cells and repair of their cell envelope. Z Allg Mikrobiol. 1981; 21(3):225-233.
[8] SAIJILAFU, ZHANG BY, ZHOU FQ. In vivo electroporation of adult mouse sensory neurons for studying peripheral axon regeneration. Methods Mol Biol. 2014;1162:167-175.
[9] LI Q, QIAN C, ZHOU FQ. Investigating Mammalian Axon Regeneration: In Vivo Electroporation of Adult Mouse Dorsal Root Ganglion. J Vis Exp. 2018;(139):58171.
[10] BASSO DM, FISHER LC, ANDERSON AJ, et al. Basso Mouse Scale for locomotion detects differences in recovery after spinal cord injury in five common mouse strains. J Neurotrauma. 2006;23(5):635-659.
[11] FILBIN MT. Myelin-associated inhibitors of axonal regeneration in the adult mammalian CNS. Nat Rev Neurosci. 2003;4(9):703-713.
[12] SAIJILAFU, HUR EM, LIU CM, et al. PI3K-GSK3 signalling regulates mammalian axon regeneration by inducing the expression of Smad1. Nat Commun. 2013;4:2690.
[13] ABE N, CAVALLI V. Nerve injury signaling. Curr Opin Neurobiol. 2008;18(3):276-283.
[14] CHOI J, KIM JH, JANG JW, et al. Decellularized sciatic nerve matrix as a biodegradable conduit for peripheral nerve regeneration. Neural Regen Res. 2018;13(10):1796-1803.
[15] CARLÉN M, MELETIS K, BARNABÉ-HEIDER F, et al. Genetic visualization of neurogenesis. Exp Cell Res. 2006; 312(15):2851-2859.
[16] CONSIGLIO A, GRITTI A, DOLCETTA D, et al. Robust in vivo gene transfer into adult mammalian neural stem cells by lentiviral vectors. Proc Natl Acad Sci U S A. 2004;101(41): 14835-14840.
[17] TASHIRO A, ZHAO C, GAGE FH. Retrovirus-mediated single-cell gene knockout technique in adult newborn neurons in vivo. Nat Protoc. 2006;1(6):3049-3055.
[18] LIU K, LU Y, LEE JK, et al. PTEN deletion enhances the regenerative ability of adult corticospinal neurons. Nat Neurosci. 2010;13(9):1075-1081.
[19] XI F, XU RJ, XU JH, et al. Calcium/calmodulin-dependent protein kinase II regulates mammalian axon growth by affecting F-actin length in growth cone. J Cell Physiol. 2019;234(12):23053-23065.
[20] HIRAKAWA M, MCCABE JT, KAWATA M. et al. Time-related changes in the labeling pattern of motor and sensory neurons innervating the gastrocnemius muscle, as revealed by the retrograde transport of the cholera toxin B subunit. Cell Tissue Res. 1992;267(3):419-427.
[21] BARNABÉ-HEIDER F, MELETIS K, ERIKSSON M, et al. Genetic manipulation of adult mouse neurogenic niches by in vivo electroporation. Nat Methods. 2008;5(2):189-196.
[22] WEAVER JC. Electroporation in cells and tissues: A biophysical phenomenon due to electromagnetic fields. Radio Science.1995;30(1):205-221.
[23] TAMURA T, NISHI T, GOTO T, et al. Intratumoral delivery of interleukin 12 expression plasmids with in vivo electroporation is effective for colon and renal cancer. Hum Gene Ther. 2001; 12(10):1265-1276.
[24] KISHIDA T, ASADA H, SATOH E, et al. In vivo electroporation-mediated transfer of interleukin-12 and interleukin-18 genes induces significant antitumor effects against melanoma in mice. Gene Ther. 2001;8(16): 1234-1240.
[25] PAVSELJ N, BREGAR Z, CUKJATI D, et al. The course of tissue permeabilization studied on a mathematical model of a subcutaneous tumor in small animals. IEEE Trans Biomed Eng. 2005;52(8):1373-1381.
[26] LOTURCO J, MANENT JB, SIDIQI F. New and improved tools for in utero electroporation studies of developing cerebral cortex. Cereb Cortex. 2009;19 Suppl 1:i120-125.
[27] DUAN RS, LIU PP, XI F, et al. Wnt3 and Gata4 regulate axon regeneration in adult mouse DRG neurons. Biochem Biophys Res Commun. 2018;499(2):246-252.
[28] MA JJ, XU RJ, QI SB, et al. Regulation of adult mammalian intrinsic axonal regeneration by NF-κB/STAT3 signaling cascade. J Cell Physiol. 2019;234(12):22517-22528.
[29] 刘颖,周玮,熊正爱,等.不可逆性电穿孔致山羊肝脏组织凋亡与坏死的实验研究[J].解放军医学杂志, 2011,36(4): 356-360.
[30] 姚陈果,孙才新,米彦,等.陡脉冲对恶性肿瘤细胞不可逆性电击穿的实验研究[J].中国生物医学工程学报, 2004,23(1): 92-97.
[31] 梁冰,牛立志,曾健滢,等.不可逆电穿孔消融兔胆囊侧肝脏病理学观察[J].介入放射学杂志,2014,23(4):320-324.
[32] 俞允.基于电穿孔技术的活细胞表面增强拉曼光谱研究[D].福州:福建师范大学,2011.
[33] STEWARD O, ZHENG B, BANOS K, et al. Response to: Kim et al. "axon regeneration in young adult mice lacking Nogo-A/B." Neuron 38, 187-199. Neuron. 2007;54(2): 191-195.
[34] SAIJILAFU, HUR EM, ZHOU FQ. Genetic dissection of axon regeneration via in vivo electroporation of adult mouse sensory neurons. Nat Commun. 2011;2:543.