Objective To investigate the effect of Fasudil in improving axon and myelin regeneration and functional recovery after peripheral nerve injury. Methods Thirty SD rats weighing (200±30) g were randomly divided into two groups: control group and Fasudil group. Normal saline and 10 mg/kg hydrochloride Fasudil were intraperitoneal given, respectively. The right sciatic nerve was transected and sutured. Two weeks after operation, the density of NF-200 positive axons at the distal end of the injury was evaluated. Four weeks after operation, the number of neurons in the L4-6 dorsal root ganglia (DRG) and sacrolumbar enlargement that sent axons to the nerve distal to the transection site was evaluated by retrograde tracing using Fluoro-gold. Twelve weeks after operation, the wet weight ratio of gastrocnemius muscle and the cross-sectional area of muscle fibers were measured; and immunohistochemistry using primary NF-200 and MPZ antibodies and transmission electron microscopy were used to measure the diameter and thickness of axons and myelin sheaths. The footprints of rats at 4, 6, 8, 10, and 12 weeks after operation were collected to evaluate the sciatic function index (SFI) of rats in the two groups. In addition, rat embryonic dorsal root ganglion (DRG) and spinal motor neurons (SMN) were cultured to evaluate the effect of Fasudil on axon growth. Results At 14 days after operation, the density of NF-200 positive axons at the distal end of the Fasudil group was significantly higher than that of control group (P=0.034). Four weeks after operation, the number of L4-6 DRG and spinal ventral horn motor neurons retrogradely labeled by Fluoro-gold in Fasudil group was significantly higher than that in control group (P<0.05). Twelve weeks after operation, the number of myelinated axons, diameter of myelinated axons, and thickness of myelin sheaths in Fasudil group was higher than those in control group (P<0.05), while G-ratio value was lower than that in control group (P<0.05). The SFI data showed that at 6, 8, 10, and 12 weeks after operation, the SFI values in Fasudil group were significantly better than those in control group (P<0.05). Twelve weeks after operation, the wet weight ratio of gastrocnemius muscle and cross-sectional area of myofibrils in control group were significantly smaller than those in Fasudil group (P<0.01). After culturing for 5 days, the axons of DRG and SMN in Fasudil group were significantly longer than those in control group (P<0.01). Conclusions Fasudil can promote axon and myelin regeneration after sciatic nerve injury in rats and improve their motor function recovery.
Key words
ROCK inhibitors; /
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Fasudil; /
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Sciatic nerve injury; /
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Axon and myelin regeneration
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References
[1] Lin J, Shi J, Min X, et al. The GDF11 promotes nerve regeneration after sciatic nerve injury in adult rats by promoting axon growth and inhibiting neuronal apoptosis[J]. Front Bioeng Biotechnol, 2022, 9:803052. DOI:10.3389/fbioe.2021.803052.
[2] Wariyar SS, Ward PJ. Application of electrical stimulation to enhance axon regeneration following peripheral nerve injury[J]. Bio Protoc, 2023, 13(19):e4833. DOI:10.21769/BioProtoc.4833.
[3] Trenton MG, Wesley NS, Kacey GM. The role of chondroitinase as an adjuvant to peripheral nerve repair[J]. Cells Tissues Organs, 2014, 200(1):59-68. DOI: 10.1159/000369449.
[4] Choi EK, Kim JG, Kim HJ, et al. Regulation of RhoA GTPase and novel target proteins for ROCK[J]. Small GTPases, 2020, 11(2):95-102. DOI:10.1080/21541248.2017.1364831.
[5] Hall A, Lalli G. Rho and Ras GTPases in axon growth, guidance, and branching[J]. Cold Spring Harb Perspect Biol, 2010, 2(2):a001818. DOI:10.1101/cshperspect.a001818.
[6] Maruhashi T, Higashi Y. An overview of pharmacotherapy for cerebral vasospasm and delayed cerebral ischemia after subarachnoid hemorrhage[J]. Expert Opin Pharmacother, 2021, 22(12):1-14. DOI:10.1080/14656566.2021.1912013.
[7] Wang H, Fang F, Chen SF et al. Dual efficacy of Fasudil at improvement of survival and reinnervation of flap through RhoA/ROCK/PI3K/Akt pathway[J]. Int Wound J, 2022, 19(8):2000-2011. DOI: 10.1111/iwj.13800.
[8] Xu ZX, Albayar A , Dollé JP, et al. Dorsal root ganglion axons facilitate and guide cortical neural outgrowth: In vitro modeling of spinal cord injury axonal regeneration[J]. Restor Neurol Neurosci, 2020, 38(1):1-9. DOI: 10.3233/RNN-190933.
[9] Graber DJ, Harris BT. Purification and culture of spinal motor neurons from rat embryos[J]. Cold Spring Harb Protoc, 2013, 2013(4):319-326. DOI:10.1101/pdb.prot074161.
[10] Benga A, Zor F, Korkmaz A, et al. The neurochemistry of peripheral nerve regeneration[J]. Indian J Plast Surg, 2017, 50(1):5-15. DOI:10.4103/ijps.IJPS1417.
[11] Lavorato A, Aruta G, De Marco R, et al. Traumatic peripheral nerve injuries: a classification proposal[J]. J Orthop Traumatol, 2023, 24(1):20. DOI:10.1186/s10195-023-00695-6.
[12] Hiraga A, Kuwabara S, Doya H, et al. Rho-kinase inhibition enhances axonal regeneration after peripheral nerve injury[J]. J Peripher Nerv Syst, 2006, 11(3):217-24. DOI: 10.1111/j.1529-8027.2006.00091.x.
[13] Joshi A, Bobylev I, Zhang G, et al. Inhibition of Rho-kinase differentially affects axon regeneration of peripheral motor and sensory nerves[J]. Exp Neurol, 2015, 263:28-38. DOI: 10.1016/j.expneurol.2014.09.012.
[14] Melendez-Vasquez CV, Einheber S, Salzer JL. Rho kinase regulates schwann cell myelination and formation of associated axonal domains[J]. J Neurosci, 2004, 24(16):3953. DOI:10.1523/JNEUROSCI.4920-03.2004.
[15] Wen J, Tan D, Li L , et al. RhoA regulates Schwann cell differentiation through JNK pathway[J]. Exp Neurol, 2018, 308:26-34. DOI:10.1016/j.expneurol.2018.06.013.
[16] Tan D, Wen J, Li L, et al. Inhibition of RhoA-Subfamily GTPases suppresses schwann cell proliferation through regulating AKT pathway rather than ROCK pathway[J]. Front Cell Neurosci, 2018, 12:437. DOI:10.3389/fncel.2018.00437.
[17] Chen M, Liu A, Ouyang Y, et al. Fasudil and its analogs: a new powerful weapon in the long war against central nervous system disorders[J]? Expert Opin Investig Drugs, 2013, 22(4):537-50. DOI:10.1517/13543784.2013.778242.
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