CaMKII-Smad1 promotes axonal regeneration of peripheral nerves

doi:10.3969/j.issn.2095-4344.2173

Abstract

Abstract: BACKGROUND: Axons do not regenerate after central nervous system injury in mammals. It is mainly caused by the inhibitory microenvironment at the site of damage and the weakened self-regeneration ability. Studies have found that peripheral nervous system has certain regeneration ability after injury, so we explore the methods of central nervous system repair by studying the genes promoting peripheral nervous system regeneration. As one of the important protein kinase families of neurons, CaMKII up-regulation can improve the ability of neuron regeneration. Similarly, acute depletion of the Smad1 protein in adult mice also prevented axon regeneration in vivo. These genes can directly or indirectly regulate neuronal axon regeneration, but exactly how they regulate neuronal regeneration is still unclear.
OBJECTIVE: To study the effects of CaMKII-Smad1 signaling pathway on axon regeneration of dorsal root ganglion neurons by intraperitoneal injection of CaMKII inhibitor and activator, and explored the mechanism of CaMKII and Smad1 in regulating axon regeneration of dorsal root ganglion neurons.
METHODS: Totally 40 ICR mice were randomly divided into four groups: KN93 control group, KN93 experimental group, CdCl2 control group and CdCl2 experimental group. Dorsal root ganglion tissue was taken for in vitro culture after 7 days of continuous administration of CaMKII inhibitor KN93 and activator CdCl2. The length of axonal regeneration of dorsal root ganglion neurons was statistically analyzed after 3 days. Protein expression of p-Smad1 in dorsal root ganglion neurons was detected using western blot assay.
RESULTS AND CONCLUSION: (1) Compared with the KN93 control group, axonal regeneration of dorsal root ganglion neurons was inhibited, and the p-Smad1 protein expression was decreased in the KN93 experimental group, showing significant differences. (2) Compared with the CdCl2 control group, axonal regeneration of dorsal root ganglion neurons was promoted, and p-Smad1 protein expression was increased in the CdCl2 experimental group, showing significant differences. (3) The results showed that the CaMKII-Smad1 signaling pathway had a regulatory effect on axonal regeneration of dorsal root ganglion neurons.

Key words: dorsal root ganglion, neurons, CaMKII, Smad1, axonal regeneration, intraperitoneal injection, cell culture, mouse

CLC Number:

Wang Feng, Zhou Liyu, Saijilafu, Qi Shibin, Ma Yanxia, Wei Shanwen. CaMKII-Smad1 promotes axonal regeneration of peripheral nerves[J]. Chinese Journal of Tissue Engineering Research, 2021, 25(7): 1064-1068.

Figures/Tables 4

References

[1] ABANKWA D, KÜRY P, MÜLLER HW. Dynamic changes in gene expression profiles following axotomy of projection fibres in the Mammalian CNS. Mol Cell Neurosci. 2002;21(3):421-435.
[2] SILVER J, SCHWAB ME, POPOVICH PG. Central nervous system regenerative failure: role of oligodendrocytes, astrocytes, and microglia. Cold Spring Harb Perspect Biol. 2014;7(3):a020602.
[3] FILBIN MT. Recapitulate development to promote axonal regeneration: good or bad approach? Philos Trans R Soc Lond B Biol Sci. 2006;361 (1473):1565-1574.
[4] GOLDBERG JL, KLASSEN MP, HUA Y, et al. Amacrine-signaled loss of intrinsic axon growth ability by retinal ganglion cells. Science. 2002; 296 (5574):1860-1864.
[5] SCHWAB ME, BARTHOLDI D. Degeneration and regeneration of axons in the lesioned spinal cord. Physiol Rev. 1996;76(2):319-370.
[6] LEE JK, GEOFFROY CG, CHAN AF, et al. Assessing spinal axon regeneration and sprouting in Nogo-, MAG-, and OMgp-deficient mice. Neuron. 2010;66(5):663-670.
[7] SILVER J, MILLER JH. Regeneration beyond the glial scar. Nat Rev Neurosci. 2004;5(2):146-156.
[8] HAMMARLUND M, NIX P, HAUTH L,et al. Axon regeneration requires a conserved MAP kinase pathway. Science. 2009;323(5915):802-806.
[9] NIX P, HISAMOTO N, MATSUMOTO K, et al. Axon regeneration requires coordinate activation of p38 and JNK MAPK pathways. Proc Natl Acad Sci U S A. 2011;108(26):10738-10743.
[10] FAAS GC, RAGHAVACHARI S, LISMAN JE, et al. Calmodulin as a direct detector of Ca2+ signals. Nat Neurosci. 2011;14(3):301-304.
[11] AVERSA Z, ALAMDARI N, CASTILLERO E, et al. CaMKII activity is reduced in skeletal muscle during sepsis. J Cell Biochem. 2013;114(6): 1294-1305.
[12] YAMAUCHI T, FUJISAWA H. Evidence for three distinct forms of calmodulin-dependent protein kinases from rat brain. FEBS Lett. 1980; 116(2):141-144.
[13] KENNEDY MB, GREENGARD P. Two calcium/calmodulin-dependent protein kinases, which are highly concentrated in brain, phosphorylate protein I at distinct sites. Proc Natl Acad Sci U S A. 1981;78(2): 1293-1297.
[14] PENG J, KIM MJ, CHENG D, et al. Semiquantitative proteomic analysis of rat forebrain postsynaptic density fractions by mass spectrometry. J Biol Chem. 2004;279(20):21003-21011.
[15] COLBRAN RJ, SODERLING TR. Calcium/calmodulin-dependent protein kinase II. Curr Top Cell Regul. 1990;31:181-221.
[16] KELLY PT. Calmodulin-dependent protein kinase II. Multifunctional roles in neuronal differentiation and synaptic plasticity. Mol Neurobiol. 1991;5(2-4):153-177.
[17] KENNEDY MB, BENNETT MK, BULLEIT RF, et al. Structure and regulation of type II calcium/calmodulin-dependent protein kinase in central nervous system neurons. Cold Spring Harb Symp Quant Biol. 1990;55:101-110.
[18] YAMAUCHI T, FUJISAWA H. Disassembly of microtubules by the action of calmodulin-dependent protein kinase (Kinase II) which occurs only in the brain tissues. Biochem Biophys Res Commun. 1983;110(1):287-291.
[19] ZOU H, HO C, WONG K, et al. Axotomy-induced Smad1 activation promotes axonal growth in adult sensory neurons. J Neurosci. 2009; 29(22):7116-7123.
[20] 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.
[21] FINELLI MJ, MURPHY KJ, CHEN L, et al. Differential phosphorylation of Smad1 integrates BMP and neurotrophin pathways through Erk/Dusp in axon development. Cell Rep. 2013;3(5):1592-1606.
[22] FARRUKH F, DAVIES E, BERRY M, et al. BMP4/Smad1 signalling promotes spinal dorsal column axon regeneration and functional recovery after injury. Mol Neurobiol. 2019;56(10):6807-6819.
[23] FORMAN DS, MCQUARRIE IG, LABORE FW, et al. Time course of the conditioning lesion effect on axonal regeneration. Brain Res. 1980; 182(1):180-185.
[24] PARIKH P, HAO Y, HOSSEINKHANI M, et al. Regeneration of axons in injured spinal cord by activation of bone morphogenetic protein/Smad1 signaling pathway in adult neurons. Proc Natl Acad Sci U S A. 2011;108(19):E99-107.
[25] YAN D, WU Z, CHISHOLM AD, et al. The DLK-1 kinase promotes mRNA stability and local translation in C. elegans synapses and axon regeneration. Cell. 2009;138(5):1005-1018.
[26] PASTUHOV SI, FUJIKI K, NIX P, et al. Endocannabinoid-Goα signalling inhibits axon regeneration in Caenorhabditis elegans by antagonizing Gqα-PKC-JNK signalling. Nat Commun. 2012;3:1136.
[27] BANGARU ML, MENG J, KAISER DJ, et al. Differential expression of CaMKII isoforms and overall kinase activity in rat dorsal root ganglia after injury. Neuroscience. 2015;300:116-127.
[28] LIU Y, TEMPLETON DM. Initiation of caspase-independent death in mouse mesangial cells by Cd2+: involvement of p38 kinase and CaMK-II. J Cell Physiol. 2008;217(2):307-318.
[29] AFSHARI FT, KAPPAGANTULA S, FAWCETT JW. Extrinsic and intrinsic factors controlling axonal regeneration after spinal cord injury. Expert Rev Mol Med. 2009;11:e37.
[30] GIGER RJ, HOLLIS ER 2ND, TUSZYNSKI MH. Guidance molecules in axon regeneration. Cold Spring Harb Perspect Biol. 2010;2(7):a001867.
[31] FILBIN MT. Myelin-associated inhibitors of axonal regeneration in the adult mammalian CNS. Nat Rev Neurosci. 2003;4(9):703-713.
[32] CHANDRAN V, COPPOLA G, NAWABI H, et al. A Systems-Level Analysis of the Peripheral Nerve Intrinsic Axonal Growth Program. Neuron. 2016;89(5):956-970.
[33] DANILOV CA, STEWARD O. Conditional genetic deletion of PTEN after a spinal cord injury enhances regenerative growth of CST axons and motor function recovery in mice. Exp Neurol. 2015;266:147-160.
[34] ZUKOR K, BELIN S, WANG C, et al. Short hairpin RNA against PTEN enhances regenerative growth of corticospinal tract axons after spinal cord injury. J Neurosci. 2013;33(39):15350-15361.
[35] HUANG Z, HU Z, XIE P, et al. Tyrosine-mutated AAV2-mediated shRNA silencing of PTEN promotes axon regeneration of adult optic nerve. PLoS One. 2017;12(3):e0174096.
[36] EASLEY CA, FAISON MO, KIRSCH TL, et al. Laminin activates CaMK-II to stabilize nascent embryonic axons. Brain Res. 2006;1092(1):59-68.
[37] TANG F, KALIL K. Netrin-1 induces axon branching in developing cortical neurons by frequency-dependent calcium signaling pathways. J Neurosci. 2005;25(28):6702-6715.
[38] JOURDAIN P, FUKUNAGA K, MULLER D. Calcium/calmodulin-dependent protein kinase II contributes to activity-dependent filopodia growth and spine formation. J Neurosci. 2003;23(33):10645-10649.