Abstract
Promoting remyelination in multiple sclerosis is important to prevent axon degeneration, given the lack of curative treatment. Although some growth factors improve this repair, unspecific delivery to cells and potential side effects limit their therapeutic use. Thus, NFL-TBS.40-63 peptide (NFL)—known to enter specifically myelinating oligodendrocytes (OL)—was used to vectorize 100 nm diameter lipid nanoparticles (LNC), and the ability of NFL-LNC to specifically target OL from newborn rat brain was assessed in vitro. Specific uptake of DiD-labeled NFL-LNC by OL characterized by CNP and myelin basic protein was observed by confocal microscopy, as well as DiD colocalization with NFL and with Rab5—a marker of early endosomes. Unvectorized LNC did not significantly penetrate OL and there was no uptake of NFL-LNC by astrocytes. Canonical maturation of OL which extended compacted myelin-like membranes was observed by transmission electron microscopy in cells grown up to 9 days with NFL-LNC. Endocytosis of NFL-LNC appeared to depend on several pathways, as demonstrated by inhibitors. In addition, vectorized NFL-LNC adsorbed on neurotrophin-3 (NT-3) potentiated the proremyelinating effects of NT-3 after demyelination by lysophosphatidyl choline, allowing noticeably decreasing NT-3 concentration. Our results if they were confirmed in vivo suggest that NFL-vectorized LNC appear safe and could be considered as putative carriers for specific drug delivery to OL in order to increase remyelination.
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References
De Stefano N, Silva DG, Barnett MH (2017) Effect of fingolimod on brain volume loss in patients with multiple sclerosis. CNS Drugs 31:289–305
Goldman SA, Nedergaard M, Windrem MS (2012) Glial progenitor cell-based treatment and modeling of neurological disease. Science 338:491–495
Myers SA, Bankston AN, Burke DA, Ohri SS, Whittemore SR (2016) Does the preclinical evidence for functional remyelination following myelinating cell engraftment into the injured spinal cord support progression to clinical trials? Exp Neurol 283(Pt B):560–572
Jean I, Lavialle C, Barthelaix-Pouplard A, Fressinaud C (2003) Neurotrophin-3 specifically increases mature oligodendrocyte population and enhances remyelination after chemical demyelination of adult rat CNS. Brain Res 972:110–118
Fressinaud C (2005) Repeated injuries dramatically affect cells of the oligodendrocyte lineage: effects of PDGF and NT-3 in vitro. Glia 49:555–566
Huang Y, Dreyfus CF (2016) The role of growth factors as a therapeutic approach to demyelinating disease. Exp Neurol 283(Pt B):531–540
Bergles DE, Richardson WD (2015) Oligodendrocyte development and plasticity. Cold Spring Harb Perspect Biol 8(2):a020453. https://doi.org/10.1101/cshperspect.a020453
Pinezich MR, Russell LN, Murphy NP, Lampe KJ (2018) Encapsulated oligodendrocyte precursor cell fate is dependent on PDGF-AA release kinetics in a 3D microparticle-hydrogel drug delivery system. J Biomed Mater Res A 106A:2402–2411
Santhosh KT, Alizadeh A, Karimi-Abdolrezaee S (2017) Design and optimization of PLGA microparticles for controlled and local delivery of Neuregulin-1 in traumatic spinal cord injury. J Control Release 261:147–162
Rittchen S, Boyd A, Burns A, Park J, Fahmy TM, Metcalfe S, Williams A (2015) Myelin repair in vivo is increased by targeting oligodendrocyte precursor cells with nanoparticles encapsulating leukaemia inhibitory factor (LIF). Biomaterials 56:78–85
Jean I, Allamargot C, Barthelaix-Pouplard A, Fressinaud C (2002) Axonal lesions and PDGF-enhanced remyelination in the rat corpus callosum after lysolecithin demyelination. NeuroReport 13:627–631
Lu P, Jones LL, Tuszynski MH (2007) Axon regeneration through scars and into sites of chronic spinal cord injury. Exp Neurol 203:8–21
Lawn S, Krishna N, Pisklakova A, Qu X, Fenstermacher DA, Fournier M, Vrionis FD, Tran N, Chan JA, Kenchappa RS, Forsyth PA (2015) Neurotrophin signaling via TrkB and TrkC receptors promotes the growth of brain tumor-initiating cells. J Biol Chem 290:3814–3824
Kamermans A, Planting KE, Jalink K, van Horssen J, de Vries HE (2018) Reactive astrocytes in multiple sclerosis impair neuronal outgrowth through TRPM7-mediated chondroitin sulfate proteoglycan production. Glia. https://doi.org/10.1002/glia.23526
Nittoli V, Sepe RM, Coppola U, D'Agostino Y, De Felice E, Palladino A, Vassalli QA, Locascio A, Ristoratore F, Spagnuolo A, D'Aniello S, Sordino P (2018) A comprehensive analysis of neurotrophins and neurotrophin tyrosine kinase receptors expression during development of zebrafish. J Comp Neurol 526:1057–1072
Vrignaud S, Anton N, Gayet P, Benoit JP, Saulnier P (2011) Reverse micelle-loaded lipid nanocarriers: a novel drug delivery system for the sustained release of doxorubicin hydrochloride. Eur J Pharm Biopharm 79:197–204
Perrier T, Saulnier P, Fouchet F, Lautram N, Benoît JP (2010) Post-insertion into Lipid NanoCapsules (LNCs): From experimental aspects to mechanisms. Int J Pharm 396:204–209
Huynh NT, Passirani C, Saulnier P, Benoit JP (2009) Lipid nanocapsules: a new platform for nanomedicine. Lipid nanocapsules: a new platform for nanomedicine. Int J Pharm 379:201–209
Carradori D, Saulnier P, Préat V, Des Rieux A, Eyer J (2016) NFL-lipid nanocapsules for brain neural stem cell targeting in vitro and in vivo. J Control Release 238:253–262
Karim R, Lepeltier E, Esnault L, Pigeon P, Lemaire L, Lépinoux-Chambaud C, Clere N, Jaouen G, Eyer J, Piel G, Passirani C (2018) Enhanced and preferential internalization of lipid nanocapsules into human glioblastoma cells: effect of a surface-functionalizing NFL peptide. Nanoscale 10:13485–13501
Dupont E, Prochiantz A, Joliot A (2007) Identification of a signal peptide for unconventional secretion. J Biol Chem 282:8994–9000
Bocquet A, Berges R, Franck R, Robert P, Peterson AC, Eyer J (2009) Neurofilaments bind tubulin and modulate its polymerization. J Neurosci 29:11043–11054
Fressinaud C, Eyer J (2014) Neurofilament-tubulin binding site peptide NFL-TBS.40-63 increases the differentiation of oligodendrocytes in vitro and partially prevents them from lysophosphatidyl choline toxiciy. J Neurosci Res 92:243–253
Fressinaud C, Eyer J (2015) Neurofilaments and NFL-TBS.40-63 peptide penetrate oligodendrocytes through clathrin-dependent endocytosis to promote their growth and survival in vitro. Neuroscience 298:42–51
Umerska A, Mouzouvi CRA, Bigot A, Saulnier P (2015) Formulation and nebulization of fluticasone propionate-loaded lipid nanocarriers. Int J Pharm 493:224–232
Fressinaud C, Berges R, Eyer J (2012) Axon cytoskeleton proteins specifically modulate oligodendrocyte growth and differentiation in vitro. Neurochem Int 60:78–90
Fressinaud C, Vallat JM, Rigaud M, Cassagne C, Labourdette G, Sarliève LL (1990) Investigation of myelination in vitro: polar lipid content and fatty acid composition of myelinating oligodendrocytes in rat oligodendrocyte cultures. Neurochem Int 16:27–39
Kirchhausen T, Macia E, Pelish HE (2008) Use of dynasore, the small molecule inhibitor of dynamin, in the regulation of endocytosis. Methods Enzymol 438:77–93
Rodal SK, Skretting G, Garred O, Vilhardt F, van Deurs B, Sandvig K (1999) Extraction of cholesterol with methyl-beta-cyclodextrin perturbs formation of clathrin-coated endocytic vesicles. Mol Biol Cell 104:961–974
Pho MT, Ashok A, Atwood WJ (2000) JC virus enters human glial cells by clathrin-dependent receptor-mediated endocytosis. J Virol 74:2288–2292
Duchardt F, Fotin-Mleczek M, Schwarz H, Fischer R, Brock R (2007) A comprehensive model for the cellular uptake of cationic cell-penetrating peptides. Traffic 8:848–866
Barres BA, Raff MC, Gaese F, Bartke I, Dechant G, Barde YA (1994) A crucial role for neurotrophin-3 in oligodendrocyte development. Nature 367:371–375
Kubista M, Akerman B, Nordén B (1987) Characterization of interaction between DNA and 4',6-diamidino-2-phenylindole by optical spectroscopy. Biochemistry 26:4545–4553
Clemente R, de la Torre JC (2009) Cell entry of Borna disease virus follows a clathrin-mediated endocytosis pathway that requires Rab5 and microtubules. J Virol 83:10406–10416
Anton N, Saulnier P, Gaillard C, Porcher E, Vrignaud S, Benoit JP (2009) Aqueous-core lipid nanocapsules for encapsulating fragile hydrophilic and/or lipophilic molecules. Langmuir 25:11413–11419
Balzeau J, Pinier M, Berges R, Saulnier P, Benoit JP, Eyer J (2013) The effect of functionalizing lipid nanocapsules with NFL-TBS. 40-63 peptide on their uptake by glioblastoma cells. Biomaterials 34:3381–3389
Hellström AK, Bordes R (2019) Reversible flocculation of nanoparticles by a carbamate surfactant. J Colloid Interface Sci 536:722–727
Trapp BD, Peterson J, Ransohof RM, Rudick R, Mörk S, Bö L (1998) Axonal transection in the lesions of multiple sclerosis. N Engl J Med 338:278–285
Rudick RA, Fisher E, Lee JC, Simon J, Jacobs L (1999) Use of the brain parenchymal fraction to measure whole brain atrophy in relapsing-remittingMS. Multiple Sclerosis Collaborative Research Group. Neurology 53:1698–1704
Wegner C, Esiri MM, Chance SA, Palace J, Matthews PM (2006) Neocortical neuronal, synaptic, and glial loss in multiple sclerosis. Neurology 67:960–967
Irvine KA, Blakemore WF (2008) Remyelination protects axons from demyelination-associated axon degeneration. Brain 131:1464–1477
Naeimi R, Safarpour F, Hashemian M, Tashakorian H, Ahmadian SR, Ashrafpour M, Ghasemi-Kasman M (2018) Curcumin-loaded nanoparticles ameliorate glial activation and improve myelin repair in lyolecithin-induced focal demyelination model of rat corpus callosum. Neurosci Lett 674:1–610
Kfoury N, Holmes BB, Jiang H, Holtzman DM, Diamond MI (2012) Trans-cellular propagation of tau aggregation by fibrillar species. J Biol Chem 287:19440–19451
Lee HJ, Suk JE, Bae EJ, Lee JH, Paik SR, Lee SJ (2008) Assembly-dependent endocytosis and clearance of extracellular alphasynuclein. Int J Biochem Cell Biol 40:1835–1849
Frost B, Jacks RL, Diamond MI (2009) Propagation of tau misfolding from the outside to the inside of a cell. J Biol Chem 284:12845–12852
Ren PH, Lauckner JE, Kachirskaia I, Heuser JE, Melki R, Kopito RR (2009) Cytoplasmic penetration and persistent infection of mammalian cells by polyglutamine aggregates. Nat Cell Biol 11:219–225
Bastiat G, Pritz CO, Roider C, Fouchet F, Lignières E, Jesacher A, Glueckert R, Ritsch-Marte M, Schrott-Fischer A, Saulnier P, Benoit JP (2013) A new tool to ensure the fluorescent dye labeling stability of nanocarriers: a real challenge for fluorescence imaging. J Control Release 170:334–342
Jovic M, Sharma M, Rahajeng J, Caplan S (2010) The early endosome: a busy sorting station for proteins at the crossroads. Histol Histopathol 25:99–112
Duncan ID, Radcliff AB, Heidari M, Kidd G, August BK, Wierenga LA (2018) The adult oligodendrocyte can participate in remyelination. Proc Natl Acad Sci USA 115:E11807–E11816
Yeung MSY, Djelloul M, Steiner E, Bernard S, Salehpour M, Possnert G, Brundin L, Frisén J (2019) Dynamics of oligodendrocyte generation in multiple sclerosis. Nature 566:538–542
Simons M, Nave KA (2016) Oligodendrocytes: myelination and axonal support. Cold Spring Harb Perspect Biol 8:a020479
Berges R, Balzeau J, Peterson AC, Eyer J (2012) A tubulin binding peptide targets glioma cells disrupting their microtubules, blocking migration, and inducing apoptosis. Mol Ther 20:1367–1377
Berges R, Balzeau J, Takahashi M, Prevost C, Eyer J (2012) Structure-function analysis of the glioma targeting NFL-TBS.40-63 peptide corresponding to the tubulin-binding site on the light neurofilament subunit. PLoS ONE 7:e49436
Ammendrup-Johnsen I, Naito Y, Craig AM, Takahashi H (2015) Neurotrophin-3 enhances the synaptic organizing function of trkc-protein tyrosine phosphatase σ in rat hippocampal neurons. J Neurosci 35:12425–12431
Steenblock ER, Fadel T, Labowsky M, Pober JS, Fahmy TM (2011) An artificial antigen-presenting cell with paracrine delivery of IL-2 impacts the magnitude and direction of the T cell response. J Biol Chem 286:34883–34892
Swiecicki JM, Di Pisa M, Burlina F, Lécorché P, Mansuy C, Chassaing G, Lavielle S (2015) Accumulation of cell-penetrating peptides in large unilamellar vesicles: a straightforward screening assay for investigating the internalization mechanism. Biopolymers 104:533–543
Alves ID, Goasdoué N, Correia I, Aubry S, Galanth C, Sagan S, Lavielle S, Chassaing G (2008) Membrane interaction and perturbation mechanisms induced by two cationic cell penetrating peptides with distinct charge distribution. Biochim Biophys Acta 1780:948–959
Carradori D, Labrak Y, Miron VE, Saulnier P, Eyer J, Préat V, desRieux A (2020) Retinoic acid-loaded NFL-lipid nanocapsules promote oligodendrogenesis in focal white matter lesion. Biomaterials 230:119653
Béduneau A, Hindré F, Clavreul A, Leroux JC, Saulnier P, Benoit JP (2008) Brain targeting using novel lipid nanovectors. J Control Release 126:44–49
Inês Teixeira M, Lopes CM, Helena Amaral M, Costa PC (2020) Current insights on lipid nanocarrier-assisted drug delivery in the treatment of neurodegenerative diseases. Eur J Pharm Biopharm S0939–6411(20):30015–30021. https://doi.org/10.1016/j.ejpb.2020.01.005
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The authors thank Dr R. Perrot for expert technical assistance with confocal microscopy, and Mrs F. Manero for electron microscopy techniques.
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Designed study: CF. Performed experiments: CF, OT, AMU. Analyzed data: CF. Wrote paper: CF, PS.
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11064_2020_3122_MOESM1_ESM.tif
Supplementary file1 (TIF 22200 kb). Supplementary Fig. 1 Transmission electron microscopy of 100 nm LNC adsorbed on NFL Scramble peptide (NFL-SCR) - composed of the same aminoacids as NFL-TBS peptide although in random order -. This peptide is inactive on oligodendrocytes. Note the filament bundles formed by NFL-SCR. Pure LNC were diluted 1/10 (v/v); NFL-SCR 0.027 µM final concentration (same concentration as NFL-TBS, see Fig. 1). Uranyl acetate negative stain. Scale bar 200 nm as indicated
11064_2020_3122_MOESM2_ESM.tif
Supplementary file2 (TIF 40345 kb). Supplementary Fig. 2 Long term treatment (9 days) with NFL-LNC-DiD does not alter OL characteristics. TEM analyses of control (left) and NFL-LNC treated cultures (right) after 9 days demonstrate numerous microtubules in OL processes. Scale bars = 200 nm
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Fressinaud, C., Thomas, O., Umerska, A.M. et al. Lipid Nanoparticles Vectorized with NFL-TBS.40-63 Peptide Target Oligodendrocytes and Promote Neurotrophin-3 Effects After Demyelination In Vitro. Neurochem Res 45, 2732–2748 (2020). https://doi.org/10.1007/s11064-020-03122-y
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DOI: https://doi.org/10.1007/s11064-020-03122-y