A neural extracellular matrix-based method for in vitro hippocampal neuron culture and dopaminergic differentiation of neural stem cells

BMC Neuroscience

Ander Izeta⁵  Adolfo López de Munain³  Antonio G García⁴  José Iñaki Álava¹  Neia Naldaiz-Gastesi³  Fabio Cavaliere²  Marcos Maroto⁴  Patricia García-Parra³ 

[1] Basque Culinary Center R&D, San Sebastian, 20009, Spain;Department of Neuroscience and CIBERNED, University of Basque Country (UPV/EHU), Zamudio, 48170, Spain;Neuroscience Area and CIBERNED, Instituto Biodonostia, Hospital Universitario Donostia, San Sebastian, 20014, Spain;Instituto Teófilo Hernando de I + D del Medicamento. Departamento de Farmacología y Terapeútica and Servicio de Farmacología Clínica del IIS Hospital Universitario de La Princesa, Facultad de Medicina, Universidad Autónoma de Madrid (UAM), Madrid, 28029, Spain;Tissue Engineering Laboratory, Department of Bioengineering, Instituto Biodonostia, Hospital Universitario Donostia, San Sebastian, 20014, Spain

关键词: Dopaminergic differentiation;    Neural progenitor cells;    Neuronal culture;    Subventricular zone;    Neural extracellular matrix;   

Others  :  1140308
DOI  :  10.1186/1471-2202-14-48

 received in 2013-01-08, accepted in 2013-04-13,  发布年份 2013

【 摘 要 】

Background

The ability to recreate an optimal cellular microenvironment is critical to understand neuronal behavior and functionality in vitro. An organized neural extracellular matrix (nECM) promotes neural cell adhesion, proliferation and differentiation. Here, we expanded previous observations on the ability of nECM to support in vitro neuronal differentiation, with the following goals: (i) to recreate complex neuronal networks of embryonic rat hippocampal cells, and (ii) to achieve improved levels of dopaminergic differentiation of subventricular zone (SVZ) neural progenitor cells.

Methods

Hippocampal cells from E18 rat embryos were seeded on PLL- and nECM-coated substrates. Neurosphere cultures were prepared from the SVZ of P4-P7 rat pups, and differentiation of neurospheres assayed on PLL- and nECM-coated substrates.

Results

When seeded on nECM-coated substrates, both hippocampal cells and SVZ progenitor cells showed neural expression patterns that were similar to their poly-L-lysine-seeded counterparts. However, nECM-based cultures of both hippocampal neurons and SVZ progenitor cells could be maintained for longer times as compared to poly-L-lysine-based cultures. As a result, nECM-based cultures gave rise to a more branched neurite arborization of hippocampal neurons. Interestingly, the prolonged differentiation time of SVZ progenitor cells in nECM allowed us to obtain a purer population of dopaminergic neurons.

Conclusions

We conclude that nECM-based coating is an efficient substrate to culture neural cells at different stages of differentiation. In addition, neural ECM-coated substrates increased neuronal survival and neuronal differentiation efficiency as compared to cationic polymers such as poly-L-lysine.

【 授权许可】

   
2013 García-Parra et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Corey JM, Feldman EL: Substrate patterning: an emerging technology for the study of neuronal behavior. Exp Neurol 2003, 184(Suppl 1):S89-S96.
• [2]Mina M, Kollar EJ, Bishop JA, Rohrbach DH: Interaction between the neural crest and extracellular matrix proteins in craniofacial skeletogenesis. Crit Rev Oral Biol Med 1990, 1(2):79-87.
• [3]Perris R, Perissinotto D: Role of the extracellular matrix during neural crest cell migration. Mech Dev 2000, 95(1–2):3-21.
• [4]Celio MR, Spreafico R, De Biasi S, Vitellaro-Zuccarello L: Perineuronal nets: past and present. Trends Neurosci 1998, 21(12):510-515.
• [5]Giamanco KA, Morawski M, Matthews RT: Perineuronal net formation and structure in aggrecan knockout mice. Neuroscience 2010, 170(4):1314-1327.
• [6]Pizzorusso T, Medini P, Berardi N, Chierzi S, Fawcett JW, Maffei L: Reactivation of ocular dominance plasticity in the adult visual cortex. Science 2002, 298(5596):1248-1251.
• [7]Morawski M, Bruckner MK, Riederer P, Bruckner G, Arendt T: Perineuronal nets potentially protect against oxidative stress. Exp Neurol 2004, 188(2):309-315.
• [8]Galvin KA, Jones DG: Adult human neural stem cells for autologous cell replacement therapies for neurodegenerative disorders. NeuroRehabilitation 2006, 21(3):255-265.
• [9]Quinones-Hinojosa A, Chaichana K: The human subventricular zone: a source of new cells and a potential source of brain tumors. Exp Neurol 2007, 205(2):313-324.
• [10]Reynolds BA, Weiss S: Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science 1992, 255(5052):1707-1710.
• [11]Gage FH: Mammalian neural stem cells. Science 2000, 287(5457):1433-1438.
• [12]Palmer TD, Willhoite AR, Gage FH: Vascular niche for adult hippocampal neurogenesis. J Comp Neurol 2000, 425(4):479-494.
• [13]Temple S: The development of neural stem cells. Nature 2001, 414(6859):112-117.
• [14]Garcia-Parra P, Cavaliere F, Maroto M, Bilbao L, Obieta I, Alava JI, Izeta A, Lopez de Munain A: Modeling neural differentiation on micropatterned substrates coated with neural matrix components. Front Cell Neurosci 2012, 6:10.
• [15]Galtrey CM, Fawcett JW: The role of chondroitin sulfate proteoglycans in regeneration and plasticity in the central nervous system. Brain Res Rev 2007, 54(1):1-18.
• [16]Zuo J, Neubauer D, Dyess K, Ferguson TA, Muir D: Degradation of chondroitin sulfate proteoglycan enhances the neurite-promoting potential of spinal cord tissue. Exp Neurol 1998, 154(2):654-662.
• [17]Kawashima H, Atarashi K, Hirose M, Hirose J, Yamada S, Sugahara K, Miyasaka M: Oversulfated chondroitin/dermatan sulfates containing GlcAbeta1/IdoAalpha1-3GalNAc(4,6-O-disulfate) interact with L- and P-selectin and chemokines. J Biol Chem 2002, 277(15):12921-12930.
• [18]Margolis RU, Margolis RK: Chondroitin sulfate proteoglycans as mediators of axon growth and pathfinding. Cell Tissue Res 1997, 290(2):343-348.
• [19]O’Donnell M, Chance RK, Bashaw GJ: Axon growth and guidance: receptor regulation and signal transduction. Annu Rev Neurosci 2009, 32:383-412.
• [20]Ravichandran R, Sundarrajan S, Venugopal JR, Mukherjee S, Ramakrishna S: Advances in Polymeric Systems for Tissue Engineering and Biomedical Applications. Macromol Biosci 2012, 12(3):286-311.
• [21]Torres-Costa V, Martinez-Munoz G, Sanchez-Vaquero V, Munoz-Noval A, Gonzalez-Mendez L, Punzon-Quijorna E, Gallach-Perez D, Manso-Silvan M, Climent-Font A, Garcia-Ruiz JP: Engineering of silicon surfaces at the micro- and nanoscales for cell adhesion and migration control. Int J Nanomedicine 2012, 7:623-630.
• [22]Discher DE, Janmey P, Wang YL: Tissue cells feel and respond to the stiffness of their substrate. Science 2005, 310(5751):1139-1143.
• [23]Martinez-Ramos C, Lainez S, Sancho F, Garcia Esparza MA, Planells-Cases R, Garcia Verdugo JM, Gomez Ribelles JL, Salmeron Sanchez M, Monleon Pradas M, Barcia JA: Differentiation of postnatal neural stem cells into glia and functional neurons on laminin-coated polymeric substrates. Tissue Eng Part A 2008, 14(8):1365-1375.
• [24]Xiong Y, Zeng YS, Zeng CG, Du BL, He LM, Quan DP, Zhang W, Wang JM, Wu JL, Li Y: Synaptic transmission of neural stem cells seeded in 3-dimensional PLGA scaffolds. Biomaterials 2009, 30(22):3711-3722.
• [25]Ma W, Tavakoli T, Derby E, Serebryakova Y, Rao MS, Mattson MP: Cell-extracellular matrix interactions regulate neural differentiation of human embryonic stem cells. BMC Dev Biol 2008, 8:90. BioMed Central Full Text
• [26]Tang X, Jang SW, Okada M, Chan CB, Feng Y, Liu Y, Luo SW, Hong Y, Rama N, Xiong WC: Netrin-1 mediates neuronal survival through PIKE-L interaction with the dependence receptor UNC5B. Nat Cell Biol 2008, 10(6):698-706.
• [27]Rhodes KE, Fawcett JW: Chondroitin sulphate proteoglycans: preventing plasticity or protecting the CNS? J Anat 2004, 204(1):33-48.
• [28]Busch SA, Silver J: The role of extracellular matrix in CNS regeneration. Curr Opin Neurobiol 2007, 17(1):120-127.
• [29]Crespo-Santiago D: The extracellular matrix of the central nervous system: chondroitin sulphate type proteoglycans and neural repair. Rev Neurol 2004, 38(9):843-851.
• [30]Yamaguchi Y: Lecticans: organizers of the brain extracellular matrix. Cell Mol Life Sci 2000, 57(2):276-289.
• [31]Martin-de-Saavedra MD, del Barrio L, Canas N, Egea J, Lorrio S, Montell E, Verges J, Garcia AG, Lopez MG: Chondroitin sulfate reduces cell death of rat hippocampal slices subjected to oxygen and glucose deprivation by inhibiting p38, NFkappaB and iNOS. Neurochem Int 2011, 58(6):676-683.
• [32]Wang H, Katagiri Y, McCann TE, Unsworth E, Goldsmith P, Yu ZX, Tan F, Santiago L, Mills EM, Wang Y: Chondroitin-4-sulfation negatively regulates axonal guidance and growth. J Cell Sci 2008, 121(Pt 18):3083-3091.
• [33]Canas N, Valero T, Villarroya M, Montell E, Verges J, Garcia AG, Lopez MG: Chondroitin sulfate protects SH-SY5Y cells from oxidative stress by inducing heme oxygenase-1 via phosphatidylinositol 3-kinase/Akt. J Pharmacol Exp Ther 2007, 323(3):946-953.
• [34]McKeon RJ, Jurynec MJ, Buck CR: The chondroitin sulfate proteoglycans neurocan and phosphacan are expressed by reactive astrocytes in the chronic CNS glial scar. J Neurosci 1999, 19(24):10778-10788.
• [35]Bradbury EJ, Carter LM: Manipulating the glial scar: chondroitinase ABC as a therapy for spinal cord injury. Brain Res Bull 2011, 84(4–5):306-316.
• [36]Bunge MB: Novel combination strategies to repair the injured mammalian spinal cord. J Spinal Cord Med 2008, 31(3):262-269.
• [37]Carter LM, Starkey ML, Akrimi SF, Davies M, McMahon SB, Bradbury EJ: The yellow fluorescent protein (YFP-H) mouse reveals neuroprotection as a novel mechanism underlying chondroitinase ABC-mediated repair after spinal cord injury. J Neurosci 2008, 28(52):14107-14120.
• [38]Cavaliere F, Urra O, Alberdi E, Matute C: Oligodendrocyte differentiation from adult multipotent stem cells is modulated by glutamate. Cell Death Dis 2012, 3:e268.
• [39]Fuccillo M, Joyner AL, Fishell G: Morphogen to mitogen: the multiple roles of hedgehog signalling in vertebrate neural development. Nat Rev Neurosci 2006, 7(10):772-783.
• [40]Machold R, Hayashi S, Rutlin M, Muzumdar MD, Nery S, Corbin JG, Gritli-Linde A, Dellovade T, Porter JA, Rubin LL: Sonic hedgehog is required for progenitor cell maintenance in telencephalic stem cell niches. Neuron 2003, 39(6):937-950.
• [41]Palma V, Dahmane N, Ruiz i Altaba A: Hedgehog-Gli signalling and the growth of the brain. Nat Rev Neurosci 2002, 3(1):24-33.
• [42]Vitalis T, Cases O, Parnavelas JG: Development of the dopaminergic neurons in the rodent brainstem. Exp Neurol 2005, 191(Suppl 1):S104-S112.
• [43]Ye W, Shimamura K, Rubenstein JL, Hynes MA, Rosenthal A: FGF and Shh signals control dopaminergic and serotonergic cell fate in the anterior neural plate. Cell 1998, 93(5):755-766.
• [44]Denizot F, Lang R: Rapid colorimetric assay for cell growth and survival. Modifications to the tetrazolium dye procedure giving improved sensitivity and reliability. J Immunol Methods 1986, 89(2):271-277.