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Microcurvature landscapes induce neural stem cell polarity and enhance neural differentiation

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Abstract

Tissue curvature has long been recognized as an important anatomical parameter that affects intracellular behaviors, and there is emerging interest in applying cell-scale curvature as a designer property to drive cell fates for tissue engineering purposes. Although neural cells are known to undergo dramatic and terminal morphological changes during development and curvature-limiting behaviors have been demonstrated in neurite outgrowth studies, there are still crucial gaps in understanding neural cell behaviors, particularly in the context of a three-dimensional (3D) curvature landscape similar to an actual tissue engineering scaffold. In this study, we fabricated two substrates of microcurvature (curvature-substrates) that present a smooth and repeating landscape with focuses of either a concave or a convex pattern. Using these curvature-substrates, we studied the properties of morphological differentiation in N2a neuroblastoma cells. In contrast to other studies where two-dimensional (2D) curvature was demonstrated to limit neurite outgrowth, we found that both the concave and convex substrates acted as continuous and uniform mechanical protrusions that significantly enhanced neural polarity and differentiation with few morphological changes in the main cell body. This enhanced differentiation was manifested in various properties, including increased neurite length, increased nuclear displacement, and upregulation of various neural markers. By demonstrating how the micron-scale curvature landscape induces neuronal polarity, we provide further insights into the design of biomaterials utilizing the influence of surface curvature in neural tissue engineering.

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References

  1. Dunn GA, Heath JP (1976) A new hypothesis of contact guidance in tissue cells. Exp Cell Res 101(1):1–14. https://doi.org/10.1016/0014-4827(76)90405-5

    Article  Google Scholar 

  2. Curtis ASG, Varde M (1964) Control of cell behavior: topological factors. J Natl Cancer Inst 33(1):15–26. https://doi.org/10.1093/jnci/33.1.15

    Article  Google Scholar 

  3. Baptista D, Teixeira L, van Blitterswijk C et al (2019) Overlooked? Underestimated? Effects of substrate curvature on cell behavior. Trends Biotechnol 37(8):838–854. https://doi.org/10.1016/j.tibtech.2019.01.006

    Article  Google Scholar 

  4. Pelham RJ Jr, Wang YL (1997) Cell locomotion and focal adhesions are regulated by substrate flexibility. Proc Natl Acad Sci USA 94(25):13661–13665. https://doi.org/10.1073/pnas.94.25.13661

    Article  Google Scholar 

  5. Ye K, Wang X, Cao L et al (2015) Matrix stiffness and nanoscale spatial organization of cell-adhesive ligands direct stem cell fate. Nano Lett 15(7):4720–4729. https://doi.org/10.1021/acs.nanolett.5b01619

    Article  Google Scholar 

  6. Cavalcanti-Adam EA, Volberg T, Micoulet A et al (2007) Cell spreading and focal adhesion dynamics are regulated by spacing of integrin ligands. Biophys J 92(8):2964–2974. https://doi.org/10.1529/biophysj.106.089730

    Article  Google Scholar 

  7. Boyan BD, Hummert TW, Dean DD et al (1996) Role of material surfaces in regulating bone and cartilage cell response. Biomaterials 17(2):137–146. https://doi.org/10.1016/0142-9612(96)85758-9

    Article  Google Scholar 

  8. Yamashita T, Kollmannsberger P, Mawatari K et al (2016) Cell sheet mechanics: how geometrical constraints induce the detachment of cell sheets from concave surfaces. Acta Biomater 45:85–97. https://doi.org/10.1016/j.actbio.2016.08.044

    Article  Google Scholar 

  9. Alias MA, Buenzli PR (2017) Modeling the effect of curvature on the collective behavior of cells growing new tissue. Biophys J 112(1):193–204. https://doi.org/10.1016/j.bpj.2016.11.3203

    Article  Google Scholar 

  10. Khan H, Beck C, Kunze A (2021) Multi-curvature micropatterns unveil distinct calcium and mitochondrial dynamics in neuronal networks. Lab Chip 21(6):1164–1174. https://doi.org/10.1039/d0lc01205j

    Article  Google Scholar 

  11. Hilgetag CC, Barbas H (2006) Role of mechanical factors in the morphology of the primate cerebral cortex. PLoS Comput Biol 2(3):e22. https://doi.org/10.1371/journal.pcbi.0020022

  12. Del Toro D, Ruff T, Cederfjäll E et al (2017) Regulation of cerebral cortex folding by controlling neuronal migration via FLRT adhesion molecules. Cell 169(4):621-635.e16. https://doi.org/10.1016/j.cell.2017.04.012

    Article  Google Scholar 

  13. Roth S, Bisbal M, Brocard J et al (2012) How morphological constraints affect axonal polarity in mouse neurons. PLoS ONE 7(3):e33623. https://doi.org/10.1371/journal.pone.0033623

  14. Smeal RM, Rabbitt R, Biran R et al (2005) Substrate curvature influences the direction of nerve outgrowth. Ann Biomed Eng 33(3):376–382. https://doi.org/10.1007/s10439-005-1740-z

    Article  Google Scholar 

  15. Smeal RM, Tresco PA (2008) The influence of substrate curvature on neurite outgrowth is cell type dependent. Exp Neurol 213(2):281–292. https://doi.org/10.1016/j.expneurol.2008.05.026

    Article  Google Scholar 

  16. Werner M, Blanquer SB, Haimi SP et al (2016) Surface curvature differentially regulates stem cell migration and differentiation via altered attachment morphology and nuclear deformation. Adv Sci 4(2):1600347. https://doi.org/10.1002/advs.201600347

    Article  Google Scholar 

  17. Pieuchot L, Marteau J, Guignandon A et al (2018) Curvotaxis directs cell migration through cell-scale curvature landscapes. Nat Commun 9(1):3995. https://doi.org/10.1038/s41467-018-06494-6

    Article  Google Scholar 

  18. Jin ZY, Zhai YS, Zhou Y et al (2022) Regulation of mesenchymal stem cell osteogenic potential via microfluidic manipulation of microcarrier surface curvature. Chem Eng J 448:137739. https://doi.org/10.1016/j.cej.2022.137739

  19. Yang Y, Xu T, Bei HP et al (2022) Gaussian curvature-driven direction of cell fate toward osteogenesis with triply periodic minimal surface scaffolds. Proc Natl Acad Sci USA 119(41):e2206684119. https://doi.org/10.1073/pnas.2206684119

  20. Moe AA, Suryana M, Marcy G et al (2012) Microarray with micro- and nano-topographies enables identification of the optimal topography for directing the differentiation of primary murine neural progenitor cells. Small 8(19):3050–3061. https://doi.org/10.1002/smll.201200490

    Article  Google Scholar 

  21. Conover JC, Notti RQ (2008) The neural stem cell niche. Cell Tissue Res 331(1):211–224. https://doi.org/10.1007/s00441-007-0503-6

    Article  Google Scholar 

  22. Gu X, Ding F, Williams DF (2014) Neural tissue engineering options for peripheral nerve regeneration. Biomaterials 35(24):6143–6156. https://doi.org/10.1016/j.biomaterials.2014.04.064

    Article  Google Scholar 

  23. Boni R, Ali A, Shavandi A et al (2018) Current and novel polymeric biomaterials for neural tissue engineering. J Biomed Sci 25(1):90. https://doi.org/10.1186/s12929-018-0491-8

    Article  Google Scholar 

  24. Khademhosseini A, Langer R, Borenstein J et al (2006) Microscale technologies for tissue engineering and biology. Proc Natl Acad Sci USA 103(8):2480–2487. https://doi.org/10.1073/pnas.0507681102

    Article  Google Scholar 

  25. Virtanen P, Gommers R, Oliphant TE et al (2020) SciPy 1.0: fundamental algorithms for scientific computing in Python. Nat Methods 17(3):261–272. https://doi.org/10.1038/s41592-019-0686-2

    Article  Google Scholar 

  26. Wu G, Fang Y, Lu ZH et al (1998) Induction of axon-like and dendrite-like processes in neuroblastoma cells. J Neurocytol 27(1):1–14. https://doi.org/10.1023/a:1006910001869

    Article  Google Scholar 

  27. Shea TB, Fischer I, Sapirstein VS (1985) Effect of retinoic acid on growth and morphological differentiation of mouse NB2a neuroblastoma cells in culture. Brain Res 353(2):307–314. https://doi.org/10.1016/0165-3806(85)90220-2

    Article  Google Scholar 

  28. Stringer C, Wang T, Michaelos M et al (2021) Cellpose: a generalist algorithm for cellular segmentation. Nat Methods 18(1):100–106. https://doi.org/10.1038/s41592-020-01018-x

    Article  Google Scholar 

  29. Ho SY, Chao CY, Huang HL et al (2011) NeurphologyJ: an automatic neuronal morphology quantification method and its application in pharmacological discovery. BMC Bioinform 12:230. https://doi.org/10.1186/1471-2105-12-230

    Article  Google Scholar 

  30. Zhang TY, Suen CY (1984) A fast parallel algorithm for thinning digital patterns. Commun ACM 27(3):236–239. https://doi.org/10.1145/357994.358023

    Article  Google Scholar 

  31. Sun J, Wang D, Guo L et al (2017) Androgen receptor regulates the growth of neuroblastoma cells in vitro and in vivo. Front Neurosci 11:116. https://doi.org/10.3389/fnins.2017.00116

    Article  Google Scholar 

  32. Tremblay RG, Sikorska M, Sandhu JK et al (2010) Differentiation of mouse Neuro 2A cells into dopamine neurons. J Neurosci Methods 186(1):60–67. https://doi.org/10.1016/j.jneumeth.2009.11.004

    Article  Google Scholar 

  33. Marzinke MA, Clagett-Dame M (2012) The all-trans retinoic acid (atRA)-regulated gene Calmin (Clmn) regulates cell cycle exit and neurite outgrowth in murine neuroblastoma (Neuro2a) cells. Exp Cell Res 318(1):85–93. https://doi.org/10.1016/j.yexcr.2011.10.002

    Article  Google Scholar 

  34. Su X, Gu X, Zhang Z et al (2020) Retinoic acid receptor gamma is targeted by microRNA-124 and inhibits neurite outgrowth. Neuropharmacology 163:107657. https://doi.org/10.1016/j.neuropharm.2019.05.034

  35. Vining KH, Mooney DJ (2017) Mechanical forces direct stem cell behaviour in development and regeneration. Nat Rev Mol Cell Biol 18(12):728–742. https://doi.org/10.1038/nrm.2017.108

    Article  Google Scholar 

  36. Tojkander S, Gateva G, Lappalainen P (2012) Actin stress fibers—assembly, dynamics and biological roles. J Cell Sci 125(Pt 8):1855–1864. https://doi.org/10.1242/jcs.098087

    Article  Google Scholar 

  37. Olson EN, Nordheim A (2010) Linking actin dynamics and gene transcription to drive cellular motile functions. Nat Rev Mol Cell Biol 11(5):353–365. https://doi.org/10.1038/nrm2890

    Article  Google Scholar 

  38. Kalukula Y, Stephens AD, Lammerding J et al (2022) Mechanics and functional consequences of nuclear deformations. Nat Rev Mol Cell Biol 23(9):583–602. https://doi.org/10.1038/s41580-022-00480-z

    Article  Google Scholar 

  39. Panciera T, Azzolin L, Cordenonsi M et al (2017) Mechanobiology of YAP and TAZ in physiology and disease. Nat Rev Mol Cell Biol 18(12):758–770. https://doi.org/10.1038/nrm.2017.87

    Article  Google Scholar 

  40. Dupont S, Morsut L, Aragona M et al (2011) Role of YAP/TAZ in mechanotransduction. Nature 474(7350):179–183. https://doi.org/10.1038/nature10137

    Article  Google Scholar 

  41. Zhang H, Deo M, Thompson RC et al (2012) Negative regulation of Yap during neuronal differentiation. Dev Biol 361(1):103–115. https://doi.org/10.1016/j.ydbio.2011.10.017

    Article  Google Scholar 

  42. Sun Y, Yong KM, Villa-Diaz LG et al (2014) Hippo/YAP-mediated rigidity-dependent motor neuron differentiation of human pluripotent stem cells. Nat Mater 13(6):599–604. https://doi.org/10.1038/nmat3945

    Article  Google Scholar 

  43. Lin YT, Ding JY, Li MY et al (2012) YAP regulates neuronal differentiation through Sonic hedgehog signaling pathway. Exp Cell Res 318(15):1877–1888. https://doi.org/10.1016/j.yexcr.2012.05.005

    Article  Google Scholar 

  44. Yamada KM, Sixt M (2019) Mechanisms of 3D cell migration. Nat Rev Mol Cell Biol 20(12):738–752. https://doi.org/10.1038/s41580-019-0172-9

    Article  Google Scholar 

  45. Gundersen GG, Worman HJ (2013) Nuclear positioning. Cell 152(6):1376–1389. https://doi.org/10.1016/j.cell.2013.02.031

    Article  Google Scholar 

  46. Davidson PM, Cadot B (2021) Actin on and around the Nucleus. Trends Cell Biol 31(3):211–223. https://doi.org/10.1016/j.tcb.2020.11.009

    Article  Google Scholar 

  47. Cáceres A, Ye B, Dotti CG (2012) Neuronal polarity: demarcation, growth and commitment. Curr Opin Cell Biol 24(4):547–553. https://doi.org/10.1016/j.ceb.2012.05.011

    Article  Google Scholar 

  48. Meiring JCM, Shneyer BI, Akhmanova A (2020) Generation and regulation of microtubule network asymmetry to drive cell polarity. Curr Opin Cell Biol 62:86–95. https://doi.org/10.1016/j.ceb.2019.10.004

    Article  Google Scholar 

  49. Lee Y, McIntire LV, Zygourakis K (1994) Analysis of endothelial cell locomotion: differential effects of motility and contact inhibition. Biotechnol Bioeng 43(7):622–634. https://doi.org/10.1002/bit.260430712

    Article  Google Scholar 

  50. Su J, Zapata PJ, Chen CC et al (2009) Local cell metrics: a novel method for analysis of cell-cell interactions. BMC Bioinform 10:350. https://doi.org/10.1186/1471-2105-10-350

    Article  Google Scholar 

  51. Moore R, Theveneau E, Pozzi S et al (2013) Par3 controls neural crest migration by promoting microtubule catastrophe during contact inhibition of locomotion. Development 140(23):4763–4775. https://doi.org/10.1242/dev.098509

    Article  Google Scholar 

  52. Dimou L, Götz M (2014) Glial cells as progenitors and stem cells: new roles in the healthy and diseased brain. Physiol Rev 94(3):709–737. https://doi.org/10.1152/physrev.00036.2013

    Article  Google Scholar 

  53. Chan KY, Baxter CF (1979) Compartments of tubulin and tubulin-like proteins in differentiating neubroblastoma cells. Brain Res 174(1):135–152. https://doi.org/10.1016/0006-8993(79)90809-6

    Article  Google Scholar 

  54. Katsetos CD, Karkavelas G, Herman MM et al (1998) Class III beta-tubulin isotype (beta III) in the adrenal medulla: I. localization in the developing human adrenal medulla. Anat Rec 250(3):335–343

    Article  Google Scholar 

  55. Wu PY, Lin YC, Chang CL et al (2009) Functional decreases in P2X7 receptors are associated with retinoic acid-induced neuronal differentiation of Neuro-2a neuroblastoma cells. Cell Signal 21(6):881–891. https://doi.org/10.1016/j.cellsig.2009.01.036

    Article  Google Scholar 

  56. Jeon WB, Park BH, Choi SK et al (2012) Functional enhancement of neuronal cell behaviors and differentiation by elastin-mimetic recombinant protein presenting Arg-Gly-Asp peptides. BMC Biotechnol 12:61. https://doi.org/10.1186/1472-6750-12-61

    Article  Google Scholar 

  57. Choi SK, Kim JH, Park JK et al (2013) Cytotoxicity and inhibition of intercellular interaction in N2a neurospheroids by perfluorooctanoic acid and perfluorooctanesulfonic acid. Food Chem Toxicol 60:520–529. https://doi.org/10.1016/j.fct.2013.07.070

    Article  Google Scholar 

  58. Dehmelt L, Halpain S (2004) Actin and microtubules in neurite initiation: are MAPs the missing link? J Neurobiol 58(1):18–33. https://doi.org/10.1002/neu.10284

    Article  Google Scholar 

  59. Winans AM, Collins SR, Meyer T (2016) Waves of actin and microtubule polymerization drive microtubule-based transport and neurite growth before single axon formation. eLife 5:e12387. https://doi.org/10.7554/eLife.12387

  60. Konietzny A, Bär J, Mikhaylova M (2017) Dendritic actin cytoskeleton: structure, functions, and regulations. Front Cell Neurosci 11:147. https://doi.org/10.3389/fncel.2017.00147

    Article  Google Scholar 

  61. Liu W, Sun Q, Zheng ZL et al (2022) Topographic cues guiding cell polarization via distinct cellular mechanosensing pathways. Small 18(2):e2104328. https://doi.org/10.1002/smll.202104328

  62. Liu Y, Yang Q, Wang Y et al (2022) Metallic scaffold with micron-scale geometrical cues promotes osteogenesis and angiogenesis via the ROCK/Myosin/YAP pathway. ACS Biomater Sci Eng 8(8):3498–3514. https://doi.org/10.1021/acsbiomaterials.2c00225

    Article  Google Scholar 

  63. Yogev S, Shen K (2017) Establishing neuronal polarity with environmental and intrinsic mechanisms. Neuron 96(3):638–650. https://doi.org/10.1016/j.neuron.2017.10.021

    Article  Google Scholar 

  64. Ferrari A, Cecchini M, Dhawan A et al (2011) Nanotopographic control of neuronal polarity. Nano Lett 11(2):505–511. https://doi.org/10.1021/nl103349s

    Article  Google Scholar 

  65. Higginbotham HR, Gleeson JG (2007) The centrosome in neuronal development. Trends Neurosci 30(6):276–283. https://doi.org/10.1016/j.tins.2007.04.001

    Article  Google Scholar 

  66. Elric J, Etienne-Manneville S (2014) Centrosome positioning in polarized cells: common themes and variations. Exp Cell Res 328(2):240–248. https://doi.org/10.1016/j.yexcr.2014.09.004

    Article  Google Scholar 

  67. Holcomb PS, Deerinck TJ, Ellisman MH et al (2013) Construction of a polarized neuron. J Physiol 591(13):3145–3150. https://doi.org/10.1113/jphysiol.2012.248542

    Article  Google Scholar 

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Acknowledgements

This work was supported by the Inter-Departmental Open Project of State Key Laboratory in Ultra-Precision Machining Technology (SKL-UPMT, No. P0033576).

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Authors and Affiliations

  1. Department of Biomedical Engineering, The Hong Kong Polytechnic University, Hung Hom, Kowloon, Hong Kong SAR, China

    Ho-Yin Yuen & Xin Zhao

  2. State Key Laboratory of Ultra-Precision Machining Technology, Department of Industrial and Systems Engineering, The Hong Kong Polytechnic University, Hung Hom, Kowloon, Hong Kong SAR, China

    Wai-Sze Yip & Suet To

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HYY was involved in the investigation, methodology, formal analysis, visualization, writing—original draft, and writing—review and editing. WSY was involved in writing—review and editing. ST was involved in funding acquisition, and writing—review and editing. XZ was involved in conceptualization, funding acquisition, supervision, and writing—review and editing.

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Correspondence to Suet To or Xin Zhao.

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XZ is an Associate Editor of Bio-Design and Manufacturing. The authors declare that they have no conflict of interest.

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Yuen, HY., Yip, WS., To, S. et al. Microcurvature landscapes induce neural stem cell polarity and enhance neural differentiation. Bio-des. Manuf. 6, 522–535 (2023). https://doi.org/10.1007/s42242-023-00243-5

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