Research Article
Non-tumor mast cells cultured in vitro on a honeycomb-like structured film proliferate with multinucleated formation

https://doi.org/10.1016/j.nano.2013.08.011Get rights and content

Abstract

Mast cells are released from bone marrow into the circulatory system as immature precursors and differentiate upon their arrival at diverse organs and tissues. Because mast cell functions can be altered in these tissues, we propose that mast cells are sensitive to their surrounding microenvironment. To examine the morphological responses of mast cells, we cultured a proliferative mouse non-tumor cell line of mast cells (NCL-2 cells) on a honeycomb-like structured polystyrene film (HCF) representing a microenvironmental scaffold. In this study, the NCL-2 cells cultured on the HCF proliferated without apoptosis. Furthermore, NCL-2 cells cultured on 3- and 5-μm HCFs exhibited multinuclear formation. These observations of different NCL-2 cell morphologies and proliferation rates on HCF scaffolds with different hole sizes suggest that mast cells undertake specific proliferative shapes depending on the surrounding microenviroment. Moreover, HCFs may lead to the regulation of mast cell differentiation.

From the Clinical Editor

This team reports on the development of a honeycomb-like structured film to study mast cell differentiation of non-cancerous origin, demonstrating that different microenvironments provided by different honeycomb hole sizes determine the morphology of the differentiated cells.

Graphical Abstract

Various steps of multinuclear NCL-2 cell (non-tumor mast cell) proliferation on the 3-μm honeycomb-like structured polystyrene film (HCF). A cell adhering to the 3-μm HCF (A), cell ballooning and division along the structure of the hole (B, C), a splitting cell migrating down into the hole of the film (D, E), and individual cells along the holes (F). The images of each stage were not obtained from the same cells. All scale bars are 5 μm.

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Section snippets

Preparation of honeycomb-like films (HCFs)

Porous HCFs of polystyrene were fabricated using water droplets as templates for regular pores. In brief, polystyrene, didecylacrylamide, and w-carboxyhexylacrylamide (CAP) (10:1 wt%) were dissolved in chloroform at a concentration of 5 g/L, and the polymer mixture was layered on cover glasses in a humid environment. The pore size was regulated by changing the casting conditions (temperature, humidity, gas flow rate, casting volume, and casting concentration) of the polymer solution.8, 9

NCL-2 cell line

Cell

NCL-2 cell proliferation on polystyrene films

NCL-2 cells are essentially non-adherent cells, but some do attach to the ECM. At 1 week after seeding at a density of 1 × 105 cells/ml, the total number of NCL-2 cells (adherent plus floating cells) gradually increased with increasing HCF hole size. After 1 week, the NCL-2 cells reached 10.72 × 105 cells/ml on the flat film, 15.36 × 105 cells/ml on the 3-μm HCF, 15.39 × 105 cells/ml on the 5-μm HCF, and 18.53 × 105 cells/ml on the 10-μm HCF. The number of NCL-2 cells adhering to the film was 0.72 × 105

Discussion

In mammalian cells, nuclear division is usually followed by cytoplasmic division. However, megakaryocytes, which produce blood platelets, and some hepatocytes and heart muscle cells become multinucleated.11 Because mast cells are derived from bone marrow hematopoietic cells, they may have the same characteristic cell division pattern as megakaryocytes. We propose that HCFs exhibiting self-organization and a suitable three-dimensional (3D) pore size could provide a physical stimulus for mast

Acknowledgments

The authors would like to thank FUJIFILM Corporation (Ltd), Kanagawa, Japan, for supplying the HCFs.

References (23)

  • M. Tanaka

    Design of novel 2D and 3D biointerfaces using self-organization to control cell behavior

    Biochim Biophys Acta

    (2011)
  • Y. Fukuhira et al.

    Biodegradable honeycomb-patterned film composed of poly(lactic acid) and dioleoylphosphatidylethanolamine

    Biomaterials

    (2006)
  • E.S.A.M. Silveira et al.

    Lipid rafts in mast cell biology

  • M. Maurer et al.

    What is the physiological function of mast cells?

    Exp Dermatol

    (2003)
  • J. Kuchler et al.

    Morphological analysis of integrin-mediated adhesion of immature human mast cells to extracellular matrix proteins

    Arch Dermatol Res

    (2006)
  • S.K. Walsh et al.

    Mast cell degranulation—a mechanism for the anti-arrhythmic effect of endothelin-1?

    Br J Pharmacol

    (2009)
  • S. Mitragotri et al.

    Physical approaches to biomaterial design

    Nat Mater

    (2009)
  • S.J. Hollister

    Porous scaffold design for tissue engineering

    Nat Mater

    (2005)
  • S. Yamamoto et al.

    Effect of honeycomb-patterned surface topography on the adhesion and signal transduction of porcine aortic endothelial cells

    Langmuir

    (2007)
  • Hide M, Okabe T, Hiragun T (invetors). New mast cell strain and method using the same strain. Japanese patent (J-STORE)...
  • B. Alberts et al.

    The cell cycle

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    Conflict of Interest: The authors declare no competing financial interests.

    All sources of support: This work was supported by JSPS KAKENHI (Grant Number 22592099) and Next Generation World-Leading Researchers (NEXT Program Number LS017) in Japan.

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