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RCCS bioreactor-based modelled microgravity induces significant changes on in vitro 3D neuroglial cell cultures.

Morabito C, Steimberg N, Mazzoleni G, Guarnieri S, Fanò-Illic G, Mariggiò MA - Biomed Res Int (2015)

Bottom Line: We propose a human-derived neuro-/glial cell three-dimensional in vitro model to investigate the effects of microgravity on cell-cell interactions.Moreover, compared to cells as traditional static monolayers, cell aggregates cultured under modelled microgravity showed increased expression of specific differentiation markers (e.g., GL15 cells: GFAP, S100B; SH-SY5Y cells: GAP43) and modulation of functional cell-cell interactions (e.g., N-CAM and Cx43 expression and localisation).In conclusion, this culture model opens a wide range of specific investigations at the molecular, biochemical, and morphological levels, and it represents an important tool for in vitro studies into dynamic interactions and responses of nervous system cell components to microgravity environmental conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Imaging and Clinical Sciences, Unit of Functional Biotechnology, Aging Research Center (Ce.S.I.), "G. d'Annunzio" University of Chieti-Pescara, Via dei Vestini 29, 66100 Chieti, Italy ; Interuniversity Institute of Myology, Italy.

ABSTRACT
We propose a human-derived neuro-/glial cell three-dimensional in vitro model to investigate the effects of microgravity on cell-cell interactions. A rotary cell-culture system (RCCS) bioreactor was used to generate a modelled microgravity environment, and morphofunctional features of glial-like GL15 and neuronal-like SH-SY5Y cells in three-dimensional individual cultures (monotypic aggregates) and cocultures (heterotypic aggregates) were analysed. Cell survival was maintained within all cell aggregates over 2 weeks of culture. Moreover, compared to cells as traditional static monolayers, cell aggregates cultured under modelled microgravity showed increased expression of specific differentiation markers (e.g., GL15 cells: GFAP, S100B; SH-SY5Y cells: GAP43) and modulation of functional cell-cell interactions (e.g., N-CAM and Cx43 expression and localisation). In conclusion, this culture model opens a wide range of specific investigations at the molecular, biochemical, and morphological levels, and it represents an important tool for in vitro studies into dynamic interactions and responses of nervous system cell components to microgravity environmental conditions.

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Glial marker localisation in GL15 cells. Representative confocal images of GL15 cells cultured as a monolayer (2D, (a), (c), and (e)) and under the modelled microgravity (RCCS bioreactor, (b), (d), and (f)) and immunostained with anti-GFAP ((a) and (b)), anti-S100B ((c) and (d)), and anti-Cx43 ((e) and (f)) antibodies (as indicated). The RCCS G-aggregate sections were also stained with propidium iodide (PI). Insets in (b), (d), and (f) show image magnification. Scale bars, 25 μm.
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fig3: Glial marker localisation in GL15 cells. Representative confocal images of GL15 cells cultured as a monolayer (2D, (a), (c), and (e)) and under the modelled microgravity (RCCS bioreactor, (b), (d), and (f)) and immunostained with anti-GFAP ((a) and (b)), anti-S100B ((c) and (d)), and anti-Cx43 ((e) and (f)) antibodies (as indicated). The RCCS G-aggregate sections were also stained with propidium iodide (PI). Insets in (b), (d), and (f) show image magnification. Scale bars, 25 μm.

Mentions: To analyse the expression of the GL15 cell specific phenotype in the G-aggregates cultured in the RCCS bioreactor for up to 2 weeks, immunostaining for glial markers was carried out. The G-aggregates showed glial-cell-specific protein expression similar to that observed in the GL15 cells cultured as monolayers under 2D conditions (Figure 3). The G-aggregates and the GL15 cells cultured as monolayers both showed cytoplasmic localisation of GFAP and S100B, as two markers of the glial cytoskeleton (Figures 3(a)–3(d)).


RCCS bioreactor-based modelled microgravity induces significant changes on in vitro 3D neuroglial cell cultures.

Morabito C, Steimberg N, Mazzoleni G, Guarnieri S, Fanò-Illic G, Mariggiò MA - Biomed Res Int (2015)

Glial marker localisation in GL15 cells. Representative confocal images of GL15 cells cultured as a monolayer (2D, (a), (c), and (e)) and under the modelled microgravity (RCCS bioreactor, (b), (d), and (f)) and immunostained with anti-GFAP ((a) and (b)), anti-S100B ((c) and (d)), and anti-Cx43 ((e) and (f)) antibodies (as indicated). The RCCS G-aggregate sections were also stained with propidium iodide (PI). Insets in (b), (d), and (f) show image magnification. Scale bars, 25 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4309310&req=5

fig3: Glial marker localisation in GL15 cells. Representative confocal images of GL15 cells cultured as a monolayer (2D, (a), (c), and (e)) and under the modelled microgravity (RCCS bioreactor, (b), (d), and (f)) and immunostained with anti-GFAP ((a) and (b)), anti-S100B ((c) and (d)), and anti-Cx43 ((e) and (f)) antibodies (as indicated). The RCCS G-aggregate sections were also stained with propidium iodide (PI). Insets in (b), (d), and (f) show image magnification. Scale bars, 25 μm.
Mentions: To analyse the expression of the GL15 cell specific phenotype in the G-aggregates cultured in the RCCS bioreactor for up to 2 weeks, immunostaining for glial markers was carried out. The G-aggregates showed glial-cell-specific protein expression similar to that observed in the GL15 cells cultured as monolayers under 2D conditions (Figure 3). The G-aggregates and the GL15 cells cultured as monolayers both showed cytoplasmic localisation of GFAP and S100B, as two markers of the glial cytoskeleton (Figures 3(a)–3(d)).

Bottom Line: We propose a human-derived neuro-/glial cell three-dimensional in vitro model to investigate the effects of microgravity on cell-cell interactions.Moreover, compared to cells as traditional static monolayers, cell aggregates cultured under modelled microgravity showed increased expression of specific differentiation markers (e.g., GL15 cells: GFAP, S100B; SH-SY5Y cells: GAP43) and modulation of functional cell-cell interactions (e.g., N-CAM and Cx43 expression and localisation).In conclusion, this culture model opens a wide range of specific investigations at the molecular, biochemical, and morphological levels, and it represents an important tool for in vitro studies into dynamic interactions and responses of nervous system cell components to microgravity environmental conditions.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience, Imaging and Clinical Sciences, Unit of Functional Biotechnology, Aging Research Center (Ce.S.I.), "G. d'Annunzio" University of Chieti-Pescara, Via dei Vestini 29, 66100 Chieti, Italy ; Interuniversity Institute of Myology, Italy.

ABSTRACT
We propose a human-derived neuro-/glial cell three-dimensional in vitro model to investigate the effects of microgravity on cell-cell interactions. A rotary cell-culture system (RCCS) bioreactor was used to generate a modelled microgravity environment, and morphofunctional features of glial-like GL15 and neuronal-like SH-SY5Y cells in three-dimensional individual cultures (monotypic aggregates) and cocultures (heterotypic aggregates) were analysed. Cell survival was maintained within all cell aggregates over 2 weeks of culture. Moreover, compared to cells as traditional static monolayers, cell aggregates cultured under modelled microgravity showed increased expression of specific differentiation markers (e.g., GL15 cells: GFAP, S100B; SH-SY5Y cells: GAP43) and modulation of functional cell-cell interactions (e.g., N-CAM and Cx43 expression and localisation). In conclusion, this culture model opens a wide range of specific investigations at the molecular, biochemical, and morphological levels, and it represents an important tool for in vitro studies into dynamic interactions and responses of nervous system cell components to microgravity environmental conditions.

Show MeSH