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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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Related in: MedlinePlus

Expression levels of functional markers in the GS-aggregates. Representative Western blotting and quantification of the levels of N-CAM and Cx43 in GS-aggregates cultured in the RCCS bioreactor for 24 h and 2 weeks (2 w). Data are densitometric ratio analyses as means ±SEM from 3 independent experiments. *P < 0.05, for 2-week versus respective 24 h cultures.
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fig10: Expression levels of functional markers in the GS-aggregates. Representative Western blotting and quantification of the levels of N-CAM and Cx43 in GS-aggregates cultured in the RCCS bioreactor for 24 h and 2 weeks (2 w). Data are densitometric ratio analyses as means ±SEM from 3 independent experiments. *P < 0.05, for 2-week versus respective 24 h cultures.

Mentions: Western blotting of N-CAM and Cx43 expression levels revealed that, in the GS-aggregates, the monomeric Cx43 protein (43 kDa) was downregulated during the RCCS bioreactor incubation. Interestingly, homogenates from the GS-aggregates showed a Cx43-positive band at 86 kDa, which demonstrates the presence of a dimeric form of Cx43, which was highly expressed in the initial phases of the coculture (over the first 24 h), and which significantly decreased over the 2 weeks of the GS-aggregates in the RCCS bioreactor (Figure 10). The N-CAM isoform expression pattern showed a slight, but not significant, decrease in N-CAM-180 levels and a significant increase in N-CAM-140 levels (Figure 10), which resembled the N-CAM-140 increase observed in the S-aggregate homogenates.


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)

Expression levels of functional markers in the GS-aggregates. Representative Western blotting and quantification of the levels of N-CAM and Cx43 in GS-aggregates cultured in the RCCS bioreactor for 24 h and 2 weeks (2 w). Data are densitometric ratio analyses as means ±SEM from 3 independent experiments. *P < 0.05, for 2-week versus respective 24 h cultures.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

fig10: Expression levels of functional markers in the GS-aggregates. Representative Western blotting and quantification of the levels of N-CAM and Cx43 in GS-aggregates cultured in the RCCS bioreactor for 24 h and 2 weeks (2 w). Data are densitometric ratio analyses as means ±SEM from 3 independent experiments. *P < 0.05, for 2-week versus respective 24 h cultures.
Mentions: Western blotting of N-CAM and Cx43 expression levels revealed that, in the GS-aggregates, the monomeric Cx43 protein (43 kDa) was downregulated during the RCCS bioreactor incubation. Interestingly, homogenates from the GS-aggregates showed a Cx43-positive band at 86 kDa, which demonstrates the presence of a dimeric form of Cx43, which was highly expressed in the initial phases of the coculture (over the first 24 h), and which significantly decreased over the 2 weeks of the GS-aggregates in the RCCS bioreactor (Figure 10). The N-CAM isoform expression pattern showed a slight, but not significant, decrease in N-CAM-180 levels and a significant increase in N-CAM-140 levels (Figure 10), which resembled the N-CAM-140 increase observed in the S-aggregate homogenates.

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
Related in: MedlinePlus