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Silica bioreplication preserves three-dimensional spheroid structures of human pluripotent stem cells and HepG2 cells.

Lou YR, Kanninen L, Kaehr B, Townson JL, Niklander J, Harjumäki R, Jeffrey Brinker C, Yliperttula M - Sci Rep (2015)

Bottom Line: It has proven challenging to stabilize spheroid architectures for detailed morphological examination.Furthermore, SBR preserves extracellular matrix-like materials and cellular proteins.These findings provide the first evidence of intact hPSC spheroid architectures and similar fine structures to 2D-cultured cells, providing a pathway to enable our understanding of morphogenesis in 3D cultures.

View Article: PubMed Central - PubMed

Affiliation: Centre for Drug Research, Division of Pharmaceutical Biosciences, Faculty of Pharmacy, the University of Helsinki, Helsinki 00014, Finland.

ABSTRACT
Three-dimensional (3D) cell cultures produce more in vivo-like multicellular structures such as spheroids that cannot be obtained in two-dimensional (2D) cell cultures. Thus, they are increasingly employed as models for cancer and drug research, as well as tissue engineering. It has proven challenging to stabilize spheroid architectures for detailed morphological examination. Here we overcome this issue using a silica bioreplication (SBR) process employed on spheroids formed from human pluripotent stem cells (hPSCs) and hepatocellular carcinoma HepG2 cells cultured in the nanofibrillar cellulose (NFC) hydrogel. The cells in the spheroids are more round and tightly interacting with each other than those in 2D cultures, and they develop microvilli-like structures on the cell membranes as seen in 2D cultures. Furthermore, SBR preserves extracellular matrix-like materials and cellular proteins. These findings provide the first evidence of intact hPSC spheroid architectures and similar fine structures to 2D-cultured cells, providing a pathway to enable our understanding of morphogenesis in 3D cultures.

No MeSH data available.


Related in: MedlinePlus

Morphology of human pluripotent stem cells and hepatocellular carcinoma cells with and without silica bioreplication.(a) SEM images of HepG2, iPS(IMR90)-4, and WA07 cell spheroids show deformation of 3D spheroids. (b) SEM images of HepG2, iPS(IMR90)-4, and WA07 cell spheroids after silica bioreplication show well-preserved spheroid architecture and tight cell-cell contact in the NFC hydrogel-based 3D cultures. Images are representative of eight biological samples.
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f2: Morphology of human pluripotent stem cells and hepatocellular carcinoma cells with and without silica bioreplication.(a) SEM images of HepG2, iPS(IMR90)-4, and WA07 cell spheroids show deformation of 3D spheroids. (b) SEM images of HepG2, iPS(IMR90)-4, and WA07 cell spheroids after silica bioreplication show well-preserved spheroid architecture and tight cell-cell contact in the NFC hydrogel-based 3D cultures. Images are representative of eight biological samples.

Mentions: To study the detailed cellular structures of the cells cultured in 2D and 3D, we prepared the cell samples and cell-silica composites for SEM. We observed dramatic differences between non-silicified spheroids and silicified spheroids. The non-silicified spheroids deformed considerably during sample preparation, presumably during the drying procedure, resulting in obscuration of surface features (Fig. 2a). In contrast, the cell spheroid-silica composites stabilized using SBR retained their spherical morphology and were well preserved (Fig. 2b). Both the hPSCs and HepG2 cells developed tight cell-cell interactions during 8-day 3D culture in the NFC hydrogel (Fig. 2b). Cells in 3D spheroids appeared more round than those in 2D culture (Fig. 2b). We observed protrusions in some elongated HepG2 cells and also in some hPSCs at the edge of the colonies, but not in the cells in 3D spheroids (Fig. 2b). Some small cracks were observed in the hPSC colonies in 2D culture (Fig. 2b), which was presumably caused by dehydration during sample preparation for SEM.


Silica bioreplication preserves three-dimensional spheroid structures of human pluripotent stem cells and HepG2 cells.

Lou YR, Kanninen L, Kaehr B, Townson JL, Niklander J, Harjumäki R, Jeffrey Brinker C, Yliperttula M - Sci Rep (2015)

Morphology of human pluripotent stem cells and hepatocellular carcinoma cells with and without silica bioreplication.(a) SEM images of HepG2, iPS(IMR90)-4, and WA07 cell spheroids show deformation of 3D spheroids. (b) SEM images of HepG2, iPS(IMR90)-4, and WA07 cell spheroids after silica bioreplication show well-preserved spheroid architecture and tight cell-cell contact in the NFC hydrogel-based 3D cultures. Images are representative of eight biological samples.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4555166&req=5

f2: Morphology of human pluripotent stem cells and hepatocellular carcinoma cells with and without silica bioreplication.(a) SEM images of HepG2, iPS(IMR90)-4, and WA07 cell spheroids show deformation of 3D spheroids. (b) SEM images of HepG2, iPS(IMR90)-4, and WA07 cell spheroids after silica bioreplication show well-preserved spheroid architecture and tight cell-cell contact in the NFC hydrogel-based 3D cultures. Images are representative of eight biological samples.
Mentions: To study the detailed cellular structures of the cells cultured in 2D and 3D, we prepared the cell samples and cell-silica composites for SEM. We observed dramatic differences between non-silicified spheroids and silicified spheroids. The non-silicified spheroids deformed considerably during sample preparation, presumably during the drying procedure, resulting in obscuration of surface features (Fig. 2a). In contrast, the cell spheroid-silica composites stabilized using SBR retained their spherical morphology and were well preserved (Fig. 2b). Both the hPSCs and HepG2 cells developed tight cell-cell interactions during 8-day 3D culture in the NFC hydrogel (Fig. 2b). Cells in 3D spheroids appeared more round than those in 2D culture (Fig. 2b). We observed protrusions in some elongated HepG2 cells and also in some hPSCs at the edge of the colonies, but not in the cells in 3D spheroids (Fig. 2b). Some small cracks were observed in the hPSC colonies in 2D culture (Fig. 2b), which was presumably caused by dehydration during sample preparation for SEM.

Bottom Line: It has proven challenging to stabilize spheroid architectures for detailed morphological examination.Furthermore, SBR preserves extracellular matrix-like materials and cellular proteins.These findings provide the first evidence of intact hPSC spheroid architectures and similar fine structures to 2D-cultured cells, providing a pathway to enable our understanding of morphogenesis in 3D cultures.

View Article: PubMed Central - PubMed

Affiliation: Centre for Drug Research, Division of Pharmaceutical Biosciences, Faculty of Pharmacy, the University of Helsinki, Helsinki 00014, Finland.

ABSTRACT
Three-dimensional (3D) cell cultures produce more in vivo-like multicellular structures such as spheroids that cannot be obtained in two-dimensional (2D) cell cultures. Thus, they are increasingly employed as models for cancer and drug research, as well as tissue engineering. It has proven challenging to stabilize spheroid architectures for detailed morphological examination. Here we overcome this issue using a silica bioreplication (SBR) process employed on spheroids formed from human pluripotent stem cells (hPSCs) and hepatocellular carcinoma HepG2 cells cultured in the nanofibrillar cellulose (NFC) hydrogel. The cells in the spheroids are more round and tightly interacting with each other than those in 2D cultures, and they develop microvilli-like structures on the cell membranes as seen in 2D cultures. Furthermore, SBR preserves extracellular matrix-like materials and cellular proteins. These findings provide the first evidence of intact hPSC spheroid architectures and similar fine structures to 2D-cultured cells, providing a pathway to enable our understanding of morphogenesis in 3D cultures.

No MeSH data available.


Related in: MedlinePlus