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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

Scanning electron microscopy of silicified cells.(a) Microvilli-like structures on the surface of HepG2, iPS(IMR90)-4, and WA07 cells in 2D culture and in 3D NFC hydrogel culture. (b) Extracellular matrix-like material on a HepG2 cell spheroid in the NFC hydrogel for 8 days. (c) Silica-replicas of WA07 spheroids (5 days in the NFC hydrogel) after calcination. Images are representative of eight biological samples.
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f3: Scanning electron microscopy of silicified cells.(a) Microvilli-like structures on the surface of HepG2, iPS(IMR90)-4, and WA07 cells in 2D culture and in 3D NFC hydrogel culture. (b) Extracellular matrix-like material on a HepG2 cell spheroid in the NFC hydrogel for 8 days. (c) Silica-replicas of WA07 spheroids (5 days in the NFC hydrogel) after calcination. Images are representative of eight biological samples.

Mentions: At higher magnification we observed fine cellular structures on the cell membrane of the cell-silica composites. Dense microvilli-like structures were observed in the hPSCs and HepG2 cells cultured in both 2D and 3D (Fig. 3a). Such structures were observed on the surface of hESCs in an earlier study15. Surprisingly, we observed abundant extracellular materials on the surfaces of HepG2 cells cultured in the NFC hydrogel (Fig. 3b). These extracellular materials are likely produced by HepG2 cells since the NFC hydrogel has been degraded by cellulase and the silicified nanofiber bundles of the NFC hydrogel appear different (Supplementary Fig. 2).


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)

Scanning electron microscopy of silicified cells.(a) Microvilli-like structures on the surface of HepG2, iPS(IMR90)-4, and WA07 cells in 2D culture and in 3D NFC hydrogel culture. (b) Extracellular matrix-like material on a HepG2 cell spheroid in the NFC hydrogel for 8 days. (c) Silica-replicas of WA07 spheroids (5 days in the NFC hydrogel) after calcination. 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

f3: Scanning electron microscopy of silicified cells.(a) Microvilli-like structures on the surface of HepG2, iPS(IMR90)-4, and WA07 cells in 2D culture and in 3D NFC hydrogel culture. (b) Extracellular matrix-like material on a HepG2 cell spheroid in the NFC hydrogel for 8 days. (c) Silica-replicas of WA07 spheroids (5 days in the NFC hydrogel) after calcination. Images are representative of eight biological samples.
Mentions: At higher magnification we observed fine cellular structures on the cell membrane of the cell-silica composites. Dense microvilli-like structures were observed in the hPSCs and HepG2 cells cultured in both 2D and 3D (Fig. 3a). Such structures were observed on the surface of hESCs in an earlier study15. Surprisingly, we observed abundant extracellular materials on the surfaces of HepG2 cells cultured in the NFC hydrogel (Fig. 3b). These extracellular materials are likely produced by HepG2 cells since the NFC hydrogel has been degraded by cellulase and the silicified nanofiber bundles of the NFC hydrogel appear different (Supplementary Fig. 2).

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