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

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.

Novel Breast Cancer Brain Metastasis Patient-Derived Orthotopic Xenograft Model for Preclinical Studies

“…Brain metastasis of one breast cancer patient (Table S1) that had been passaged three passages as brain PDX was used for this experiment. 2 µL of RPMI-1640 with collagenase and hyaluronidase at commonly used concentrations [34] were added to a tumor fragment (1 mm 3 ) and the mix was incubated at 37 • C for an hour, after which the tumor preparation was washed and resuspended to 3 µL with PBS for Digestion method. Post-operative mortality was zero regardless of enzymatic digestion technique.…”

Nanofibrillar cellulose hydrogels and reconstructed hydrogels as matrices for controlled drug release

“…Nanofibrillated cellulose (NFC) hydrogel has been widely investigated in pharmaceutical and biomedical fields, such as in drug delivery (Laurén, et al 2014), scaffold synthesis (Borges, et al 2011), and as cell and drug carriers (Thérien-Aubin, et al 2016;Valo, et al 2013). NFC hydrogel is also applicable as a cell culture scaffold for three dimensional cell culturing (Bhattacharya, et al 2012;Lou, et al 2014;Lou, et al 2015;Malinen, et al 2014). It is produced from the natural biopolymer cellulose, and wood pulp is a commonly used starting material in the production (Klemm, et al 2011).…”

Effects of nanofibrillated cellulose hydrogels on adipose tissue extract and hepatocellular carcinoma cell spheroids in freeze-drying

“…Surface structures and cell membranes of freeze-dried HepG2 cell spheroids observed with SEM appeared intact and resembled the reference images [36]. It appears that the cell membrane is intact in the dry state and damaged during the rehydration phase.…”