Construction of Hybrid Cell Spheroids Using Cell-Sized Cross-Linked Nanogel Microspheres as an Artificial Extracellular Matrix.

The introduction of functional material supports or spacers into cell spheroids increases the free volume, allowing oxygen, nutrients, and waste products to diffuse in and out more freely. Here, a biocompatible polysaccharide spacer material was investigated. Microspheres were prepared by cross-linking cholesterol-modified pullulan (CHP) nanogels with poly(ethylene glycol) (PEG). The ratio of modified CHP nanogel to PEG cross-linker was optimized to give uniform microspheres with an average diameter of approximately 14 µm. Rhodamine B-labeled microspheres showed a homogeneous assembly with bone marrow-derived mesenchymal stem cells (1:1 ratio) to create hybrid cell spheroids. The addition of the cross-linked nanogel spacers did not affect the cell viability, indicating that the microspheres provided a biocompatible scaffold that supported cell proliferation. In addition, the microspheres were stable under culture conditions over 14 days. The hybrid cell spheroids were scaled up to millimeter size to demonstrate their potential as a transplantable treatment, and the cells were found to maintain their high viability. The hybrid cell spheroids are expected to support the production of organoids.

New approach for the establishment of an hepatocyte cell line derived from rat early embryonic stem cells.

A cell line with the characteristics of hepatocytes was established from rat early embryonic stem cells (REES). This cell line was established using a new novel method of Ishiwata et al. from two cell embryos taken from the spontaneous dwarf rat (SDR). The hepatocyte cell line (REES-hep) was instituted from dark red colored tissue in embryos during embryogenesis using REES cell line cultured in the presence of embryotrophic factors. These cell lines were cultured with DMEM/F12 medium supplemented 10% FBS and 1 ng/ml of LIF. They were found to maintain their diploid state, were characterized with 42 normal chromosomes and proliferated to confluence; contact inhibition was also present. These cells produced albumin when cultured using a collagen sponge gel system and reconstructed in a funicular form resembling the cell cords of liver. The cells also produced albumin and bilirubin when transplanted into the spleen of SDR Reconstruction of a REES-hep cell line from early embryonic stem cells should help in treating hepatic insufficient patients. It will be valuable for further research, as an introduction to cell transplantation and application for use in a bio-hybrid typed liver apparatus.