Journal: Cell transplantation

Article Title: Serum-Free Medium and Mesenchymal Stromal Cells Enhance Functionality and Stabilize Integrity of Rat Hepatocyte Spheroids

doi: 10.3727/096368912X656054

Figure Lengend Snippet: Experiment design. Rats were provided normal protein diet (NPD) or high protein diet (HPD) for 7 days before hepatocyte harvest. Freshly isolated hepatocytes were suspended in serum-free medium (SFM) or serum-containing medium (SCM) [Williams-E medium supplemented with 10% fetal bovine serum (FBS)] at a concentration 1 × 106 viable cells/ml. In heterotypic coculture system, hepatocytes (1 × 106/ml) were mixed with mesenchymal stromal cells (MSCs; 1 × 105/ml) and cultured in spheroid dishes. 20 ml suspended hepatocytes were inoculated into spheroid dishes (10 × 8 × 2 cm) and placed in the 37°C incubator with 5% CO2 and rocked continuously at 10 cycles per minute to induce spheroid formation and maintain suspension of spheroids. Culture medium was first changed at 24 h and subsequently at 3-day intervals. Dishes were sampled at 24 h after each medium change. Four culture groups were studied as follows: HPD + SFM + MSC group, hepatocytes harvested from high protein dietary rat and cultured in serum-free medium and cocultured with MSCs; HPD + SFM group, hepatocytes harvested from high protein dietary rat and cultured in serum-free medium; NPD + SFM group, hepatocytes harvested from normal protein dietary rat and cultured in serum-free medium; NPD + SCM group, hepatocytes harvested from normal protein dietary rat and cultured in serum-containing medium.

Article Snippet: Spheroid Culture Freshly isolated hepatocytes were suspended in serum-free medium (SFM) ( ) or serum-containing medium (SCM) [Williams-E medium supplemented with 10% fetal bovine serum (FBS; Mediatech, Inc., Herndon, VA), 10 U/ml penicillin G, 100 μg/ml streptomycin, 10 μg/ml insulin, 5.5 μg/ml transferrin, 5 ng/ml sodium selenite] at a concentration of 1 × 10 6 viable cells/ml.

Techniques: Isolation, Concentration Assay, Cell Culture, Suspension

Journal: Cell transplantation

Article Title: Serum-Free Medium and Mesenchymal Stromal Cells Enhance Functionality and Stabilize Integrity of Rat Hepatocyte Spheroids

doi: 10.3727/096368912X656054

Figure Lengend Snippet: Hepatocyte count in suspension, incorporation into spheroids and viability. Cell counts were measured by Multisizer 3 Beckman Coulter counter. Cells were inoculated (day 0) at 1 × 106/ml. (A) Proportion of cells present in spheroids (>60 μm diameter). (B) Percent of hepatocyte mass left at each time point versus original cells inoculated of NPD + SCM. (C) Percent viability of cells in spheroids at each time point. Values represent the averages of three independent experiments. **p < 0.001; *p < 0.05 versus NPD + SCM at each time point.

Article Snippet: Spheroid Culture Freshly isolated hepatocytes were suspended in serum-free medium (SFM) ( ) or serum-containing medium (SCM) [Williams-E medium supplemented with 10% fetal bovine serum (FBS; Mediatech, Inc., Herndon, VA), 10 U/ml penicillin G, 100 μg/ml streptomycin, 10 μg/ml insulin, 5.5 μg/ml transferrin, 5 ng/ml sodium selenite] at a concentration of 1 × 10 6 viable cells/ml.

Techniques: Suspension