High-density culture of hepatocytes in a packed-bed bioreactor using a fibrous scaffold from plant

A vegetable sponge (loofa sponge, LS) from the fibrous interior of dried fruits (Luffa cylindrica) was used as a scaffold in a packed-bed bioreactor for dynamic seeding and perfusion culture of rat hepatocytes. Hepatocytes were immobilized within the highly complex macroscopic architecture of LS by attaching to its rough fiber surface. The cell seeding efficiency was independent of the initial loading cell number up to 2 × 10⁸ hepatocytes (1 × 10⁷ cells/cm³ scaffold) and cell distribution within the bioreactor was uniform. Liver-specific functions of the hepatocytes depended on cell seeding density and medium flow rate with the optimum cell density at 3.5 × 10⁶ cells/cm³ scaffold. Spheroids with diameters around 100 μm and high viability from FDA staining were observed at day 3 and spheroids as large as 400 μm could be found at day 4. Cell damage was enhanced due to direct contacting of cells with the flow stream. Nonetheless, hepatocytes metabolic rates were substantially improved over stationary culture on TCPS dishes with production rates of albumin and urea at day 7 maintained at 90% and 67% of their initial values, respectively.