Optimal Number of Hepatocytes per Microcarrier in Spheroid Culture using Cytodex 3 Microcarrier

Shin, Woo Young; Lee, Kuhn Uk; Lee, Hae Won; Cho, Eung Ho; Yi, Nam Joon; Suh, Kyung Suk

Abstract

PURPOSE: The mass cultivation of functional hepatocytes is a key factor of a bioartificial liver. Combining spheroid and microcarrier cultures has been applied for enhancing the cell viability and metabolic activities. Hence, the optimal number of hepatocytes per microcarrier was investigated.
METHODS
Firstly, spheroid cultures were carried out with 1 g Cytodex 3 microcarrier plus 2 x 10(9), 4 x 10(8) and 8 x 10(7) viable hepatocytes per flask. The numbers of hepatocytes per microcarrier were approximately 666.7, 133.3 and 26.7, respectively. The control group consisted of a spheroid culture of 4 x 10(8) hepatocytes without any microcarrier. According to the primary experimental results, spheroid cultures with 1 x 10(8) of hepatocytes plus 1 g, 2 g and 3 g of the Cytodex 3 microcarrier were performed. The numbers of hepatocytes per microcarrier were approximately 33.3, 16.7 and 11.1, respectively. The control group consisted of a spheroid culture of 1 x 10(8) hepatocytes. The cell viabilities were assayed using a Cell Counting Kit-8; with the albumin production assayed using ELISA.
RESULTS
According to the primary experiment, the group consisting of 26.7 hepatocytes per microcarrier showed the highest viability (P<0.01). However, there was no statistical difference in the albumin production between the groups (P=0.744). The second Experiment showed the groups consisting of 11.1 and 16.7 hepatocytes per microcarrier had higher viabilities than the other hepatocyte and control groups (P<0.01). The albumin production was similar for each group (P=0.187).
CONCLUSION
With respect to their application to a bioartificial liver, about 130 hepatocytes per microcarrier was appeared to be good for the mass cultivation of a hepatocytes spheroid culture using the Cytodex 3 microcarrier.