Rat hepatocyte spheroids formed by rocked technique maintain differentiated hepatocyte gene expression and function

Area of Science:

• Hepatology
• Biotechnology
• Tissue Engineering

Background:

• Primary hepatocyte culture in 3D spheroids offers a promising in vitro model for liver studies.
• Hepatocyte spheroids exhibit structural polarity, functional bile canaliculi, and differentiated functions.
• These spheroids are being explored for diagnostic, discovery, and therapeutic applications, including bioartificial livers.

Purpose of the Study:

• To evaluate a novel rocking technique for inducing hepatocyte spheroid formation.
• To assess the kinetics, cell-cell adhesion, gene expression, and biochemical activities of rat hepatocyte spheroids cultured over 14 days.
• To compare the rocker technique with traditional rotational systems for spheroid culture.

Main Methods:

• Primary rat hepatocytes were cultured using a novel rocking technique to form spheroids.
• Kinetics of spheroid formation, cell-cell adhesion, gene expression (including liver-related genes), and biochemical activities were monitored over 14 days.
• Comparison was made with hepatocytes cultured using a traditional rotational system and monolayer cultures.

Main Results:

• The rocking technique resulted in more rapid spheroid formation with reduced nonadherent cells compared to rotational methods.
• Hepatocyte spheroids maintained stable expression of over 80% of tested liver-related genes, including albumin synthesis, urea cycle, and metabolic enzymes.
• Biochemical activity of spheroids cultured by the rocker technique was superior to that of hepatocytes in monolayer cultures.

Conclusions:

• Spheroid formation using the rocker technique is more efficient and faster than using the rotational technique.
• Rocker-formed hepatocyte spheroids are suitable for use as in vitro liver tissue constructs and in bioartificial liver applications.