Substrate stiffness regulates primary hepatocyte functions.

Area of Science:

• Biomedical Engineering
• Hepatology
• Cell Biology

Background:

• Chronic liver injury leads to fibrosis, characterized by increased liver stiffness due to extracellular matrix deposition.
• Liver stiffness is a critical regulator of hepatic cell function, but its precise effects are not fully understood.
• Engineered in vitro models are needed to mimic disease-specific liver stiffness and study cellular responses.

Purpose of the Study:

• To investigate the impact of substrate stiffness on primary rat hepatocyte behavior and function.
• To develop a polydimethylsiloxane (PDMS)-based model system that recreates healthy (2 kPa) and diseased (55 kPa) liver stiffness.

Main Methods:

• Primary rat hepatocytes were cultured on PDMS substrates with tunable stiffness (2 kPa, 55 kPa) and standard tissue culture plate surfaces (TCPS).
• Hepatocyte phenotype, function (urea and albumin synthesis, CYP activity), gene expression (drug transporters), and cell markers were assessed over time.

Main Results:

• Hepatocytes on soft substrates (2 kPa) maintained a more differentiated and functional phenotype for longer durations compared to stiff substrates (55 kPa) and TCPS.
• Softer substrates promoted higher urea and albumin synthesis, and significantly higher Cytochrome P450 (CYP) activity.
• Increased stiffness downregulated key drug transporter genes and impaired the epithelial cell phenotype, while soft substrates maintained better cell marker expression.

Conclusions:

• Substrate stiffness is a significant factor modulating hepatocyte behavior, differentiation, and function.
• PDMS-based models with tunable stiffness can effectively mimic liver microenvironments and are valuable tools for studying liver disease progression.
• Understanding hepatocyte-liver microenvironment communication through substrate stiffness is crucial for developing new therapeutic strategies for liver diseases.