Hepatocytes Cultured on Collagen Modified Micropatterned Agarose for Evaluating Inflammatory and Oxidative Stress Responses

Introduction: Evaluation of hepatocyte response to inflammatory and oxidative stress inducing agents has been hampered by limited availability of appropriate in vitro testing methods. There is an unmet need for in vitro culture techniques to identify potential deleterious agents or stimuli to liver cells. Materials and Methods: We designed a culture system consisting of collagen covalently attached to agarose patterned with microchannels and determined whether this system could be used to assess hepatocyte inflammatory and oxidative stress responses. Results and Discussion: Hepatocytes cultured in collagen modified agarose gels displayed features of differentiated phenotype compared to control cell cultures in nonpatterned agarose. These cells responded to inflammatory interleukin-1 beta (IL-1β) and oxidative stress hydrogen peroxide (H2O2) stimuli. IL-1β induced production of pro-inflammatory prostaglandin E2 (PGE2), IL-8, and macrophage chemotactic protein-1 (MCP-1, also known as CCL-2). H2O2 induced PGE2 production, and intracellular levels of the antioxidant peptide reduced glutathione. Hepatocyte response to stimuli was associated with translocation of nuclear factor-kappa beta (NF-κB) and nuclear factor erythroid 2-related factor 2 (Nrf2). NF-κB and Nrf2 are transcription factors that modulate the inflammatory and oxidative stress response, respectively. Conclusion: The present study demonstrates that our culture design facilitated hepatocyte adhesion, survival, and function. This in vitro model may be useful in studying response to inflammatory and oxidative stimuli, the mechanisms involved in this response, and their potential inhibitors.