The role of connexin32 and connexin43 in the control of hepatocellular homeostasis: development of a long-term primary hepatocyte culture system

Since the liver, and the hepatocyte in particular, plays a key role in generating the pharmaco-toxicological outcome of drugs in the body, primary cultures of hepatocytes are well-acknowledged in vitro tools to investigate potential adverse effects of candidate pharmaceuticals in the early phases of drug development. Their use, however, is mainly restricted to short-term applications, because of the progressive loss of hepatocyte-specific in vivo features followed by the onset of spontaneous cell death during cultivation. A hallmark of this so-called dedifferentiation process is the decreased capacity of hepatocytes to communicate via gap junctions, being intercellular channels that are composed of connexin (Cx) proteins. In vivo, hepatocytes abundantly express Cx32, but upon their cultivation, Cx32 production drastically decreases, whereas Cx43 becomes increasingly detectable. The biological relevance of this connexin switch phenomenon is unclear and will be addressed in the current project. Preliminary research from the host laboratory suggests a protective role for Cx43 against spontaneous cell death and this research track will therefore be pursued in the first part of the present project. In the second part, two innovative anti-dedifferentiation strategies, focused on the enhancement of Cx32 production in primary hepatocyte cultures, will be developed and tested for their efficiency in maintaining functional hepatocyte-specific features in vitro. By doing so, this research project directly contributes to the development of a primary hepatocyte culture system that can serve a broad range of long-term pharmaco-toxicological purposes.