Human Hepatic HepaRG Cells Maintain an Organotypic Phenotype with High Intrinsic CYP450 Activity/Metabolism and Significantly Outperform Standard HepG2/C3A Cells for Pharmaceutical and Therapeutic Applications.

Conventional in vitro human hepatic models for drug testing are based on the use of standard cell lines derived from hepatomas or primary human hepatocytes (PHHs). Limited availability, interdonor functional variability and early phenotypic alterations in PHHs restrict their use, whilst standard cell lines such as HepG2 lack a substantial and variable set of liver-specific functions such as CYP450 activity. Alternatives include the HepG2-derivative C3A cells selected as a more differentiated and metabolically active hepatic phenotype. Human HepaRG cells are an alternative organotypic co-culture model of hepatocytes and cholangiocytes reported to maintain in vivo-like liver-specific functions, including intact Phase I-III drug metabolism. In this study, we compared C3A and human HepaRG cells using phenotypic profiling, CYP450 activity and drug metabolism parameters to assess their value as hepatic models for pre-clinical drug testing or therapeutics. Compared with C3As, HepaRG co-cultures exhibit a more organotypic phenotype, including evidence of hepatic polarity with the strong expression of CYP3A4, the major isoform involved in the metabolism of over 60% of marketed drugs. Significantly greater CYP450 activity and expression of CYP1A2, CYP2E1 and CYP3A4 genes in HepaRG cells (comparable with that of human liver tissue) was demonstrated. Moreover, HepaRG cells also preferentially expressed the hepatic integrin α5 ß1 - an important modulator of cell behaviour including growth and survival, differentiation and polarity. Drug metabolite profiling of phenacetin (CYP1A2) and testosterone (CYP3A4) using LC-MS/MS and HPLC, respectively, revealed that HepaRGs had more intact (Phase I-II) metabolism profile. Thus, HepaRG cells significantly outperform C3A cells for the potential pharmaceutical and therapeutic applications.

Combined decellularisation and dehydration improves the mechanical properties of tissue-engineered sinews.

Novel sources of replacement sinews are needed to repair damaged tissue after injury. The current methods of repair ultilise autografts, allografts or xenografts, although each method has distinct disadvantages that limit their success. Decellularisation of harvested tissues has been previously investigated for sinew repair with the long-term aim of repopulating the structure with autologous cells. Although this procedure shows promise, the demand for donor scaffolds will always outweigh supply. Here, we report the fabrication of fibrin-based tissue-engineered sinews, which can be decellularised, dehydrated and stored. The sinews may then be rehydrated and repopulated with an autologous cell population. In addition to enabling production of patient-specific implants, interestingly, the process of combined decellularisation, dehydration and rehydration enhanced the mechanical properties of the sinew. The treated sinews exhibited a 2.6-fold increase in maximum load and 8-fold increase in ultimate tensile strength when compared with the control group (p < 0.05 in both cases).