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Hepatocytes in primary cultures are still the gold standard in the pharmaceutical industry ; however they are expensive and have a very short lifespan with a progressive loss of specific functions over time. The dipolar aprotic solvent, dimethyl sulfoxide (DMSO) is wildly used to stabilize the liver specific function of human hepatocytes in vitro. It is also used to potentialize the dfferentiation of the human HepaRG-hepatocytes-like cells that share functional characteristics with human hepatocytes. Besides its free radicals scavenging properties that contribute to hepatoprotection, DMSO has been shown to interfere with apoptosis signalling pathways. To avoid this disadvantage, we have analysed the functional properties of freshly isolated human hepatocytes (FIH) and HepaRG-hepatocyte-like cells in 3D cultures performed using Ultra Low Attachment Technology (ULA) plates.