Hepatoprotective Effect of HUVEC Cells in an In Vitro Hepatic Co-Culture model of Acetaminophen Toxicity

Background and Aims: In vitro models of drug hepatotoxicity using hepatic cell lines, and more recently co-cultures, are highly desirable platforms for screening toxins and understanding underlying mechanisms. In this study, we investigated whether the presence of Human Umbilical Vein Endothelial Cells (HUVEC) can protect hepatic cells against acetaminophen toxicity in an in vitro co-culture model using HepG2/C3A cells with HUVEC.

Methods: HepG2/C3A cells and HUVEC were cultured separately or co-cultured at a ratio of 1:1. After 2 days, cultures were exposed to 0mM, 5mM, 10mM, and 20mM of acetaminophen for 24h. C3A viability was assessed by Cell Viability assay for ATP levels (Promega), total nitric oxide (R&D systems) and total Glutathione levels were measured using an enzymatic method.

Results: In HepG2/C3A monoculture, high concentrations (>10mM) of acetaminophen led to a significant reduction in cellular ATP content associated with cell death and depletion of intracellular glutathione and total nitric oxide. By contrast, HepG2/C3A cells co-cultured with HUVEC showed a significantly higher ATP content when exposed to >10mM of acetaminophen with a relative increase
in nitric oxide and glutathione.

Conclusions: In this study, we have demonstrated an interaction between hepatocytes and endothelial cells in response to the hepatotoxic drug acetaminophen. In the presence of HUVEC, cell viability was maintained and synthesis of total nitric oxide and glutathione was increased in response to toxic insult. The application of this hepatoprotective effect of endothelial cells in vivo
could enhance the integrity of the hepatic sinusoid endothelium, in addition to administration of N-acetyl-L-cysteine in acetaminophen poisoning.