Gut-liver crosstalk and gut homeostasis in paracetamol toxicity: In Vitro Study Using Impedence-Based Assay

Introduction: Drug induced liver injury accounts for approximately one half of all acute liver failure cases, with Paracetamol (APAP) being one of the most reported hepatotoxins [1]. We used an in vitro approach to model APAP transport from gut to liver where metabolic products (NAPQI) can cause hepatocellular damage. Investigating the mechanisms of a gut-liver axis in vitro may contribute to a better understanding of APAP toxicity.
Methods: The cellular impedance ECIS Z &theta1; platform was used to study the gut-liver axis and effects of APAP. Caco-2 and HepaRGs were seeded on separate 8W10+E ibidi arrays. The Caco-2 array was collagen coated. Step 1: Previously established concentrations of APAP [2] were applied to confluent Caco-2s (day 10), and impedance monitored for 24 hrs. Step 2: This ‘preconditioned’ culture medium was then transferred to HepaRGs (day 11) for 24 hrs. Step 3: preconditioned culture medium was then transferred from HepaRGs to fresh Caco-2s (day 12) for 24 hrs.
Results: Step 1: using an impedance based cellular assay, we show a tightening of barrier function in Caco-2s treated with 20 mM APAP. Step 2: only the highest concentration (20 mM APAP) shows substantial loss of impedance on hepatocytes. Step 3: we demonstrate that barrier function has become sensitized on Caco-2s with an increase in impedance at 10 mM
Discussion and Conclusions: This study demonstrates a potential cross-talk between hepatocytes and enterocytes and reveals a homeostatic effect of gut barrier function in presence of APAP. Exposing hepatocytes to toxic levels of APAP taken from Caco-2’s attenuates toxicity at 5 and 10 mM concentrations compared to previous study [2] where a dose-dependent loss of tight junctions was observed (5–20 mM APAP). Further exposure of preconditioned media on fresh Caco-2s (step 3) shows that tightening of barrier function is now achieved with lower APAP concentration (10 mM) to reduce absorption of APAP. It is possible a liver-gut axis regulates APAP absorption through paracrine signals, though more work is needed. This may differ between population phenotypes and potentially be responsible for variability of toxic response in patients with APAP overdose.