An important function of the liver is to excrete xenobiotics and endogenous metabolites via bile into feces. Bile represents one of the most toxic body fluids and its leakage from the biliary tract into the parenchyma causes severe tissue injury (Jansen et al., 2017). Thus, bile flux and mechanisms that guarantee a safe excretion are critical for tissue integrity. Traditionally, bile excretion has been described by the mechano-osmotic concept, according to which bile acids are actively secreted into the bile canaliculi; since they are osmotically active they draw water into canalicular lumen, causing liquid flow within the bile canaliculi that finally generates extrahepatic bile flow.
However, flow within the bile canaliculi has never been measured since these structures are too thin to allow analysis by existing methods. The group developed new methods that allow the intravital analysis of flow and diffusion in extremely thin microvessels, in collaboration with the ImageTox group. Surprisingly, the researchers observed that solutes predominantly move by diffusion in the canaliculi, following a concentration gradient, and only in the interlobular bile ducts liquid flow sets in, leading to the novel diffusion-flow concept of bile flux (Vartak et al., 2021a,b). Importantly, this concept implies that the diffusion dominated tissue zone of canaliculi is particularly susceptible when the bile-barrier becomes leaky, which may have important consequences for intoxications that compromise the integrity of this barrier.
To study the vulnerability of the diffusion-dominated tissue zone of the liver, the group developed methods to characterize the bile-blood barrier (Ghallab et al., 2022) and identified a so far unrecognised mechanism of hepatotoxicity in the diffusion-dominated canalicular domain: Oxidative stress inducing compounds, for example paracetamol compromise the tight junctions of bile canaliculi, leading to leakage of bile acids into the sinusoidal blood, followed by uptake into the hepatocytes, secretion into canaliculi and repeated cycling.
This, ‘futile cycling’ of bile acids leads to increased intracellular bile acid concentrations, high enough to cause cell death. Importantly, the pharmacologic interruption of ‘futile cycling’ strongly reduced paracetamol induced hepatotoxicity and represents a therapeutic option even after the time window, where standard therapy is possible (Ghallab et al., 2022). The technique was developed further to additionally allow the intravital analysis of the barrier of cholangiocytes in the ducts and the analysis of gut-liver interactions, showing that the modification of bile acid concentration and composition by altered gut microbiota influences the degree by which bile acids leak into the peri-ductular space to promote cholangitis (Schneider et al., 2021, 2023).
Extending the intravital imaging for functional analysis of cell death, an unexpected key result was that concomitant necrosome and NF-κB activation in hepatocytes forced hepatocytes into a prolonged ‘sublethal’ state with leaky membranes but still intact mitochondrial metabolism, in which cells release DAMPs and cytokines that drive inflammation and carcinogenesis (Vucur et al., 2023). Since LPS is a critical cell death mediator in end-stage liver disease, the researchers established intravital two-photon imaging techniques that allow the direct observation, how sinusoidal lipopolysaccharide (LPS) is cleared by Kupffer cells (Remetic et al., 2022). Importantly, cholestasis leading to increased serum bile acids compromise this Kupffer cell function, thereby offering an explanation for the increased susceptibility to LPS in advanced liver disease.
The group leader of SysTox is chair of the Permanent Senate Commission on Food Safety (SKLM) of the Deutsche Forschungsgemeinschaft (DFG). One important activity is the establishment of a public database, where information on compounds including threshold values and their relationship to currently observed human exposure is documented.
In the applied research, the researchers addressed the limitations of current models to estimate hepatotoxic doses of compounds and predict drug-induced liver injury (DILI), developing novel in vitro/in silico techniques that allowed to successfully predict organ toxicity in relation to oral doses and blood concentrations (Albrecht et al., 2019). To address the limited availability of primary human hepatocytes, they established stem cell-based technologies to generate hepatocyte-like cells for toxicity testing (Nell et al., 2022), which is important since stem cells offer the perspective of an unlimited supply of hepatocytes.