Viktoriia Starokozhko

Human Precision-Cut Liver Slices as an ex Vivo Model to Study Idiosyncratic Drug-Induced Liver Injury

Idiosyncratic drug-induced liver injury (IDILI) is a major problem during drug development and has caused drug withdrawal and black-box warnings. Because of the low concordance of the hepatotoxicity of drugs in animals and humans, robust screening methods using human tissue are needed to predict IDILI in humans. According to the inflammatory stress hypothesis, the effects of inflammation interact with the effects of a drug or its reactive metabolite, precipitating toxic reactions in the liver. As a follow-up to our recently published mouse precision-cut liver slices model, an ex vivo model involving human precision-cut liver slices (hPCLS), co-incubated for 24 h with IDILI-related drugs and lipopolysaccharide (LPS), was developed to study IDILI mechanisms related to inflammatory stress in humans and to detect potential biomarkers. LPS exacerbated the effects of ketoconazole and clozapine toxicity but not those of their non-IDILI-related comparators, voriconazole and olanzapine. However, the IDILI-related drugs diclofenac, carbamazepine, and troglitazone did not show synergistic toxicity with LPS after incubation for 24 h. Co-incubation of ketoconazole and clozapine with LPS decreased the levels of glutathione in hPCLS, but this was not seen for the other drugs. All drugs affected LPS-induced cytokine release, but interestingly, only ketoconazole and clozapine increased the level of LPS-induced TNF release. Decreased levels of glutathione and cysteine conjugates of clozapine were detected in IDILI-responding livers following cotreatment with LPS. In conclusion, we identified ketoconazole and clozapine as drugs that exhibited synergistic toxicity with LPS, while glutathione and TNF were found to be potential biomarkers for IDILI-inducing drugs mediated by inflammatory stress. hPCLS appear to be suitable for further unraveling the mechanisms of inflammatory stress-associated IDILI.

Judging the value of ‘liver-on-a-chip’ devices for prediction of toxicity

Despite a continuous increase in the application of in vitro tests for absorption, metabolism, distribution and excretion (ADME) and toxicity testing, conventional in vitro models have not been powerful enough to fully replace animal models to predict toxicity in human. In general, they lack sufficient functionality and predictivity for human ADME and toxicity. Organ-on-a-chip technologies have been proposed as a new generation of in vitro models for drug candidate screening in the preclinical phase of drug development. Most ADME and toxicity in vitro tests are liver-based as the liver is the most important organ in xenobiotic metabolism and is one of the major targets for drugand chemical-induced toxicity. Liver-on-a-chip models, defined here as three-dimensional (3D) microfluidic cell or tissue culture systems that aim tomimic closely the anatomy, physiology, and functionality of the liver, are gaining increasing attention as in vitro alternative for in vivo testing. The chips onwhich cells are seeded aremostly made fromoptically transparent polymers and usually contain channels the size of which ranges between 50 and 500 μm. They can contain monocultures of hepatocytes, twodimensional or 3D co-cultures of hepatocytes with several other hepatic non-parenchymal cells (NPC) or other stromal cells, hepatocyte spheroids, or organoids formed by monoor co-cultures and intact tissue such as precision-cut liver slices (PCLS).

Differentiation of human-induced pluripotent stem cell under flow conditions to mature hepatocytes for liver tissue engineering

Hepatic differentiation of human‐induced pluripotent stem cells (hiPSCs) under flow conditions in a 3D scaffold is expected to be a major step forward for construction of bioartificial livers. The aims of this study were to induce hepatic differentiation of hiPSCs under perfusion conditions and to perform functional comparisons with fresh human precision‐cut liver slices (hPCLS), an excellent benchmark for the human liver in vivo. The majority of the mRNA expression of CYP isoenzymes and transporters and the tested CYP activities, Phase II metabolism, and albumin, urea, and bile acid synthesis in the hiPSC‐derived cells reached values that overlap those of hPCLS, which indicates a higher degree of hepatic differentiation than observed until now. Differentiation under flow compared with static conditions had a strong inducing effect on Phase II metabolism and suppressed AFP expression but resulted in slightly lower activity of some of the Phase I metabolism enzymes. Gene expression data indicate that hiPSCs differentiated into both hepatic and biliary directions. In conclusion, the hiPSC differentiated under flow conditions towards hepatocytes express a wide spectrum of liver functions at levels comparable with hPCLS indicating excellent future perspectives for the development of a bioartificial liver system for toxicity testing or as liver support device for patients.

Challenges on the road to a multicellular bioartificial liver

Over recent years, the progress in the development of a bioartificial liver (BAL) as an extracorporeal device or as a tissue engineered transplantable organ has been immense. However, many important BAL characteristics that are necessary to meet clinical demands have not been sufficiently addressed. This review describes the existing challenges in the development of a BAL for clinical applications, highlighting multicellularity and sinusoidal microarchitecture as crucial BAL characteristics to fulfil various liver functions. Currently available sources of nonparenchymal liver cells, such as endothelial cells, cholangiocytes and macrophages, used in BAL development are defined. Also, we discuss the recent studies on the reconstruction of the complex liver sinusoid microarchitecture using various liver cell types. Moreover, we highlight some other aspects of a BAL, such as liver zonation and formation of a vascular as well as biliary network for an adequate delivery, biotransformation and removal of substrates and waste products. Finally, the benefits of a multicellular BAL for the pharmaceutical industry are briefly described. Copyright

How microtechnologies enable organs-on-a-chip

Engineering cellular microenvironments that more accurately reflect the in vivo situation is now recognized as being crucial for the improvement of the in vitro viability and in vivo-like function of cells or tissues. Microfluidic technologies have been increasingly applied since the late 1990s for this purpose, with a growing number of examples of perfused cell and tissue cultures in microfluidic chambers and channels. More recently, additional microfabricated features have been implemented in microfluidic structures to achieve 3-D cell culture systems which mimic not only in vivo fluid flows, but also the structure, transport, and mechanical properties of tissue in, for example, the lung or the intestine. The ultimate challenge becomes the combination of different organ functions into single, linked-compartment devices - the body-on-the-chip.