Inflammation-on-a-chip: a microphysiological human liver mode

Microphysiological organ-on-a-chip models enter the stage as an enhanced in vitro method to emulate physiological processes ex vivo. This approach offers new possibilities to study systemic inflammatory diseases like sepsis and could be a promising tool to investigate the pathophysiological role of organs, such as the liver, during inflammation under humanized conditions. The objective of this work was the establishment of a human liver-on-a-chip and its application as a model of inflammatory hepatic dysfunction. In the first study, four major cell types of the liver were integrated in the Multi Organ Tissue Flow (MOTiF) biochip and examined by functional as well as morphological characteristics. To verify the applicability of the liver-on-a-chip model as a new biomedical research tool we investigated toll-like receptor (TLR)-mediated inflammation. Interestingly, hepatic damage was prevented by integration of circulating monocytes, which resulted in inflammatory resolution. To improve utilization of organ-on-a-chip applications we developed novel protocols for hypothermic storage of liver-on-a-chip models to maintain their microstructure after assembly. The study confirmed that the preservation of the liver model in an adapted formulation of the TiProtec® storage solution is possible for up to two days at 4 °C. Our hepatic model featured characteristics of human tissue samples and clinical observations during liver inflammation. This model has the potential to become a valuable alternative to overcome the limitations in translational research and will help to close the gap between conventional cell culture and animal experimentation

Functional integration of natural killer cells in a microfluidically perfused liver on-a-chip model.

OBJECTIVE The liver acts as an innate immunity-dominant organ and natural killer (NK) cells, are the main lymphocyte population in the human liver. NK cells are in close interaction with other immune cells, acting as the first line of defense against pathogens, infections, and injury. A previously developed, three-dimensional, perfused liver-on-a-chip comprised of human cells was used to integrate NK cells, representing pivotal immune cells during liver injury and regeneration. The objective of this study was to integrate functional NK cells in an in vitro model of the human liver and assess utilization of the model for NK cell-dependent studies of liver inflammation. RESULTS NK cells from human blood and liver specimen were isolated by Percoll separation with subsequent magnetic cell separation (MACS), yielding highly purified blood and liver derived NK cells. After stimulation with toll-like-receptor (TLR) agonists (lipopolysaccharides, Pam3CSK4), isolated NK cells showed increased interferon (IFN)-gamma secretion. To study the role of NK cells in a complex hepatic environment, these cells were integrated in the vascular compartment of a microfluidically supported liver-on-a-chip model in close interaction with endothelial and resident macrophages. Successful, functional integration of NK cells was verified by immunofluorescence staining (NKp46), flow cytometry analysis and TLR agonist-dependent secretion of interleukin (IL)-6 and tumor necrosis factor (TNF)-alpha. Lastly, we observed that inflammatory activation of NK cells in the liver-on-a-chip led to a loss of vascular barrier integrity. Overall, our data shows the first successful, functional integration of NK cells in a liver-on-a-chip model that can be utilized to investigate NK cell-dependent effects on liver inflammation in vitro

CAAP48, a New Sepsis Biomarker, Induces Hepatic Dysfunction in an in vitro Liver-on-Chip Model

Sepsis is a leading cause of mortality in the critically ill, characterized by life-threatening organ dysfunctions due to dysregulation of the host response to infection. Using mass spectrometry, we identified a C-terminal fragment of alpha-1-antitrypsin, designated CAAP48, as a new sepsis biomarker that actively participates in the pathophysiology of sepsis. It is well-known that liver dysfunction is an early event in sepsis-associated multi-organ failure, thus we analyzed the pathophysiological function of CAAP48 in a microfluidic-supported in vitro liver-on-chip model. Hepatocytes were stimulated with synthetic CAAP48 and several control peptides. CAAP48-treatment resulted in an accumulation of the hepatocyte-specific intracellular enzymes aspartate- and alanine-transaminase and impaired the activity of the hepatic multidrug resistant-associated protein 2 and cytochrome P450 3A4. Moreover, CAAP48 reduced hepatic expression of the multidrug resistant-associated protein 2 and disrupted the endothelial structural integrity as demonstrated by reduced expression of VE-cadherin, F-actin and alteration of the tight junction protein zonula occludens-1, which resulted in a loss of the endothelial barrier function. Furthermore, CAAP48 induced the release of adhesion molecules and pro- and anti-inflammatory cytokines. Our results show that CAAP48 triggers inflammation-related endothelial barrier disruption as well as hepatocellular dysfunction in a liver-on-chip model emulating the pathophysiological conditions of inflammation. Besides its function as new sepsis biomarker, CAAP48 thus might play an important role in the development of liver dysfunction as a consequence of the dysregulated host immune-inflammatory response in sepsis

Human iPSC‐Derived Proinflammatory Macrophages cause Insulin Resistance in an Isogenic White Adipose Tissue Microphysiological System

Chronic white adipose tissue (WAT) inflammation has been recognized as a critical early event in the pathogenesis of obesity-related disorders. This process is characterized by the increased residency of proinflammatory M1 macrophages in WAT. However, the lack of an isogenic human macrophage-adipocyte model has limited biological studies and drug discovery efforts, highlighting the need for human stem cell-based approaches. Here, human induced pluripotent stem cell (iPSC) derived macrophages (iMACs) and adipocytes (iADIPOs) are cocultured in a microphysiological system (MPS). iMACs migrate toward and infiltrate into the 3D iADIPOs cluster to form crown-like structures (CLSs)-like morphology around damaged iADIPOs, recreating classic histological features of WAT inflammation seen in obesity. Significantly more CLS-like morphologies formed in aged and palmitic acid-treated iMAC-iADIPO-MPS, showing the ability to mimic inflammatory severity. Importantly, M1 (proinflammatory) but not M2 (tissue repair) iMACs induced insulin resistance and dysregulated lipolysis in iADIPOs. Both RNAseq and cytokines analyses revealed a reciprocal proinflammatory loop in the interactions of M1 iMACs and iADIPOs. This iMAC-iADIPO-MPS thus successfully recreates pathological conditions of chronically inflamed human WAT, opening a door to study the dynamic inflammatory progression and identify clinically relevant therapies

Opportunities and challenges for combining chemo- and biocatalysis

Rudroff F, Mihovilovic MD, Gröger H, Snajdrova R, Iding H, Bornscheuer UT. Opportunities and challenges for combining chemo- and biocatalysis. NATURE CATALYSIS. 2018;1(1):12-22.The past decade has seen a substantial increase in successful examples of the combination of chemo-and biocatalysis for multistep syntheses. This is driven by obvious advantages such as higher yields, decreased costs, environmental benefits and high selectivity. On the downside, efforts must be undertaken to combine the divergent reaction conditions, reagent tolerance and solvent systems of these 'different worlds of catalysis'. Owing to progress in enzyme discovery and engineering, as well as in the development of milder and more compatible conditions for operating with various chemocatalysts, many historical limitations can already be overcome. This Review highlights the opportunities available in the chemical space of combined syntheses using prominent examples, but also discusses the current challenges and emerging solutions, keeping in mind the fast progress in transition metal-, organo-, photo-, electro-, hetero- and biocatalysis