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Background: Francisella tularensis, a gram-negative bacterium replicates intracellularly within macrophages and efficiently evades the innate immune response. It is able to infect and replicate within Kupffer cells, specialized tissue macrophages of the liver, and to modulate the immune response upon infection to its own advantage. Studies on Francisella tularensis liver infection were mostly performed in animal models and difficult to extrapolate to the human situation, since human infections and clinical observations are rare. Results: Using a human co-culture model of macrophages and hepatocytes we investigated the course of infection of three Francisella tularensis strains (subspecies holarctica – wildtype and live vaccine strain, and mediasiatica - wildtype) and analyzed the immune response triggered upon infection. We observed that hepatocytes support the intracellular replication of Franciscella species in macrophages accompanied by a specific immune response inducing TNFα, IL-1β, IL-6 and fractalkine (CX3CL1) secretion and the induction of apoptosis. Conclusions: We could demonstrate that this human macrophage / hepatocyte co-culture model reflects strain-specific virulence of Francisella tularensis. We developed a suitable tool for more detailed in vitro studies on the immune response upon liver cell infection by F. tularensis. [ABSTRACT FROM AUTHOR]

Pneumonia is the leading cause of hospitalization worldwide. Besides viruses, bacterial co-infections dramatically exacerbate infection. In general, surfactant protein-A (SP-A) represents a first line of immune defense. In this study, we analyzed whether influenza A virus (IAV) and/or Staphylococcus aureus (S. aureus) infections affect SP-A expression. To closely reflect the situation in the lung, we used a human alveolus-on-a-chip model and a murine pneumonia model. Our results show that S. aureus can reduce extracellular levels of SP-A, most likely attributed to bacterial proteases. Mono-epithelial cell culture experiments reveal that the expression of SP-A is not directly affected by IAV or S. aureus. Yet, the mRNA expression of SP-A is strongly down-regulated by TNF-α, which is highly produced by professional phagocytes in response to bacterial infection. By using the human alveolus-on-a-chip model, we show that the down-regulation of SP-A is strongly dependent on macrophages. In a murine model of pneumonia, we can confirm that S. aureus decreases SP-A levels in vivo. These findings indicate that (I) complex interactions of epithelial and immune cells induce down-regulation of SP-A expression and (II) bacterial mono- and super-infections reduce SP-A expression in the lung, which might contribute to a severe outcome of bacterial pneumonia. [ABSTRACT FROM AUTHOR]

Harvesting cultivated macrophages for tissue engineering purposes by enzymatic digestion of cell adhesion molecules can potentially result in unintended activation, altered function, or behavior of these cells. Thermo-responsive polymer is a promising tool that allows for gentle macrophage detachment without artificial activation prior to subculture within engineered tissue constructs. We therefore characterized different species of thermo-responsive polymers for their suitability as cell substrate and to mediate gentle macrophage detachment by temperature shift. Primary human monocyte- and THP-1-derived macrophages were cultured on thermo-responsive polymers and characterized for phagocytosis and cytokine secretion in response to lipopolysaccharide stimulation. We found that both cell types differentially respond in dependence of culture and stimulation on thermo-responsive polymers. In contrast to THP-1 macrophages, primary monocyte–derived macrophages showed no signs of impaired viability, artificial activation, or altered functionality due to culture on thermo-responsive polymers compared to conventional cell culture. Our study demonstrates that along with commercially available UpCell carriers, two other thermo-responsive polymers based on poly(vinyl methyl ether) blends are attractive candidates for differentiation and gentle detachment of primary monocyte–derived macrophages. In summary, we observed similar functionality and viability of primary monocyte–derived macrophages cultured on thermo-responsive polymers compared to standard cell culture surfaces. While this first generation of custom-made thermo-responsive polymers does not yet outperform standard culture approaches, our results are very promising and provide the basis for exploiting the unique advantages offered by custom-made thermo-responsive polymers to further improve macrophage culture and recovery in the future, including the covalent binding of signaling molecules and the reduction of centrifugation and washing steps. Optimizing these and other benefits of thermo-responsive polymers could greatly improve the culture of macrophages for tissue engineering applications. [ABSTRACT FROM AUTHOR]

Within the liver, non-parenchymal cells (NPCs) are critically involved in the regulation of hepatocyte polarization and maintenance of metabolic function. We here report the establishment of a liver organoid that integrates NPCs in a vascular layer composed of endothelial cells and tissue macrophages and a hepatic layer comprising stellate cells co-cultured with hepatocytes. The three-dimensional liver organoid is embedded in a microfluidically perfused biochip that enables sufficient nutrition supply and resembles morphological aspects of the human liver sinusoid. It utilizes a suspended membrane as a cell substrate mimicking the space of Disse. Luminescence-based sensor spots were integrated into the chip to allow online measurement of cellular oxygen consumption. Application of microfluidic flow induces defined expression of ZO-1, transferrin, ASGPR-1 along with an increased expression of MRP-2 transporter protein within the liver organoids. Moreover, perfusion was accompanied by an increased hepatobiliary secretion of 5(6)-carboxy-2′,7′-dichlorofluorescein and an enhanced formation of hepatocyte microvilli. From this we conclude that the perfused liver organoid shares relevant morphological and functional characteristics with the human liver and represents a new in vitro research tool to study human hepatocellular physiology at the cellular level under conditions close to the physiological situation. [ABSTRACT FROM AUTHOR]

Infections with Staphylococcus aureus (S. aureus) have been reported from various organs ranging from asymptomatic colonization to severe infections and sepsis. Although considered an extracellular pathogen, S. aureus can invade and persist in professional phagocytes such as monocytes and macrophages. Its capability to persist and manipulate macrophages is considered a critical step to evade host antimicrobial reactions. We leveraged a recently established human liver-on-chip model to demonstrate that S. aureus specifically targets macrophages as essential niche facilitating bacterial persistence and phenotype switching to small colony variants (SCVs). In vitro, M2 polarization was found to favor SCV-formation and was associated with increased intracellular bacterial loads in macrophages, increased cell death, and impaired recruitment of circulating monocytes to sites of infection. These findings expand the knowledge about macrophage activation in the liver and its impact on bacterial persistence and dissemination in the course of infection. [ABSTRACT FROM AUTHOR]