Multi-compartment 3D-cultured organ-on-a-chip: Towards a biomimetic lymph node for drug development

The interaction of immune cells with drugs and/or with other cell types should be mechanistically investigated in order to reduce attrition of new drug development. However, they are currently only limited technologies that address this need. In our work, we developed initial but significant building blocks that enable such immune-drug studies. We developed a novel microfluidic platform replicating the Lymph Node (LN) microenvironment called LN-on-a-chip, starting from design all the way to microfabrication, characterization and validation in terms of architectural features, fluidics, cytocompatibility, and usability. To prove the biomimetics of this microenvironment, we inserted different immune cell types in a microfluidic device, which showed an in-vivo-like spatial distribution. We demonstrated that the developed LN-on-a-chip incorporates key features of the native human LN, namely, (i) similarity in extracellular matrix composition, morphology, porosity, stiffness, and permeability, (ii) compartmentalization of immune cells within distinct structural domains, (iii) replication of the lymphatic fluid flow pattern, (iv) viability of encapsulated cells in collagen over the typical timeframe of immunotoxicity experiments, and (v) interaction among different cell types across chamber boundaries. Further studies with this platform may assess the immune cell function as a step forward to disclose the effects of pharmaceutics to downstream immunology in more physiologically relevant microenvironments

Alterations in Cell Mechanics by Actin Cytoskeletal Changes Correlate with Strain-Specific Rubella Virus Phenotypes for Cell Migration and Induction of Apoptosis

The cellular cytoskeleton is central for key cellular functions, and as such is a marker for diseased and infected cell states. Here we analyzed infection with rubella virus (RV) strains with respect to phenotypes in cellular mechanical properties, cell movement, and viral cytopathogenicity. Real-time deformability cytometry (RT-DC), as a high-throughput platform for the assessment of cell mechanics, revealed a correlation of an increase in cortical filamentous-actin (F-actin) with a higher cellular stiffness. The additional reduction of stress fibers noted for only some RV strains as the most severe actin rearrangement lowered cell stiffness. Furthermore, a reduced collective and single cell migration speed in a wound healing assay was detected in addition to severe changes in cell morphology. The latter was followed by activation of caspase 3/7 as a sign for induction of apoptosis. Our study emphasizes RT-DC technology as a sensitive means to characterize viral cell populations and to implicate alterations of cell mechanical properties with cell functions. These interdependent events are not only promising options to elucidate viral spread and to understand viral pathologies within the infected host. They also contribute to any diseased cell state, as exemplified by RV as a representative agent for cytoskeletal alterations involved in a cytopathological outcome

Hydrogels for Engineering the Immune System

Human immune system has evolved as one of the most powerful defense systems to protect against invading pathogens and mutated cells. However, when persistent immune suppression or activation occurs, it can lead to adverse, chronic physiological effects including cancer and arthritis. Hydrogels are soft materials that can be engineered to modulate immune responses through controlled biomolecule release/adsorption, regeneration of lymphoid tissues, and enhanced antigen presentations. This is achieved by programming hydrogels to exhibit optimal properties such as porosity, biodegradability, and biocompatibility to interface seamlessly with the immune system. Herein, recent innovations and future challenges are described using programmable hydrogels to regenerate the lymphatic system, modulate inflammation, and enhance cancer immunotherapy. Key properties of hydrogels are also highlighted for engineering the immune system and techniques to characterize these properties

Molecular weight of surface immobilized hyaluronic acid infuences CD44-mediated binding of gastric cancer cells

The physiological importance of the interactions between hyaluronic acid (HA) and its main membrane receptor, CD44, in pathological processes, e.g. cancer, is well recognized. However, these interactions are mainly studied in solution, whereas HA in the extracellular matrix (ECM) is partially immobilized via its interactions with other ECM components. We therefore, developed substrates in which HA is presented in an ECM-relevant manner. We immobilized HA with different molecular weights (Mw) in a Layer-by-Layer (LbL) fashion and studied the interactions of the substrates with CD44 and two human gastric cancer cell lines that overexpress this receptor, namely AGS and MKN45. We demonstrate that MKN45 cells are more sensitive to the LbL substrates as compared with AGS. This difference is due to different CD44 expression: while CD44 is detected mainly in the cytoplasm of AGS, MKN45 express CD44 predominantly at the cell membrane where it is involved in the recognition and binding of HA. The invasiveness of the studied cell lines was also evaluated as a function of HA Mw. Invasive profile characterized by low cell adhesion, high cell motility, high expression of cortactin, formation of invadopodia and cell clusters was observed for MKN45 cells when they are in contact with substrates presenting HA of high Mw.Te authors acknowledge the fnancial support from the European Commission’s H2020 programme, under grant agreements H2020-TWINN-2015-692333-CHEM2NATURE, H2020-WIDESPREAD-2014-668983- FORECAST and H2020-WIDESPREAD-01-2016-2017-739572-THE DISCOVERIES CTR. S.A., D.S.C. and I.P. also acknowledge the Portuguese Foundation for Science and Technology (FCT) for fnancial support under grants SFRH/BD/112075/2015, SFRH/BPD/85790/2012 and IF/00032/2013, respectively.info:eu-repo/semantics/publishedVersio