Using Rapid Prototyping to Develop a Cell-Based Platform with Electrical Impedance Sensor Membranes for In Vitro RPMI2650 Nasal Nanotoxicology Monitoring.

Due to advances in additive manufacturing and prototyping, affordable and rapid microfluidic sensor-integrated assays can be fabricated using additive manufacturing, xurography and electrode shadow masking to create versatile platform technologies aimed toward qualitative assessment of acute cytotoxic or cytolytic events using stand-alone biochip platforms in the context of environmental risk assessment. In the current study, we established a nasal mucosa biosensing platform using RPMI2650 mucosa cells inside a membrane-integrated impedance-sensing biochip using exclusively rapid prototyping technologies. In a final proof-of-concept, we applied this biosensing platform to create human cell models of nasal mucosa for monitoring the acute cytotoxic effect of zinc oxide reference nanoparticles. Our data generated with the biochip platform successfully monitored the acute toxicity and cytolytic activity of 6 mM zinc oxide nanoparticles, which was non-invasively monitored as a negative impedance slope on nasal epithelial models, demonstrating the feasibility of rapid prototyping technologies such as additive manufacturing and xurography for cell-based platform development.

The Cultivation Modality and Barrier Maturity Modulate the Toxicity of Industrial Zinc Oxide and Titanium Dioxide Nanoparticles on Nasal, Buccal, Bronchial, and Alveolar Mucosa Cell-Derived Barrier Models.

As common industrial by-products, airborne engineered nanomaterials are considered important environmental toxins to monitor due to their potential health risks to humans and animals. The main uptake routes of airborne nanoparticles are nasal and/or oral inhalation, which are known to enable the transfer of nanomaterials into the bloodstream resulting in the rapid distribution throughout the human body. Consequently, mucosal barriers present in the nose, buccal, and lung have been identified and intensively studied as the key tissue barrier to nanoparticle translocation. Despite decades of research, surprisingly little is known about the differences among various mucosa tissue types to tolerate nanoparticle exposures. One limitation in comparing nanotoxicological data sets can be linked to a lack of harmonization and standardization of cell-based assays, where (a) different cultivation conditions such as an air-liquid interface or submerged cultures, (b) varying barrier maturity, and (c) diverse media substitutes have been used. The current comparative nanotoxicological study, therefore, aims at analyzing the toxic effects of nanomaterials on four human mucosa barrier models including nasal (RPMI2650), buccal (TR146), alveolar (A549), and bronchial (Calu-3) mucosal cell lines to better understand the modulating effects of tissue maturity, cultivation conditions, and tissue type using standard transwell cultivations at liquid-liquid and air-liquid interfaces. Overall, cell size, confluency, tight junction localization, and cell viability as well as barrier formation using 50% and 100% confluency was monitored using trans-epithelial-electrical resistance (TEER) measurements and resazurin-based Presto Blue assays of immature (e.g., 5 days) and mature (e.g., 22 days) cultures in the presence and absence of corticosteroids such as hydrocortisone. Results of our study show that cellular viability in response to increasing nanoparticle exposure scenarios is highly compound and cell-type specific (TR146 6 ± 0.7% at 2 mM ZnO (ZnO) vs. ~90% at 2 mM TiO2 (TiO2) for 24 h; Calu3 93.9 ± 4.21% at 2 mM ZnO vs. ~100% at 2 mM TiO2). Nanoparticle-induced cytotoxic effects under air-liquid cultivation conditions declined in RPMI2650, A549, TR146, and Calu-3 cells (~0.7 to ~0.2-fold), with increasing 50 to 100% barrier maturity under the influence of ZnO (2 mM). Cell viability in early and late mucosa barriers where hardly influenced by TiO2 as well as most cell types did not fall below 77% viability when added to Individual ALI cultures. Fully maturated bronchial mucosal cell barrier models cultivated under ALI conditions showed less tolerance to acute ZnO nanoparticle exposures (~50% remaining viability at 2 mM ZnO for 24 h) than the similarly treated but more robust nasal (~74%), buccal (~73%), and alveolar (~82%) cell-based models.

A Progress Report and Roadmap for Microphysiological Systems and Organ-On-A-Chip Technologies to Be More Predictive Models in Human (Knee) Osteoarthritis.

Osteoarthritis (OA), a chronic debilitating joint disease affecting hundreds of million people globally, is associated with significant pain and socioeconomic costs. Current treatment modalities are palliative and unable to stop the progressive degeneration of articular cartilage in OA. Scientific attention has shifted from the historical view of OA as a wear-and-tear cartilage disorder to its recognition as a whole-joint disease, highlighting the contribution of other knee joint tissues in OA pathogenesis. Despite much progress in the field of microfluidic systems/organs-on-a-chip in other research fields, current in vitro models in use do not yet accurately reflect the complexity of the OA pathophenotype. In this review, we provide: 1) a detailed overview of the most significant recent developments in the field of microsystems approaches for OA modeling, and 2) an OA-pathophysiology-based bioengineering roadmap for the requirements of the next generation of more predictive and authentic microscale systems fit for the purpose of not only disease modeling but also of drug screening to potentially allow OA animal model reduction and replacement in the near future.

Recent Advances in Additive Manufacturing and 3D Bioprinting for Organs-On-A-Chip and Microphysiological Systems.

The re-creation of physiological cellular microenvironments that truly resemble complex in vivo architectures is the key aspect in the development of advanced in vitro organotypic tissue constructs. Among others, organ-on-a-chip technology has been increasingly used in recent years to create improved models for organs and tissues in human health and disease, because of its ability to provide spatio-temporal control over soluble cues, biophysical signals and biomechanical forces necessary to maintain proper organotypic functions. While media supply and waste removal are controlled by microfluidic channel by a network the formation of tissue-like architectures in designated micro-structured hydrogel compartments is commonly achieved by cellular self-assembly and intrinsic biological reorganization mechanisms. The recent combination of organ-on-a-chip technology with three-dimensional (3D) bioprinting and additive manufacturing techniques allows for an unprecedented control over tissue structures with the ability to also generate anisotropic constructs as often seen in in vivo tissue architectures. This review highlights progress made in bioprinting applications for organ-on-a-chip technology, and discusses synergies and limitations between organ-on-a-chip technology and 3D bioprinting in the creation of next generation biomimetic in vitro tissue models.

Establishment of a human three-dimensional chip-based chondro-synovial coculture joint model for reciprocal cross talk studies in arthritis research.

Rheumatoid arthritis is characterised by a progressive, intermittent inflammation at the synovial membrane, which ultimately leads to the destruction of the synovial joint. The synovial membrane as the joint capsule's inner layer is lined with fibroblast-like synoviocytes that are the key player supporting persistent arthritis leading to bone erosion and cartilage destruction. While microfluidic models that model molecular aspects of bone erosion between bone-derived cells and synoviocytes have been established, RA's synovial-chondral axis has not yet been realised using a microfluidic 3D model based on human patient in vitro cultures. Consequently, we established a chip-based three-dimensional tissue coculture model that simulates the reciprocal cross talk between individual synovial and chondral organoids. When co-cultivated with synovial organoids, we could demonstrate that chondral organoids induce a higher degree of cartilage physiology and architecture and show differential cytokine response compared to their respective monocultures highlighting the importance of reciprocal tissue-level cross talk in the modelling of arthritic diseases.