ABSTRACT
Nanoparticle toxicity assessments have moved closer to physiological conditions while trying to avoid the use of animal models. An example of new in vitro exposure techniques developed is the exposure of cultured cells at the air-liquid interface (ALI), particularly in the case of respiratory airways. While the commercially available VITROCELL® Cloud System has been applied for the delivery of aerosolized substances to adherent cells under ALI conditions, it has not yet been tested on lung surfactant and semi-adherent cells such as alveolar macrophages, which are playing a pivotal role in the nanoparticle-induced immune response. OBJECTIVES: In this work, we developed a comprehensive methodology for coating semi-adherent lung cells cultured at the ALI with aerosolized surfactant and subsequent dose-controlled exposure to nanoparticles (NPs). This protocol is optimized for subsequent transcriptomic studies. METHODS: Semi-adherent rat alveolar macrophages NR8383 were grown at the ALI and coated with lung surfactant through nebulization using the VITROCELL® Cloud 6 System before being exposed to TiO2 NM105 NPs. After NP exposures, RNA was extracted and its quantity and quality were measured. RESULTS: The VITROCELL® Cloud system allowed for uniform and ultrathin coating of cells with aerosolized surfactant mimicking physiological conditions in the lung. While nebulization of 57 µL of 30 mg/mL TiO2 and 114 µL of 15 mg/mL TiO2 nanoparticles yielded identical cell delivered dose, the reproducibility of dose as well as the quality of RNA extracted were better for 114 µL.
ABSTRACT
Hyaline cartilage is deficient in self-healing properties. The early treatment of focal cartilage lesions is a public health challenge to prevent long-term degradation and the occurrence of osteoarthritis. Cartilage tissue engineering represents a promising alternative to the current insufficient surgical solutions. 3D printing is a thriving technology and offers new possibilities for personalized regenerative medicine. Extrusion-based processes permit the deposition of cell-seeded bioinks, in a layer-by-layer manner, allowing mimicry of the native zonal organization of hyaline cartilage. Mesenchymal stem cells (MSCs) are a promising cell source for cartilage tissue engineering. Originally isolated from bone marrow, they can now be derived from many different cell sources (e.g., synovium, dental pulp, Wharton's jelly). Their proliferation and differentiation potential are well characterized, and they possess good chondrogenic potential, making them appropriate candidates for cartilage reconstruction. This review summarizes the different sources, origins, and densities of MSCs used in extrusion-based bioprinting (EBB) processes, as alternatives to chondrocytes. The different bioink constituents and their advantages for producing substitutes mimicking healthy hyaline cartilage is also discussed.
