Development of a microfluidic device to test nanoparticle toxicity

Recent years have seen a growth in the manufacturing of nanoparticles for their uses in various fields of science and technology. However, this explosion in the production and use of nanoparticles has in turn resulted in growing concerns regarding their impact on public health and the environment (Hoet, 2004). One major route of entry into the human body is through the air-blood barrier in the lungs. The air-blood barrier at the alveolar region in most mammals is normally about 500-600 nm in thickness (Bartels, 1979) and is mainly responsible for the selective transport of gases and certain vital solutes across the membrane (Theodore, 1975). This selective transport across these barriers is regulated by tight junction protein complexes that bind two adjacent cells in the tissue. This particular selective transport mechanism is highly attractive for the drugs industry due to which the lung epithelial barriers could provide a novel mode for delivery to patients (such a system already exist for patients suffering from asthama, where they use an inhaler to deliver their dosage). However, to develop such drug delivery systems it is necessary to study the effects either through in vivo and/or in vitro research methods. For the purpose of this thesis an in vitro system using the Calu-3 cell line (cultured on two types of membrane systems) was used in the attempt at mimicing certain barrier properties (mainly transport of solutes across the membrane and integrity/tightness of the cell monolayers) present in the in vivo state. Calu-3 cells were maintained on two different sets of porous membrane types, one was the commercially available Transwell® membranes (Costar/Fisher) and the other was the self-fabricated (at CSEM, Switzerland) silicon nitride membranes. The silicon nitride membranes were particularly unique, in the sense that their thickness was only 500nm (compared to the polymer Transwell® membranes) and also presented the possibility of miniaturising the Calu-3 in vitro system. Miniaturisation helps reduce the use of test solutions and allow the development of high throughput screening devices for biological applications (Beebe, 2002). When the possibility of miniaturisation occurs along side a biological application it is often the case that microflows would be necessary for maintaining cell culture within small areas inside the devices. Therefore, microfluidics was vital in providing the opportunity for miniaturised cell based systems. In this study PDMS (Polydimethyl siloxane) based microfluidic devices were used for developing the cell culture and concentration gradient devices. The final purpose of this poject was to create a scaleable modular integrated device allowing the analysis of the induced effects on Calu-3 cells against nanoparticle/solute translocation and assessing cell monolayer integrity using a real time TEER measurement system. This miniaturised in vitro multilayered microfluidic setup consisted of three main components, a top layer micro channel (fabricated in PDMS), the middle silicon wafer bearing the silicon nitride membrane (also bearing the electrodes for measuring TEER of the cultured cell monolayer) and the bottom layer micro channel. This modular device would help assess Calu-3 cell monolayers resposes to toxic solutions and hopefully assist towards developing a novel analysis system device to study the effects of such toxic solutions in real time.