Tight junction proteins seal the space between adjacent epithelial and endothelial cells and form paracellular pores that regulate the paracellular transport: pore-forming claudins. Unfortunately, biophysics of these pore-forming claudins remains incompletely undefined. Mutations in pore-forming claudins cause human disease, including familial hypomagnesemia and hypercalcuria, neonatal sclerosing cholangitis associated with ichthyosis, and others. Underlying structure-function relationships will fundamentally advance our understanding of paracellular permeability and may, ultimately, lead to novel therapeutic approaches. The Dynchan project aims to develop a new technology that will enable high resolution analyses of claudin channel activity and regulation. The technology based on silicon chips with array of nanopillar electrodes is expected to provide the tool needed to develop foundational understanding of claudin biology. During the Dynchan project, we established conditions for cultivating functional, mature, properly assembled epithelial monolayers on high aspect ratio pillars localized to lateral intercellular spaces. We found that the pillar size strongly affected the assembling of tight junctions. For MDCK I cells, pillar structures of 1 µm in diameter and approximately up to 6 µm in height facilitated the formation of tight junctions in close proximity with the tips of the pillar structures. Additionally, through the application of micro and nanotechnologies, we have successfully fabricated miniature electrodes atop these pillars. This breakthrough allows us to record electrical currents crossing the tight junctions with an very low spatial resolution in the one micron range.
