Topographically-patterned porous membranes in a microfluidic device as an in vitro model of renal reabsorptive barriers.

Models of reabsorptive barriers require both a means to provide realistic physiologic cues to and quantify transport across a layer of cells forming the barrier. Here we have topographically-patterned porous membranes with several user-defined pattern types. To demonstrate the utility of the patterned membranes, we selected one type of pattern and applied it to a membrane to serve as a cell culture support in a microfluidic model of a renal reabsorptive barrier. The topographic cues in the model resemble physiological cues found in vivo while the porous structure allows quantification of transport across the cell layer. Sub-micron surface topography generated via hot-embossing onto a track-etched polycarbonate membrane, fully replicated topographical features and preserved porous architecture. Pore size and shape were analyzed with SEM and image analysis to determine the effect of hot embossing on pore morphology. The membrane was assembled into a bilayer microfluidic device and a human kidney proximal tubule epithelial cell line (HK-2) and primary renal proximal tubule epithelial cells (RPTEC) were cultured to confluency on the membrane. Immunofluorescent staining of both cell types revealed protein expression indicative of the formation of a reabsorptive barrier responsive to mechanical stimulation: ZO-1 (tight junction), paxillin (focal adhesions) and acetylated α-tubulin (primary cilia). HK-2 and RPTEC aligned in the direction of ridge/groove topography of the membrane in the device, evidence that the device has mechanical control over cell response. This topographically-patterned porous membrane provides an in vitro platform on which to model reabsorptive barriers with meaningful applications for understanding biological transport phenomenon, underlying disease mechanisms, and drug toxicity.

The use of controlled surface topography and flow-induced shear stress to influence renal epithelial cell function.

Physiologically-representative and well-controlled in vitro models of human tissue provide a means to safely, accurately, and rapidly develop therapies for disease. Current in vitro models do not possess appropriate levels of cell function, resulting in an inaccurate representation of in vivo physiology. Mechanical parameters, such as sub-micron topography and flow-induced shear stress (FSS), influence cell functions such as alignment, migration, differentiation and phenotypic expression. Combining, and independently controlling, biomaterial surface topography and FSS in a cell culture device would provide a means to control cell function resulting in more physiologically-representative in vitro models of human tissue. Here we develop the Microscale Tissue Modeling Device (MTMD) which couples a topographically-patterned substrate with a microfluidic chamber to control both topographic and FSS cues to cells. Cells from the human renal proximal tubule cell line HK-2 were cultured in the MTMD and exposed to topographic patterns and several levels of FSS simultaneously. Results show that the biomaterial property of surface topography and FSS work in concert to elicit cell alignment and influence tight junction (TJ) formation, with topography enhancing cell response to FSS. By administering independently-controlled mechanical parameters to cell populations, the MTMD creates a more realistic in vitro model of human renal tissue.