Porous membranes are frequently used as supports of cell monolayers in functional studies of epithelial and endothelial barriers. However, commonly used conventional polymer-based membranes such as those made of polycarbonate (transwells) do not mimic the structural and biochemical properties of native basement membranes, which may limit cellular differentiation and function. Here, we use a nanofibrillar membrane that mimics the microenvironment of the basement membrane and is made of recombinant spider silk functionalized with the integrin-binding RGD motif of fibronectin and coated with human kidney-specific laminin-521 (FN-silk). The FN-silk membranes were evaluated as culture substrates for renal epithelial cells (RPTEC/TERT1), a cell type notoriously difficult to differentiate in culture. FN-silk membrane structure, cellular morphology, mRNA expression, barrier properties and transporter activity were assessed using scanning and transmission electron microscopy, RNA-sequencing, permeability and transport assays. Both FN-silk and conventional membranes supported barrier integrity and tight junction expression. In contrast to cultures on conventional membranes, the RPTEC monolayers on FN-silk exhibited a differentiated morphology, low expression of cell death markers, and, directional anion and cation transport. Further, the commonly used conventional membranes released endocrine-disrupting bisphenols that activated estrogen-mediated signaling. In summary, these findings indicate that FN-silk membranes with kidney-specific basement membrane laminin 521 reduces cellular stress, supports and maintains cellular differentiation and preserves important cellular functions. Our work establishes FN-silk membranes as a next-generation biomaterial that enables the differentiation of renal epithelial monolayers into a physiologically relevantin vitromodel.
Meyer et al. (Wed,) studied this question.