Biomimetic fractal topography enhances podocyte maturation in vitro

Cells and tissues in their native environment are organized into intricate fractal structures, which are rarely recapitulated in their culture in vitro. The extent to which fractal patterns that resemble complex topography in vivo influence cell maturation, and the cellular responses to such shape stimulation remain inadequately elucidated. Yet, the application of fractal cues (topographical stimulation via self-similar patterns) as an external input may offer a much-needed solution to the challenge of improving the differentiated cell phenotype in vitro. Here, we established fractality in the kidney glomerulus and podocytes, branching highly differentiated kidney cells within the glomerulus. Biomimetic fractal patterns derived from glomerular histology were used to generate topographical (2.5-D) substrates for cell culture. Podocytes grown on fractal topography expressed higher levels of functional markers and exhibited enhanced cell polarity. To track morphological complexities of differentiated podocytes, we employed a fluorescent labelling assay where labelled individual cells are tracked within otherwise optically silent confluent cell monolayer to reveal single-cell borders. RNAseq analysis suggested enhanced ECM deposition and remodeling in podocytes grown on fractal substrates versus flat surfaces, and enhanced maturation accompanied by stress in podocytes grown on fractal versus non-fractal topography. The incorporation of fractal topography into standard tissue culture well plates as demonstrated here may serve as a user-friendly bioengineered platform for high-fidelity cell culture. Overall design: RNA-seq profiling of E11 murine porodyctes cultured on polydimethylsiloxane (PDMS) substrates with No Topography (NT), Round Topography (RT), and High Fractal (HF) topography where NT and RT serve as non-fractal non-topographic control and non-fractal topographic control groups, respectively, and HF refers to a biomimetic high fractal topographic group derived from healthy histological glomerular tracing