Partnerzy

Streszczenie:
Epithelial folding is a fundamental process where initially flat monolayers transform into functional 3D structures. This protocol details fabrication steps for a polycarbonate microfluidic platform which enables triggering epithelial folds that recapitulate stereotypical cell shape changes and folding-associated mechanical stresses. We describe the steps for cell seeding to form a monolayer on the chip, and subsequent approach to trigger calcium waves in the epithelial monolayer through local epithelial deformation. Lastly, we outline quantitative analysis steps of the epithelial response.

Streszczenie:
Cell shape dynamics during development is tightly regulated and coordinated with cell fate determination. Triggered by an interplay between biochemical and mechanical signals, epithelia form complex tissues by undergoing coordinated cell shape changes, but how such spatiotemporal coordination is controlled remains an open question. To dissect biochemical signaling from purely mechanical cues, we developed a microfluidic system that experimentally triggers epithelial folding to recapitulate stereotypic deformations observed in vivo. Using this system, we observe that the apical or basal direction of folding results in strikingly different mechanical states at the fold boundary, where the balance between tissue tension and torque (arising from the imposed curvature) controls the spread of folding-induced calcium waves at a short timescale and induces spatial patterns of gene expression at longer timescales. Our work uncovers that folding-associated gradients of cell shape and their resulting mechanical stresses direct spatially distinct biochemical responses within the monolayer.

Słowa kluczowe:
epithelial morphogenesis, epithelial folding, tension, calcium waves, microfluidics, RNAseq