Hypergravity-induced multicellular spheroid generation with different morphological patterns precisely controlled on a centrifugal microfluidic platform

Abstract

In living tissue, cells exist in three-dimensional (3D) microenvironments with intricate cell-cell interactions. To model these cellular environments, numerous techniques for generating cell spheroids have been proposed and improved. However, previously reported methods still have limitations in uniformity, reproducibility, scalability, throughput, etc. Here, we present a centrifugal microfluidic-based spheroid (CMS) formation method for generating both co-culture and mono-culture 3D spheroids in a highly controlled manner. Wedesigned circularly arrayed microwells to allow the even distribution of cells introduced at the center of a rotating platform and to provide identical hypergravity conditions at each well by the centrifugal forces generated. Compared with conventional well plate-based spheroid formation, the CMS formation method significantly promotes sphericity and consistency in both size and shape with high production yields. In addition to mono-culture spheroids, we successfully generated co-culture spheroids in concentric, Janus, and sandwich shapes using human adipose-derived stem cells and human lung fibroblasts, demonstrating the versatility of our CMS formation method. We believe that our new method for generating 3D spheroids will become one of the essential technologies in the field of 3D cell culture. We also expect that we are providing an innovative means to assess cellular responses, including cell motility under different hypergravity conditions.