Networked concave microwell arrays for constructing 3D cell spheroids

  The engineered three-dimensional (3D) cell cultivation system for the production ofmulticellular spheroids has attracted considerable attention due to its improved in vivo relevance to cellular communications compared with the traditional two-dimensional (2D) cell culture platform. The formation and maintenance of cell spheroids in a healthy condition is the critical factor for tissue engineering applications such as the repair of damaged tissues, the development of organ replacement parts and preclinical drug tests. However, culturing spheroids in conventional isolated single wells shows limited yield and reduced maintenance periods due to the lack of proper supplies of nutrition aswell as intercellular chemical signaling.Here, we develop novel networked concave microwell arrays for the effective construction of 3Dmulti-cellular spheroids. The proposed method provides a suitable structure for the diffusion of oxygen, water-soluble nutrients and cytokines for cell–cell interactions between the spheroids in neighboringmicrowells. Wehave further demonstrated that hepatocyte spheroid culturednetworkedconcavemicrowells show enhanced cell viability and albumin secretion compared to the un-networked control group over twoweeks.Our results reveal thatmulti-cellular functionality can be tuned up by networking individual 3D spheroidswithout supplying additional chemicals or biological supplements.We anticipate our result to be useful in high-throughput cellular screening platforms to study cell–cell interactions, in response to diverse chemical stimuli as well as the development of the in vivo mimicking of the customized 3D tissue culture system.