3D Microtissues for Injectable Regenerative Therapy and High-throughput Drug Screening.

To upgrade traditional 2D cell culture to 3D cell culture, we have integrated microfabrication with cryogelation technology to produce macroporous microscale cryogels (microcryogels), which can be loaded with a variety of cell types to form 3D microtissues. Herein, we present the protocol to fabricate versatile 3D microtissues and their applications in regenerative therapy and drug screening. Size and shape-controllable microcryogels can be fabricated on an array chip, which can be harvested off-chip as individual cell-loaded carriers for injectable regenerative therapy or be further assembled on-chip into 3D microtissue arrays for high-throughput drug screening. Due to the high elastic nature of these microscale cryogels, the 3D microtissues exhibit great injectability for minimally invasive cell therapy by protecting cells from mechanical shear force during injection. This ensures enhanced cell survival and therapeutic effect in the mouse limb ischemia model. Meanwhile, assembly of 3D microtissue arrays in a standard 384-multi-well format facilitates the use of common laboratory facilities and equipment, enabling high-throughput drug screening on this versatile 3D cell culture platform.

Bi-content micro-collagen chip provides contractility-based biomechanical readout for phenotypic drug screening with expanded and profiled targets.

Phenotypic screening has regained momentum in the pharmaceutical industry owing to its success over target-based screening. Most phenotypic screening relies on nonspecific biochemical readouts regarding cellular viability, which hampers the discovery of novel drug mechanisms of action (MOAs). Here we present a Contractility-based bi-Content micro-Collagen Chip (3CChip), which establishes cellular contractility as a biomechanics-related phenotype for drug screening. Bi-content analysis of cell contractility (imaged by iPhone) and viability suggests that the label-free contractility-based analysis exhibits superior sensitivity to compounds targeting contractile elements (e.g. focal adhesion, cytoskeleton), resulting in a enlarged target pool for drug assessment. Six typical readout patterns of drug response are summarized according to the relative positions of the contraction/viability curves, and drug targets are profiled into three categories (biomechanical, biochemical and housekeeping) by 3CChip, which will benefit subsequent target identification. The simple-to-use and effective 3CChip offers a robust platform for micro-tissue-based functional screening and may lead to a new era of mechanism-informed phenotypic drug discovery.