Injectable therapeutic organoids using sacrificial hydrogels

Abstract

<jats:title>Abstract</jats:title><jats:p>Organoids, by promoting self-organization of cells into native-like structures, are becoming widespread in drug-screening technologies, but have so far been used sparingly for cell therapy as current approaches for producing self-organized cell clusters lack scalability or reproducibility in size and cellular organization. We introduce a method of using hydrogels as sacrificial scaffolds, which allow cells to form self-organized clusters followed by gentle release, resulting in highly reproducible multicellular structures on a large scale. We demonstrated this strategy for endothelial cells and mesenchymal stem cells to self-organize into blood-vessel units, which were injected into mice using hypodermic needles, and observed in real time to rapidly form perfusing vasculature. As cell therapy transforms into a new class of therapeutic modality, this simple method – by making use of the dynamic nature of hydrogels – could offer high yields of self-organized multicellular aggregates with reproducible sizes and cellular architectures.</jats:p>

Towards organoid culture without Matrigel

“…In addition to organoids grown in naturally-derived proteins, several laboratories have grown a wide range of organoids in polysaccharides such as alginate or alginate-chitosan mixtures. Organoid types grown in alginate include human lung 99,100 , human brain 101 , murine intestinal 102 , human intestinal 102,103 , human pancreatic 104 , and human and murine vascular 105 . Other blood vessels are structurally intact, despite cell removal (e), with the structural details apparent even at high magnification (f).…”

“…The authors hypothesized that cells create their own niche within the alginate hydrogel by secreting basement membrane proteins and forming mesenchyme, allowing cellular survival and differentiation into HIOs 103 . Rossen and colleagues demonstrated the development of murine and human vascular organoids in a non-functionalized alginate setting 105 . Alginate has a number of advantages that make it attractive as a material for further study; it is inexpensive, relatively easy to modify and functionalize, biocompatible, and has been used in a wide range of biological and materials applications [106][107][108] .…”

“…While the backbone materials of synthetic hydrogels are cheap and can be produced on an industrial scale, functionalization of these materials with precisely-placed, custom-made peptides significantly increases cost and requires expertise in materials science, making these materials less attractive to cell biology labs. There have been a number of studies showing that organoids can be grown on unmodified surfaces such as alginate, but this requires more research 103,105,110,175,183 . Further, synthetic hydrogels may degrade into cytotoxic by-products 184 or require cytotoxic initiators 185 , limiting the types of polymers that can be used in cell culture 186 .…”

Therapeutic strategies of three-dimensional stem cell spheroids and organoids for tissue repair and regeneration

“…The bio-inspired square microwells spontaneously formed spheroids with high viability, and fibronectins conjugated to the hydrogel successfully gripped the spheroids, similar to the funiculus gripping seeds in the lotus seedpod. Dissolvable microwell scaffolds generated using alginate hydrogels were utilized to allow cells to form self-organized clusters and then be gently released, resulting in highly reproducible multicellular structures that can be produced on a large scale [ 134 ]. The PEG hydrogel-based microwell array, containing 1225 wells (200 μm in diameter) composed of cylindrical microwells with inverted-pyramidal openings, was fabricated to prevent cell loss during the mass-production of hMSC-spheroids [ 135 ].…”

Translating Embryogenesis to Generate Organoids: Novel Approaches to Personalized Medicine

“…Endothelial cells can be mixed with lineage progenitors to generate vascularization in organoids. Injectable organoids have recently been developed in hydrogels that can also generate perfusive vasculature in mice in vivo ( Rossen et al., 2020 ). Although improved vasculature is currently used, engineering organoids with other surrounding tissue types are necessary.…”