Stem cells for organoids

Abstract

Organoids are three-dimensional (3D) cell culture systems that simulate the structures and functions of organs, involving applications in disease modeling, drug screening, and cellular developmental biology. The material matrix in organoids can provide a 3D environment for stem cells to differentiate into different cell types and continuously self-renew, thereby realizing the in vitro culture of organs, which has received extensive attention in recent years. However, some challenges still exist in organoids, including low maturity, high heterogeneity, and lack of spatiotemporal regulation. Therefore, in this review, we summarized the culturing protocols and various applications of stem cell-derived organoids and proposed insightful thoughts for engineering stem cells into organoids in view of the current shortcomings, to achieve the further application and clinical translation of stem cells and engineered stem cells in organoid research.

Keywords: cell surface engineering; gene editing; organoid; stem cell.

FIGURE 1

Schematic diagram of the development history of organoids. Organoids have been widely used in disease modeling, drug research, cell development research, and tissue regeneration. Created by FigDraw.

FIGURE 2

Cultivation procedure, applications, and readout evaluation of organoids. Created by Biorender.

FIGURE 3

Extracellular matrix hydrogel derived from decellularized tissues. Reproduced under terms of the CC‐BY license. Copyright 2019, The Authors, published by Springer Nature. (A) The gelation preparation protocol. (B) SEM images of the ECM gel displaying the interconnected fibrous network. Scale bars 1 µm. (C, D) Storage modulus and loss modulus of the ECM gel and Matrigel. (E) Bright field and H&E images of mouse intestinal organoids in ECM gel and Matrigel. Scale bars 100 µm. (F) Culture of liver ductal and hepatocyte human organoids in ECM gel, BME, and Matrigel. Scale bar 500 µm. (G) Immunofluorescence staining of SI organoids in ECM gel after 4 weeks in vivo. Scale bars 100 µm. BME, basement membrane extract; ECM, extracellular matrix; SEM, scanning electron microscope; SI, small intestine.

FIGURE 4

Programing the reconstitution of fully ECM‐embedded 3D microtissue arrays by DNA‐programed assembly (DPAC). Reproduced with permission. Copyright 2015, Springer Nature. (A) Scheme showing the relationship between DNA spots (colored squares), DNA‐programed connectivity (colored lines), and multistep assembly. (B) Images of fully embedded aggregates of human luminal and myoepithelial cells. (C) Heat map illustrating differences in global cell position in two dimensions versus three dimensions relative to the pattern center. (D) Representative images of human mammary luminal and myoepithelial cells assembled through identical four‐step synthetic schemes and then transferred to Matrigel or collagen I. (E) Maximum‐intensity projection of a center‐patterned microtissue after processing using CLARITY. Insets are single confocal sections of the indicated region of the microtissue. ECM, extracellular matrix.

FIGURE 5

A computational model of PSC dynamics, enabling a machine learning optimization approach to predict experimental conditions. (A) Space–time relationships are captured with velocity characterizations, time–protein expression is captured characterizing the relative protein expression for several days after knockdown, and protein–space relationships are characterized by confocal microscopy imaging of spatial behavior due to cell mechanical perturbations. (B) Paired in vitro and in silico, schematic images of spatial patterning after CDH1 (blue) and ROCK1 (red) knockdown in stem cells. Reproduced with permission. Copyright 2020, The Authors, published by the National Academy of Sciences of the United States of America PSC, pluripotent stem cells.