Engineering organoids

Abstract

Organoids are in vitro miniaturized and simplified model systems of organs that have gained enormous interest for modelling tissue development and disease, and for personalized medicine, drug screening and cell therapy. Despite considerable success in culturing physiologically relevant organoids, challenges remain to achieve real-life applications. In particular, the high variability of self-organizing growth and restricted experimental and analytical access hamper the translatability of organoid systems. In this Review, we argue that many limitations of traditional organoid culture can be addressed by engineering approaches at all levels of organoid systems. We investigate cell surface and genetic engineering approaches, and discuss stem cell niche engineering based on the design of matrices that allow spatiotemporal control of organoid growth and shape-guided morphogenesis. We examine how microfluidic approaches and lessons learnt from organs-on-a-chip enable the integration of mechano-physiological parameters and increase accessibility of organoids to improve functional readouts. Applying engineering principles to organoids increases reproducibility and provides experimental control, which will, ultimately, be required to enable clinical translation.

Keywords: Morphogenesis; Organogenesis; Stem cells; Tissue engineering.

Fig. 1. Tissue-derived organoids.

Organoids can be generated from tissue samples for a variety of organs. A1AT, alpha-1 antitrypsin; IBD, inflammatory bowel disease; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.

Fig. 2. Pluripotent stem-cell-derived organoids.

Organoids can be generated from pluripotent stem cells for a variety of organs. AD, Alzheimer disease; ALI, air–liquid interface; ASD, autism spectrum disorders; DKC, dyskeratosis congenita; EA, enteric anendocrinosis; EB, embryoid bodies; ESCs, embryonic stem cells; iPSCs, induced pluripotent stem cells; LCA, Leber congenital amaurosis; PD, Parkinson disease; PKD, polycystic kidney disease; RP, retinitis pigmentosa; RSV, respiratory syncytial virus; ZIKV, Zika virus.

Fig. 3. Engineering approaches for organoids.

Engineering approaches can be applied at several levels to organoids, including at the cellular level, niche level, multi-tissue level and to improve functional readouts.

Fig. 4. Engineering cells for guided morphogenesis.

Two populations of genetically engineered cells self-organize into multilayer tissues when mixed together. The microscopy images show cells of type A (blue, BFP+) and cell type B (grey) mixed together and their development into a three-layered structure with cells of type A, C (green, GFP+) and D (pink, mCherry+) by mutual activation and repression of E-cadherin (E-cad) expression. First, in the presence of A-type cells, B-type cells are converted to C-type cells that self-aggregate owing to their high E-cad expression. These cells present GFP at their surface, in turn, leading adjacent A-type cells to express low levels of E-cad and mCherry protein through activation of their synthetic anti-GFP receptor, which will lead to their conversion to D-type cells. BFP, blue fluorescent protein; GFP, green fluorescent protein; HI, high; LO, low. Microscopy images reprinted with permission from ref., AAAS.

Fig. 5. Engineering the niche.

Intestinal organoids grown in a patterned tubular matrix organize into a crypt-like structure akin to organoids, following the predefined architecture. Stem and progenitor cells (pink, Sox9+), as well as Paneth cells (light pink, Lysozyme+), are found in the crypt structures, whereas enterocytes (yellow, L-FABP+) are present towards the lumen. The integration into a microfluidic device allows the growth for extended periods of time. FABP, fatty-acid-binding protein. Adapted from ref., Springer Nature Limited.

Fig. 6. Systemic context engineering for organoid cultures.

Kidney organoids grown in the presence of flow-induced shear stresses show substantially enhanced vascularization and maturation. Confocal microscopy images show organoids under different flow conditions: nuclei (DAPI, blue), vasculature (MCAM, yellow, and PECAM1, red) and podocytes (PODXL, cyan) are stained. DAPI, 4′,6-diamidino-2-phenylindole; MCAM, melanoma cell adhesion molecule; PECAM1, platelet and endothelial cell adhesion molecule 1; PODXL, podocalyxin-like protein 1. Reprinted from ref., Springer Nature Limited.

Fig. 7. Microengineered platforms for live organoid monitoring.

Automated and continuous in situ monitoring of physical and biological parameters in organoid cultures can be achieved using microfluidic devices. The presented device contains multiple compartments for simultaneous growth of different organoids, as well as sensors to monitor various biological (for example, metabolites and biomarkers) and biophysical (temperature, pH and oxygen levels) parameters. The biosensor was designed to be regenerated, allowing continuous measurements (inset). APAP, acetaminophen; GST, glutathione S-transferase. Adapted with permission from ref., National Academy of Sciences.