Human heart-forming organoids recapitulate early heart and foregut development

Lika Drakhlis,Henning Kempf,Ulrich Martin,Jan Hegermann,Lena Nolte,Katja Schenke‐Layland ,Christian Wahl‐Schott ,Victoria Lupanow,Jana Teske,Katharina Ritzenhoff,Clara-Milena Farr,Felix Manstein,Heiko Meyer,Annika Franke,Stefan Thiemann,Emiliano Bolesani,Santoshi Biswanath,Simone Liebscher,Robert Zweigerdt

doi:10.1038/s41587-021-00815-9

Lika Drakhlis, Henning Kempf + Show 17 more

Open Access

https://doi.org/10.1038/s41587-021-00815-9

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Abstract

Organoid models of early tissue development have been produced for the intestine, brain, kidney and other organs, but similar approaches for the heart have been lacking. Here we generate complex, highly structured, three-dimensional heart-forming organoids (HFOs) by embedding human pluripotent stem cell aggregates in Matrigel followed by directed cardiac differentiation via biphasic WNT pathway modulation with small molecules. HFOs are composed of a myocardial layer lined by endocardial-like cells and surrounded by septum-transversum-like anlagen; they further contain spatially and molecularly distinct anterior versus posterior foregut endoderm tissues and a vascular network. The architecture of HFOs closely resembles aspects of early native heart anlagen before heart tube formation, which is known to require an interplay with foregut endoderm development. We apply HFOs to study genetic defects in vitro by demonstrating that NKX2.5-knockout HFOs show a phenotype reminiscent of cardiac malformations previously observed in transgenic mice.

Highlights

Human pluripotent stem cells, including embryonic stem cells and induced PSCs, have the capability to self-organize into three-dimensional (3D) structures called organoids resembling embryo-like tissue patterns in vitro
Free-floating Human pluripotent stem cells (hPSCs) aggregates efficiently differentiate into highly pure cardiomyocytes in suspension culture in response to canonical WNT pathway modulation by small molecules such as CHIR99021 (CHIR; WNT pathway activator) and IWP2 (WNT pathway inhibitor)[15,16]
For heart-forming organoids (HFOs) formation, we applied an equivalent protocol but using individual hPSC aggregates of a defined size (based on the cell number seeded for aggregate formation on differentiation day minus 4 (d−4)) embedded in Matrigel (Fig. 1a)

Summary

Introduction

Human pluripotent stem cells (hPSCs), including embryonic stem cells (hESCs) and induced PSCs (hiPSCs), have the capability to self-organize into three-dimensional (3D) structures called organoids resembling embryo-like tissue patterns in vitro. Mills et al.[7] and Voges et al.[9] generated organoids by embedding hPSC-derived cardiomyocytes in a collagen I–Matrigel mixture These organoids reproduce some aspects of adult heart tissue, including stromal cells, an endothelial network and an epicardial layer, but do not reflect the processes of early heart development. Using mouse PSCs, Andersen et al derived ‘precardiac organoids’ forming two heart fields, which showed high similarities to their in vivo counterparts at the corresponding developmental stage[12] These studies did not demonstrate the spatiotemporal patterning of early cardiogenesis, including the interplay with foregut endoderm. We demonstrate an application of the model to monitor gene-knockout (KO) phenotypes in vitro

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