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Human pluripotent stem cell fate trajectories toward lung and hepatocyte progenitors.

ABSTRACT: In this study, we interrogate molecular mechanisms underlying the specification of lung progenitors from human pluripotent stem cells (hPSCs). We employ single-cell RNA-sequencing with high temporal precision, alongside an optimized differentiation protocol, to elucidate the transcriptional hierarchy of lung specification to chart the associated single-cell trajectories. Our findings indicate that Sonic hedgehog, TGF-β, and Notch activation are essential within an ISL1/NKX2-1 trajectory, leading to the emergence of lung progenitors during the foregut endoderm phase. Additionally, the induction of HHEX delineates an alternate trajectory at the early definitive endoderm stage, preceding the lung pathway and giving rise to a significant hepatoblast population. Intriguingly, neither KDR+ nor mesendoderm progenitors manifest as intermediate stages in the lung and hepatic lineage development. Our multistep model offers insights into lung organogenesis and provides a foundation for in-depth study of early human lung development and modeling using hPSCs.

SUBMITTER: Ori C

PROVIDER: S-EPMC10663741 | biostudies-literature | 2023 Nov

REPOSITORIES: biostudies-literature

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Human pluripotent stem cell fate trajectories toward lung and hepatocyte progenitors.

Ori Chaido C Ansari Meshal M Angelidis Ilias I Olmer Ruth R Martin Ulrich U Theis Fabian J FJ Schiller Herbert B HB Drukker Micha M

iScience 20231014 11

In this study, we interrogate molecular mechanisms underlying the specification of lung progenitors from human pluripotent stem cells (hPSCs). We employ single-cell RNA-sequencing with high temporal precision, alongside an optimized differentiation protocol, to elucidate the transcriptional hierarchy of lung specification to chart the associated single-cell trajectories. Our findings indicate that Sonic hedgehog, TGF-β, and Notch activation are essential within an ISL1/NKX2-1 trajectory, leading ...[more]

PMID: 38026193

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Project description:Mast cells (MCs) are derived from CD34+ hematopoietic progenitors, consist of different subtypes, and are involved in several inflammatory conditions. However, our understanding of human MC developmental trajectories and subtypes has been limited by a scarcity of suitable cellular model systems. Herein, we developed an in vitro model of human MC differentiation from induced pluripotent stem cells (iPSC) to study human MC differentiation trajectories. Flow cytometry characterization of hemopoietic cells derived from the myeloid cells-forming complex (MCFC) revealed an initial increase in Lin- CD34+ hematopoietic progenitors within Weeks 1-3, followed by an increase in CD34- CD45RA- SSClow and SSChigh hematopoietic cells. The Lin- CD34+ hematopoietic progenitors consisted of SSClow CD45RA- CD123± c-Kit+ FcεRI+ populations that were β7-integrinhigh CD203c+ and β7-integrinhigh CD203c- cells consistent with CMPFcεRI+ cells. Flow cytometry and cytologic analyses of the CD34- Lin- (SSClow) population revealed hypogranular cell populations, predominantly characterized by CD45RA- CD123± c-Kit+ FcεRI- β7-integrinlow and CD45RA- CD123± c-Kit- FcεRI+ β7-integrinMid cells. Analyses of hypergranular SSChigh cells identified Lin- CD34- CD45RA- c-Kit+ FcεRI- and Lin- CD34- CD45RA- c-Kit+ FcεRI+ cells. scRNA-seq analysis of the cells harvested at week 4 of the MCFC culture revealed the presence of monocyte and granulocyte progenitors (n = 547 cells, 26.7 %), Erythrocyte / unknown (n = 85, 4.1 %), neutrophils / myelocytes (n = 211 cells, 10.2 %), mast cell progenitor 1 (n = 599, 29.1 %), mast cell progenitor 2 (n = 152, 7.4 %), committed mast cell precursor (n = 113, 5.5 %), and MCs (n = 353, 17.1 %). In silico analyses of the MC precursor and mature MC populations revealed transcriptionally distinct MC precursor subtype and mature MC states (CMA1+ and CMA1- subtypes). Culturing MC precursor populations in MC maturation media (mast cell media II) led to homogenous mature MC populations as evidenced by high expression of high-affinity IgE receptor, metachromatic granules, presence of MC granule proteins (Tryptase and Chymase) and activation following substance P stimulation and FcεRI crosslinking. This human iPSC-based approach generates MC precursors and phenotypically mature and functional MC populations. This system will be a useful model to generate human MC populations and broaden our understanding of MC biology and transcriptional regulation of MC differentiation trajectories.

Project description:Oligodendrogenesis encompasses lineage specification of neural progenitor cells (NPCs) and differentiation into oligodendrocytes that ultimately culminates in the myelination of central nervous system axons. Each individual process must be tightly regulated by extracellular and cell-intrinsic mechanisms, whose identities are barely understood. We had previously demonstrated that soluble factors derived from rat mesenchymal stem cells (MSCs) induce oligodendrogenesis in differentiating adult NPCs under differentiation conditions. However, since lineage specification predominantly occurs in proliferating progenitors and not necessarily during early differentiation, we investigated if soluble factors derived from MSCs are able to prime NPCs to the oligodendroglial fate already under proliferation conditions. Therefore, we analyzed the effects of a 3 weeks stimulation of adult NPCs under proliferation conditions with conditioned media derived from MSCs (MSC-CM) in terms of cell morphology, proliferation, cell-specific marker expression profile, response to growth factor withdrawal (GFW), cell-lineage restriction, and expression of glial fate determinants. While MSC-CM did not affect the proliferation rate of NPCs, it boosted the formation of 2', 3'-cyclic-nucleotide-3'-phosphodieesterase (CNPase)- and myelin basic protein-expressing oligodendrocytes after GFW, even when cells were exposed to an astrogenic milieu. Moreover, it reinforced the proper development of oligodendrocytes, since it ensured a sustained expression of the functional marker CNPase. Finally, the presence of MSC-CM reduced the anti-oligodendrogenic determinant Id2 in proliferating NPCs, thus increasing the relative proportion of the pro-oligodendrogenic factor Olig2 expression. In summary, MSCs prime proliferating progenitors and, thus, reinforce cell fate choice and accelerate differentiation toward the oligodendrocyte lineage. The present findings underscore the potential use of MSCs in cell therapies for remyelination such as in multiple sclerosis and spinal cord injury. Moreover, they urge the identification of the oligodendrogenic activity(ies) derived from MSCs to develop novel molecular therapies for demyelinating diseases.

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Human pluripotent stem cell fate trajectories toward lung and hepatocyte progenitors.

Publications

Human pluripotent stem cell fate trajectories toward lung and hepatocyte progenitors.

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