Format

Send to

Choose Destination
BMC Biol. 2018 Jun 5;16(1):62. doi: 10.1186/s12915-018-0527-2.

Harnessing single-cell genomics to improve the physiological fidelity of organoid-derived cell types.

Mead BE1,2,3,4,5, Ordovas-Montanes J6,7,8,9,10, Braun AP11,6,12,7, Levy LE11,6,12, Bhargava P7,8, Szucs MJ7, Ammendolia DA11,12, MacMullan MA7, Yin X11,13,6,12, Hughes TK13,6,7,8,9, Wadsworth MH 2nd6,7,8,9, Ahmad R7, Rakoff-Nahoum S10, Carr SA7, Langer R13,6,14, Collins JJ13,7,8,15,16,17,18, Shalek AK13,6,7,8,9,19, Karp JM20,21,22,23.

Author information

1
Division of Engineering in Medicine, Department of Medicine, Brigham & Women's Hospital, Harvard Medical School, Boston, MA, USA. bemead@mit.edu.
2
Harvard-MIT Division of Health Sciences & Technology, Cambridge, MA, USA. bemead@mit.edu.
3
Koch Institute for Integrative Cancer Research, MIT, Cambridge, MA, USA. bemead@mit.edu.
4
Harvard Stem Cell Institute, Cambridge, MA, USA. bemead@mit.edu.
5
Broad Institute of Harvard and MIT, Cambridge, MA, USA. bemead@mit.edu.
6
Koch Institute for Integrative Cancer Research, MIT, Cambridge, MA, USA.
7
Broad Institute of Harvard and MIT, Cambridge, MA, USA.
8
Institute for Medical Engineering and Science, MIT, Cambridge, MA, USA.
9
Department of Chemistry, MIT, Cambridge, MA, USA.
10
Divisions of Infectious Diseases and Gastroenterology, Boston Children's Hospital and Harvard Medical School, Boston, MA, USA.
11
Division of Engineering in Medicine, Department of Medicine, Brigham & Women's Hospital, Harvard Medical School, Boston, MA, USA.
12
Harvard Stem Cell Institute, Cambridge, MA, USA.
13
Harvard-MIT Division of Health Sciences & Technology, Cambridge, MA, USA.
14
Department of Chemical Engineering, MIT, Cambridge, MA,, USA.
15
Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA, USA.
16
Department of Biological Engineering, MIT, Cambridge, MA, USA.
17
Synthetic Biology Center, MIT, Cambridge, MA, USA.
18
Center for Microbiome Informatics and Therapeutics, MIT, Cambridge, MA, USA.
19
Ragon Institute of MGH, MIT and Harvard, Cambridge, MA, USA.
20
Division of Engineering in Medicine, Department of Medicine, Brigham & Women's Hospital, Harvard Medical School, Boston, MA, USA. jmkarp@bwh.harvard.edu.
21
Harvard-MIT Division of Health Sciences & Technology, Cambridge, MA, USA. jmkarp@bwh.harvard.edu.
22
Harvard Stem Cell Institute, Cambridge, MA, USA. jmkarp@bwh.harvard.edu.
23
Broad Institute of Harvard and MIT, Cambridge, MA, USA. jmkarp@bwh.harvard.edu.

Abstract

BACKGROUND:

Single-cell genomic methods now provide unprecedented resolution for characterizing the component cell types and states of tissues such as the epithelial subsets of the gastrointestinal tract. Nevertheless, functional studies of these subsets at scale require faithful in vitro models of identified in vivo biology. While intestinal organoids have been invaluable in providing mechanistic insights in vitro, the extent to which organoid-derived cell types recapitulate their in vivo counterparts remains formally untested, with no systematic approach for improving model fidelity.

RESULTS:

Here, we present a generally applicable framework that utilizes massively parallel single-cell RNA-seq to compare cell types and states found in vivo to those of in vitro models such as organoids. Furthermore, we leverage identified discrepancies to improve model fidelity. Using the Paneth cell (PC), which supports the stem cell niche and produces the largest diversity of antimicrobials in the small intestine, as an exemplar, we uncover fundamental gene expression differences in lineage-defining genes between in vivo PCs and those of the current in vitro organoid model. With this information, we nominate a molecular intervention to rationally improve the physiological fidelity of our in vitro PCs. We then perform transcriptomic, cytometric, morphologic and proteomic characterization, and demonstrate functional (antimicrobial activity, niche support) improvements in PC physiology.

CONCLUSIONS:

Our systematic approach provides a simple workflow for identifying the limitations of in vitro models and enhancing their physiological fidelity. Using adult stem cell-derived PCs within intestinal organoids as a model system, we successfully benchmark organoid representation, relative to that in vivo, of a specialized cell type and use this comparison to generate a functionally improved in vitro PC population. We predict that the generation of rationally improved cellular models will facilitate mechanistic exploration of specific disease-associated genes in their respective cell types.

KEYWORDS:

Chemical biology; Differentiation; Intestinal organoid; Intestinal stem cell; Paneth cell; Single-cell RNA-seq; Stem cell-derived models; Systems biology

PMID:
29871632
PMCID:
PMC5989470
DOI:
10.1186/s12915-018-0527-2
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for BioMed Central Icon for PubMed Central
Loading ...
Support Center