Spatial patterning of liver progenitor cell differentiation mediated by cellular contractility and Notch signaling

Abstract

The progenitor cells of the developing liver can differentiate toward both hepatocyte and biliary cell fates. In addition to the established roles of TGFβ and Notch signaling in this fate specification process, there is increasing evidence that liver progenitors are sensitive to mechanical cues. Here, we utilized microarrayed patterns to provide a controlled biochemical and biomechanical microenvironment for mouse liver progenitor cell differentiation. In these defined circular geometries, we observed biliary differentiation at the periphery and hepatocytic differentiation in the center. Parallel measurements obtained by traction force microscopy showed substantial stresses at the periphery, coincident with maximal biliary differentiation. We investigated the impact of downstream signaling, showing that peripheral biliary differentiation is dependent not only on Notch and TGFβ but also E-cadherin, myosin-mediated cell contractility, and ERK. We have therefore identified distinct combinations of microenvironmental cues which guide fate specification of mouse liver progenitors toward both hepatocyte and biliary fates.

Data availability

Source data tables (9 total) for the immunofluorescence and TFM array experiments have been attached to this submission and are associated with the relevant figures. Source code files (11 total) have been uploaded for the TFM analysis (Figure 4-6), FEM simulations (Figure 4), and Notch simulations (Figure 5). A detailed protocol for our array analysis technique together with source code has been made available elsewhere, see Kaylan et al. (J Vis Exp, 2017, e55362, http://dx.doi.org/10.3791/55362).

Article and author information

Author details

  1. Kerim B Kaylan

    Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-7147-0614
  2. Ian C Berg

    Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Matthew J Biehl

    Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Aidan Brougham-Cook

    Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Ishita Jain

    Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Sameed M Jamil

    Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Lauren H Sargeant

    Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Nicholas J Cornell

    Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  9. Lori T Raetzman

    Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, Urbana, United States
    Competing interests
    The authors declare that no competing interests exist.
  10. Gregory H Underhill

    Department of Bioengineering, University of Illinois at Urbana-Champaign, Urbana, United States
    For correspondence
    gunderhi@illinois.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1002-5335

Funding

National Institute of Biomedical Imaging and Bioengineering (5R03EB022254-02)

  • Gregory H Underhill

National Science Foundation (1636175)

  • Gregory H Underhill

National Institute of Biomedical Imaging and Bioengineering (T32EB019944)

  • Ian C Berg

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

Reviewing Editor

  1. Gordana Vunjak-Novakovic, Columbia University, United States

Publication history

  1. Received: May 21, 2018
  2. Accepted: December 24, 2018
  3. Accepted Manuscript published: December 27, 2018 (version 1)
  4. Version of Record published: January 22, 2019 (version 2)

Copyright

© 2018, Kaylan et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 3,171
    Page views
  • 411
    Downloads
  • 18
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Kerim B Kaylan
  2. Ian C Berg
  3. Matthew J Biehl
  4. Aidan Brougham-Cook
  5. Ishita Jain
  6. Sameed M Jamil
  7. Lauren H Sargeant
  8. Nicholas J Cornell
  9. Lori T Raetzman
  10. Gregory H Underhill
(2018)
Spatial patterning of liver progenitor cell differentiation mediated by cellular contractility and Notch signaling
eLife 7:e38536.
https://doi.org/10.7554/eLife.38536

Further reading

    1. Developmental Biology
    2. Stem Cells and Regenerative Medicine
    Brian Silver, Kevin Gerrish, Erik Tokar
    Research Article

    Cell-free DNA (cfDNA) present in the bloodstream or other bodily fluids holds potential as a non-invasive diagnostic for early disease detection. However, it remains unclear what cfDNA markers might be produced in response to specific tissue-level events. Organoid systems present a tractable and efficient method for screening cfDNA markers. However, research investigating the release of cfDNA from organoids is limited. Here, we present a scalable method for high-throughput screening of cfDNA from cardiac organoids. We demonstrate that cfDNA is recoverable from cardiac organoids, and that cfDNA release is highest early in differentiation. Intriguingly, we observed that the fraction of cell-free mitochondrial DNA appeared to decrease as the organoids developed, suggesting a possible signature of cardiac organoid maturation, or other cardiac growth-related tissue-level events. We also observe alterations in the prevalence of specific genomic regions in cardiac organoid-derived cfDNA at different timepoints during growth. In addition, we identify cfDNA markers that were increased upon addition of cardiotoxic drugs, prior to the onset of tissue demise. Together, these results indicate that cardiac organoids may be a useful system towards the identification of candidate predictive cfDNA markers of cardiac tissue development and demise.

    1. Developmental Biology
    2. Neuroscience
    Sweta Parab, Olivia A Card ... Ryota L Matsuoka
    Research Article Updated

    Fenestrated and blood-brain barrier (BBB)-forming endothelial cells constitute major brain capillaries, and this vascular heterogeneity is crucial for region-specific neural function and brain homeostasis. How these capillary types emerge in a brain region-specific manner and subsequently establish intra-brain vascular heterogeneity remains unclear. Here, we performed a comparative analysis of vascularization across the zebrafish choroid plexuses (CPs), circumventricular organs (CVOs), and retinal choroid, and show common angiogenic mechanisms critical for fenestrated brain capillary formation. We found that zebrafish deficient for Gpr124, Reck, or Wnt7aa exhibit severely impaired BBB angiogenesis without any apparent defect in fenestrated capillary formation in the CPs, CVOs, and retinal choroid. Conversely, genetic loss of various Vegf combinations caused significant disruptions in Wnt7/Gpr124/Reck signaling-independent vascularization of these organs. The phenotypic variation and specificity revealed heterogeneous endothelial requirements for Vegfs-dependent angiogenesis during CP and CVO vascularization, identifying unexpected interplay of Vegfc/d and Vegfa in this process. Mechanistically, expression analysis and paracrine activity-deficient vegfc mutant characterization suggest that endothelial cells and non-neuronal specialized cell types present in the CPs and CVOs are major sources of Vegfs responsible for regionally restricted angiogenic interplay. Thus, brain region-specific presentations and interplay of Vegfc/d and Vegfa control emergence of fenestrated capillaries, providing insight into the mechanisms driving intra-brain vascular heterogeneity and fenestrated vessel formation in other organs.