An improved organ explant culture method reveals stem cell lineage dynamics in the adult Drosophila intestine

  1. Marco Marchetti
  2. Chenge Zhang
  3. Bruce A Edgar  Is a corresponding author
  1. University of Utah, United States

Abstract

In recent years, live-imaging techniques have been developed for the adult midgut of Drosophila melanogaster that allow temporal characterization of key processes involved in stem cell and tissue homeostasis. However, these organ culture techniques have been limited to imaging sessions of <16 hours, an interval too short to track dynamic processes such as damage responses and regeneration, which can unfold over several days. Therefore, we developed an organ explant culture protocol capable of sustaining midguts ex vivo for up to 3 days. This was made possible by the formulation of a culture medium specifically designed for adult Drosophila tissues with an increased Na+/K+ ratio and trehalose concentration, and by placing midguts at an air-liquid interface for enhanced oxygenation. We show that midgut progenitor cells can respond to gut epithelial damage ex vivo, proliferating and differentiating to replace lost cells, but are quiescent in healthy intestines. Using ex vivo gene induction to promote stem cell proliferation using RasG12V or string and Cyclin E overexpression, we demonstrate that progenitor cell lineages can be traced through multiple cell divisions using live imaging. We show that the same culture set-up is useful for imaging adult renal tubules and ovaries for up to 3 days and hearts for up to 10 days. By enabling both long-term imaging and real-time ex vivo gene manipulation, our simple culture protocol provides a powerful tool for studies of epithelial biology and cell lineage behavior.

Data availability

Source Data files have been provided for Figures 2, 3, 3 - figure supplement 1, 4, 6, 7, 7 - figure supplement 1, 8

Article and author information

Author details

  1. Marco Marchetti

    Department of Oncological Sciences, University of Utah, Salt Lake City, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Chenge Zhang

    Department of Oncological Sciences, University of Utah, Salt Lake City, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Bruce A Edgar

    Department of Oncological Sciences, University of Utah, Salt Lake City, United States
    For correspondence
    bruce.edgar@hci.utah.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3383-2044

Funding

National Institutes of Health (R35 140900)

  • Bruce A Edgar

National Institutes of Health (R01 GM124434)

  • Bruce A Edgar

National Institutes of Health (P30 CA042014)

  • Bruce A Edgar

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Lucy Erin O'Brien, Stanford University School of Medicine, United States

Version history

  1. Received: December 1, 2021
  2. Preprint posted: December 19, 2021 (view preprint)
  3. Accepted: August 24, 2022
  4. Accepted Manuscript published: August 25, 2022 (version 1)
  5. Version of Record published: October 17, 2022 (version 2)

Copyright

© 2022, Marchetti 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.

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  1. Marco Marchetti
  2. Chenge Zhang
  3. Bruce A Edgar
(2022)
An improved organ explant culture method reveals stem cell lineage dynamics in the adult Drosophila intestine
eLife 11:e76010.
https://doi.org/10.7554/eLife.76010

Share this article

https://doi.org/10.7554/eLife.76010

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