Injury-induced pulmonary tuft cells are heterogenous, arise independent of key Type 2 cytokines, and are dispensable for dysplastic repair

  1. Justinn Barr
  2. Maria Elena Gentile
  3. Sunyoung Lee
  4. Maya E Kotas
  5. Maria Fernanda de Mello Costa
  6. Nicolas P Holcomb
  7. Abigail Jaquish
  8. Gargi Palashikar
  9. Marcella Soewignjo
  10. Margaret McDaniel
  11. Ichiro Matsumoto
  12. Robert Margolskee
  13. Jakob Von Moltke
  14. Noam A Cohen
  15. Xin Sun  Is a corresponding author
  16. Andrew E Vaughan  Is a corresponding author
  1. University of California, San Diego, United States
  2. University of Pennsylvania, United States
  3. University of California, San Francisco, United States
  4. University of Washington, United States
  5. Monell Chemical Senses Center, United States

Abstract

While the lung bears significant regenerative capacity, severe viral pneumonia can chronically impair lung function by triggering dysplastic remodeling. The connection between these enduring changes and chronic disease remains poorly understood. We recently described the emergence of tuft cells within Krt5+ dysplastic regions after influenza injury. Using bulk and single cell transcriptomics, we characterized and delineated multiple distinct tuft cell populations that arise following influenza clearance. Distinct from intestinal tuft cells which rely on Type 2 immune signals for their expansion, neither IL-25 nor IL-4ra signaling are required to drive tuft cell development in dysplastic/injured lungs. In addition, tuft cell expansion occurred independently of type I or type III interferon signalling. Furthermore, tuft cells were also observed upon bleomycin injury, suggesting that their development may be a general response to severe lung injury. While intestinal tuft cells promote growth and differentiation of surrounding epithelial cells, in the lungs of tuft cell deficient mice, Krt5+ dysplasia still occurs, goblet cell production is unchanged, and there remains no appreciable contribution of Krt5+ cells into more regionally appropriate alveolar Type 2 cells. Together, these findings highlight unexpected differences in signals necessary for murine lung tuft cell amplification and establish a framework for future elucidation of tuft cell functions in pulmonary health and disease.

Data availability

Sequencing data have been deposited in GEO under accession code GSE197163.In addition to the deposited sequencing data, raw numerical data is available as excel files corresponding to each figure, e.g. Figure 1 - Source Data.xls.

The following data sets were generated

Article and author information

Author details

  1. Justinn Barr

    Department of Pediatrics, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Maria Elena Gentile

    Department of Biomedical Sciences, University of Pennsylvania, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Sunyoung Lee

    Department of Pediatrics, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Maya E Kotas

    Division of Pulmonary, Critical Care, Allergy & Sleep Medicine, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Maria Fernanda de Mello Costa

    Department of Biomedical Sciences, University of Pennsylvania, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Nicolas P Holcomb

    Department of Biomedical Sciences, University of Pennsylvania, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Abigail Jaquish

    Department of Pediatrics, University of California, San Diego, San Diego, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Gargi Palashikar

    Department of Biomedical Sciences, University of Pennsylvania, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  9. Marcella Soewignjo

    Department of Biomedical Sciences, University of Pennsylvania, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  10. Margaret McDaniel

    Department of Immunology, University of Washington, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  11. Ichiro Matsumoto

    Monell Chemical Senses Center, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  12. Robert Margolskee

    Monell Chemical Senses Center, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  13. Jakob Von Moltke

    Department of Immunology, University of Washington, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  14. Noam A Cohen

    Department of Otorhinolaryngology-Head and Neck Surgery, University of Pennsylvania, Philadelphia, United States
    Competing interests
    The authors declare that no competing interests exist.
  15. Xin Sun

    Department of Pediatrics, University of California, San Diego, La Jolla, United States
    For correspondence
    xinsun@health.ucsd.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8387-4966
  16. Andrew E Vaughan

    Department of Biomedical Sciences, University of Pennsylvania, Philadelphia, United States
    For correspondence
    andrewva@vet.upenn.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5740-643X

Funding

National Institutes of Health (R01HL153539)

  • Andrew E Vaughan

U.S. Department of Veterans Affairs (CX001617)

  • Noam A Cohen

Fonds de Recherche du Québec - Santé

  • Maria Elena Gentile

Lisa Dean Moseley Foundation

  • Andrew E Vaughan

National Institutes of Health (R01HL142215)

  • Xin Sun

National Institutes of Health (1R01AT011676)

  • Xin Sun

National Institutes of Health (T29IR0475)

  • Xin Sun

National Institutes of Health (F32HL151168)

  • Justinn Barr

National Institutes of Health (F32HL140868)

  • Maya E Kotas

National Institutes of Health (T32HL007185)

  • Maya E Kotas

A.P. Giannini Foundation

  • Maya E Kotas

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

Ethics

Animal experimentation: All animal procedures were approved by the Institutional Animal Care and Use Committee (IACUC) of the University of Pennsylvania, the University of California - San Diego, and the University of California, San Francisco. All experiments were performed with every effort to minimize suffering. The protocol number associated with the ethical approval of this work is 806262 (University of Pennsylvania) and S16187 (University of California San Diego).

Copyright

© 2022, Barr 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

  • 2,545
    views
  • 520
    downloads
  • 32
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

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. Justinn Barr
  2. Maria Elena Gentile
  3. Sunyoung Lee
  4. Maya E Kotas
  5. Maria Fernanda de Mello Costa
  6. Nicolas P Holcomb
  7. Abigail Jaquish
  8. Gargi Palashikar
  9. Marcella Soewignjo
  10. Margaret McDaniel
  11. Ichiro Matsumoto
  12. Robert Margolskee
  13. Jakob Von Moltke
  14. Noam A Cohen
  15. Xin Sun
  16. Andrew E Vaughan
(2022)
Injury-induced pulmonary tuft cells are heterogenous, arise independent of key Type 2 cytokines, and are dispensable for dysplastic repair
eLife 11:e78074.
https://doi.org/10.7554/eLife.78074

Share this article

https://doi.org/10.7554/eLife.78074

Further reading

    1. Stem Cells and Regenerative Medicine
    Zhifei Wu, Baoshan Liao ... Wenjun Xiong
    Research Article

    Harnessing the regenerative potential of endogenous stem cells to restore lost neurons is a promising strategy for treating neurodegenerative disorders. Müller glia (MG), the primary glial cell type in the retina, exhibit extraordinary regenerative abilities in zebrafish, proliferating and differentiating into neurons post-injury. However, the regenerative potential of mouse MG is limited by their inherent inability to re-enter the cell cycle, constrained by high levels of the cell cycle inhibitor p27Kip1 and low levels of cyclin D1. Here, we report a method to drive robust MG proliferation by adeno-associated virus (AAV)-mediated cyclin D1 overexpression and p27Kip1 knockdown. MG proliferation induced by this dual targeting vector was self-limiting, as MG re-entered cell cycle only once. As shown by single-cell RNA-sequencing, cell cycle reactivation led to suppression of interferon signaling, activation of reactive gliosis, and downregulation of glial genes in MG. Over time, the majority of the MG daughter cells retained the glial fate, resulting in an expanded MG pool. Interestingly, about 1% MG daughter cells expressed markers for retinal interneurons, suggesting latent neurogenic potential in a small MG subset. By establishing a safe, controlled method to promote MG proliferation in vivo while preserving retinal integrity, this work provides a valuable tool for combinatorial therapies integrating neurogenic stimuli to promote neuron regeneration.

    1. Stem Cells and Regenerative Medicine
    Sujeethkumar Prithiviraj, Alejandro Garcia Garcia ... Paul E Bourgine
    Research Article

    Tissue engineering strategies predominantly rely on the production of living substitutes, whereby implanted cells actively participate in the regenerative process. Beyond cost and delayed graft availability, the patient-specific performance of engineered tissues poses serious concerns on their clinical translation ability. A more exciting paradigm consists in exploiting cell-laid, engineered extracellular matrices (eECMs), which can be used as off-the-shelf materials. Here, the regenerative capacity solely relies on the preservation of the eECM structure and embedded signals to instruct an endogenous repair. We recently described the possibility to exploit custom human stem cell lines for eECM manufacturing. In addition to the conferred standardization, the availability of such cell lines opened avenues for the design of tailored eECMs by applying dedicated genetic tools. In this study, we demonstrated the exploitation of CRISPR/Cas9 as a high precision system for editing the composition and function of eECMs. Human mesenchymal stromal/stem cell (hMSC) lines were modified to knock out vascular endothelial growth factor (VEGF) and Runt-related transcription factor 2 (RUNX2) and assessed for their capacity to generate osteoinductive cartilage matrices. We report the successful editing of hMSCs, subsequently leading to targeted VEGF and RUNX2-knockout cartilage eECMs. Despite the absence of VEGF, eECMs retained full capacity to instruct ectopic endochondral ossification. Conversely, RUNX2-edited eECMs exhibited impaired hypertrophy, reduced ectopic ossification, and superior cartilage repair in a rat osteochondral defect. In summary, our approach can be harnessed to identify the necessary eECM factors driving endogenous repair. Our work paves the road toward the compositional eECMs editing and their exploitation in broad regenerative contexts.