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DevelopmentDevelopmental BiologyPeer-reviewed journal article

Fibroblast response to burn injury in larval zebrafish mirrors developmental maturation and is inhibited by infiltrating neutrophils

Adam J. HorniD, Yiran HouiD, Jayne M. SquirrelliD, Alexandra M FisteriD, Julie RindyiD, Veronika MiskolciiD, Jonathan H. SchropeiD, Russell BurkeiD, Colin N. DeweyiD, Kevin W. EliceiriiD, Anna Huttenlocher AiD

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Abstract

Fibroblasts mediate tissue repair after damage, but aberrant fibroblast behavior in response to injury can result in impaired wound healing. Severe burn injury often results in tissue scarring, but the underlying mechanisms by which fibroblasts respond to burn injury and the role of inflammation in fibrosis are not well understood. Here we developed fluorescent reporters of collagen expressing mesenchymal cells enabling real time imaging of fibroblasts during homeostatic development and in response to burn injury using larval zebrafish. We find that fibroblasts derived from the mesenchyme respond to burn injury by engaging in a maturation process, characterized by the expression of vimentin, which is reminiscent of larval development. In burned tissue, fibroblast maturation is perturbed by prolonged neutrophil infiltration, resulting in disorganized extracellular matrix (ECM) and delayed ECM remodeling, which can be rescued by neutrophil depletion. This work adds to our understanding of fibroblast development in zebrafish and shows that collagen expressing mesenchymal cells regulate ECM remodeling in coordination with immune cells during burn wound healing. Summary StatementVimentin-positive fibroblasts are required for normal wound healing in larval zebrafish, but the presence of inflammatory neutrophils after burn injury impairs fibroblast maturation and collagen remodeling.

In-depth summary

Claude Sonnet 4.6 · xhigh effort · 8 Nov 2026 · from the full text of version 1 (22 Jun 2026) · CC BY 4.0

Vimentin-driven maturation of col1a1a-positive fibroblasts after larval zebrafish burn parallels fin development, but prolonged neutrophil presence delays fibroblast maturation and organized collagen regrowth.

Burn scarring mechanisms are poorly understood, and the interplay between infiltrating neutrophils and fibroblasts in the wound is unresolved, since neutrophils can both promote and impede fibrosis. This work adds in vivo fibroblast reporters and shows that neutrophil presence during the remodeling phase inhibits fibroblast maturation and collagen organization.

Key results

  1. SuggestedDuring tail fin development, vimentin is absent from col1a1a-positive cells nearest the midline and rises progressively toward the fin edge, while the double-positive fraction grows over time, consistent with a maturation program with altered cell shape.

    Double-positive fibroblasts were 55.7±2.4% at 3 days post-fertilization and 67.4±1.9 at 6 days post-fertilization (N ≥ 16 larvae per day). Total col1a1a-positive cell number did not increase over this period. · Mesenchymal cells express col1a1a and vimentin…; Figure legends

  2. ShownAfter burn, the fibroblast population is lost and then repopulates, and vimentin-positive (mature-like) fibroblasts accumulate slowly with morphology changes resembling development. Recovery is incomplete.

    Double-positive cells were 4.3 ± 0.7 (24 hpb), 11.4 ± 1.1 (48 hpb) and 20.6 ± 1.3 (72 hpb); about 21% double positive at 72 hpb vs 60% before burn. Not restored at 7 days post-burn (N ≥ 10). · Burn wounding triggers vimentin expression and…; Figure legends

  3. ShownReducing vimentin protein with a morpholino impairs fibroblast maturation after burn and slows tail fin regrowth, without altering fibroblast number or fin size in unwounded larvae.

    At 72 hpb col1a1a-positive cells were 61.3 ± 4.1 in vimentin MO larvae vs 86.6 ± 4.7 in controls (N ≥ 18). Tailfin length showed a 15% decrease (N ≥ 21). Perimeter was reduced and circularity increased. · Vimentin expression is required for fibroblast…; Figure legends

  4. ShownBlocking neutrophil recruitment (neutrophil-specific Rac2D57N) leaves early fibroblast morphology unchanged but yields more mature vimentin-positive fibroblast morphology at 72 hpb, implicating late neutrophil presence in inhibiting maturation.

    No morphology difference vs Rac2WT at 24 hpb. At 72 hpb Rac2D57N vimentin-positive cells had greater perimeter and reduced circularity, with total vimentin area unchanged (N ≥ 20 larvae per genotype, independent t-test). · Prolonged neutrophil inflammation inhibits…; Figure legends

  5. ShownPreventing neutrophil infiltration improves the extent and organization of collagen regrowth after burn, measured label-free by second harmonic generation (SHG) imaging with blinded ranking and automated fiber-angle analysis.

    92.1% (35 out of 38 total rankings) of no-defect (category 1) images were Rac2 D57N. Severe-defect images were 18.5% of rankings with no distribution difference. WT showed larger fiber deviation angles (N ≥ 25 larvae per genotype). · Neutrophil-fibroblast interactions inhibit…; Figure legends

Methods, data and reproducibility

Bulk RNA-seq (DESeq2, WGCNA) of burned caudal fins from 6 to 72 hpb, plus live imaging of col1a1a and vimentin reporter larvae after cautery burn, vimentin morpholino knockdown, Rac2D57N neutrophil-migration mutants, and SHG collagen imaging.

  • Bulk RNA-seq of caudal fin tissue, burned vs unwounded age-matched controls, at 6, 24, 48 and 72 hpb · zebrafish (AB background) · n = 3 biological replicates per time point; 50 larvae pooled per sample · GSE315881
  • Live imaging of col1a1a and vimentin reporter larvae during tail fin development · zebrafish (AB background) · n = N ≥ 16 larvae per day post-fertilization
  • Burned col1a1a/vimentin double-reporter larvae imaged 24 to 72 hpb and at 7 days post-burn · zebrafish (AB background) · n = N ≥ 19 larvae per time point (24 to 72 hpb); N ≥ 10 per condition at 7 dpb
  • Vimentin morpholino vs water-injected control larvae, unwounded and 72 hpb · zebrafish (AB background) · n = N ≥ 8 larvae (unwounded); N ≥ 18 (cells, 72 hpb); N ≥ 21 (fin regrowth)
  • Rac2WT vs Rac2D57N neutrophil lines crossed to vimentin reporter, morphology at 24 and 72 hpb · zebrafish (AB background) · n = N ≥ 20 larvae per genotype
  • SHG imaging of collagen in Rac2WT vs Rac2D57N burned fins at 72 hpb · zebrafish (AB background) · n = N ≥ 25 larvae per genotype
Methods & tools
RSEM, STAR 2.7.11b, DESeq2 v1.38.3, WGCNA, Metascape V3.5.20250701, ImageJ, CT-FIRE module of CurveAlign v5.0 Beta, OpenScan, GraphPad Prism 10.3.0, GRCz11 zebrafish genome, Second harmonic generation multiphoton microscopy, Vimentin morpholino
Software & environment
STAR 2.7.11b, DESeq2 v1.38.3, Metascape V3.5.20250701, CurveAlign v5.0 Beta on Matlab 2018b, GraphPad Prism 10.3.0; no container or lockfile reported.
Reproducibility gaps
Newly generated fish lines and all other data are available only on request; the GEO series was accessible to reviewers via a token at submission.
Limitations (5)
  • Bulk RNA-seq cannot resolve cell-type-specific expression, so results must be interpreted with caution. · stated by the authors
  • Vimentin-positive but col1a1a-negative cells at the fin edge were excluded from analysis, and a contribution of these cells to ECM remodeling cannot be completely ruled out. · stated by the authors
  • The mechanism by which neutrophils alter ECM remodeling is unclear, and MMP9-mediated degradation is only proposed. The authors chose to focus on neutrophils rather than macrophages. · stated by the authors
  • Maturation is inferred from vimentin reporter expression and cell shape in reporter lines; the morpholino knockdown lacks a reported rescue or second independent loss-of-function allele. · noted in this summary
  • The Rac2D57N manipulation blocks neutrophil recruitment broadly, so a direct neutrophil-to-fibroblast effect is not separated from indirect effects on other cells or the wound environment. · noted in this summary

Read this if you study fibroblast heterogeneity, burn wound healing, or neutrophil-fibroblast crosstalk using live-imaging zebrafish models.

Citations
1
FWCI
2.70
Field percentile
top 8%

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