Adult Prg4+ progenitors repair long-term articular cartilage wounds in vivo
Mei Massengale, Justin L. Massengale, Catherine R. Benson, Ninib Baryawno, Toshihiko Oki, Matthew L. Steinhauser, Alissa Wang, Deepak Balani, Luke S. Oh, Mark A. Randolph, Thomas J. Gill III, Henry M. Kronenberg, David T. Scadden
Mei Massengale, Justin L. Massengale, Catherine R. Benson, Ninib Baryawno, Toshihiko Oki, Matthew L. Steinhauser, Alissa Wang, Deepak Balani, Luke S. Oh, Mark A. Randolph, Thomas J. Gill III, Henry M. Kronenberg, David T. Scadden
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Research Article Cell biology

Adult Prg4+ progenitors repair long-term articular cartilage wounds in vivo

Abstract

The identity and origin of the stem/progenitor cells for adult joint cartilage repair remain unknown, impeding therapeutic development. Simulating the common therapeutic modality for cartilage repair in humans, i.e., full-thickness microfracture joint surgery, we combined the mouse full-thickness injury model with lineage tracing and identified a distinct skeletal progenitor cell type enabling long-term (beyond 7 days after injury) articular cartilage repair in vivo. Deriving from a population with active Prg4 expression in adulthood while lacking aggrecan expression, these progenitors proliferate, differentiate to express aggrecan and type II collagen, and predominate in long-term articular cartilage wounds, where they represent the principal repair progenitors in situ under native repair conditions without cellular transplantation. They originate outside the adult bone marrow or superficial zone articular cartilage. These findings have implications for skeletal biology and regenerative medicine for joint injury repair.

Authors

Mei Massengale, Justin L. Massengale, Catherine R. Benson, Ninib Baryawno, Toshihiko Oki, Matthew L. Steinhauser, Alissa Wang, Deepak Balani, Luke S. Oh, Mark A. Randolph, Thomas J. Gill III, Henry M. Kronenberg, David T. Scadden

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Figure 4

mRNA FISH analysis of Prg4-lineage wound cells.

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mRNA FISH analysis of Prg4-lineage wound cells. (A and B) mRNA FISH assa...
(A and B) mRNA FISH assay of Prg4 CreERt;tdTomato knee wounds (using punch), given 12 mg tamoxifen, showing colocalization of aggrecan (A) or Col2 (B) mRNA transcripts with tdTomato protein in wound cells. Dotted lines: the wound border. Blue: DAPI. Red: tdTomato. Red was pseudocolored based on the staining signals of anti-tdTomato secondary antibody. Green: aggrecan or Col2 mRNA probe. Green was pseudocolored based on the staining results of mRNA FISH using far-red fluorophores to eliminate background associated with cartilage matrix autofluorescence. Scale bars: 50 μm. Quantification: Percentage tdTomato+ wound cells expressing aggrecan mRNA or Col2 mRNA transcripts. Sample size: n = 3 per group. Aggrecan: paired t test, 2-tailed, t = 29.13, degrees of freedom = 2, **P = 0.0012. Col2: paired t test, 2-tailed, t = 51.99, degrees of freedom = 2, ***P = 0.0004. (C) EdU proliferation assay of Prg4 CreERt;tdTomato knee wound cells: 4 mg tamoxifen followed by washout and injury as in Figure 2, followed by 5 daily EdU injections (1.25 mg/animal) prior to euthanasia at 10 dpi, and the last dose within 48 hours of euthanasia. Inset: maximum intensity projection (projection of brightest signals only). Red: tomato. Blue: DAPI. Green: EdU. Green was pseudocolored based on the staining signals of far-red fluorophores to eliminate the background associated with cartilage matrix autofluorescence. Scale bars: 50 μm.