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 5

Abundance of adult Prg4-lineage progenitors in pre- and postinjury bone marrow and cartilage wounds.

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Abundance of adult Prg4-lineage progenitors in pre- and postinjury bone ...
(A and B) Confocal fluorescence images with maximum intensity projection of Prg4 CreERt;tdTomato knee wound (using punch) at 21 dpi, having received 6 mg tamoxifen before injury and 4 mg after injury (2 mg at injury, 1 mg at 3 dpi, and 1 mg at 14 dpi). Dotted lines: the wound border. Repaired cartilage matrix in aggrecan protein immunofluorescent staining (green). Green was pseudocolored based on staining signals of far-red fluorophores conjugated to secondary antibodies to eliminate the background associated with cartilage matrix autofluorescence. Scale bars: 50 μm. (C) Quantification: Percentage tdTomato+ cells in marrow and wounds with different tamoxifen timing and dosing: marrow homeostasis control (12 mg tamoxifen before injury with data taken from experiments related to Supplemental Figure 4 as described in the text), marrow and wound injury control (injury first, then tamoxifen, reversing the sequence of tamoxifen and injury protocol used in Figure 2, and euthanasia within 2–4 days to detect active Prg4-expressing cells in postinjury marrow and wounds, with data taken from Supplemental Figure 9). Single-dose tamoxifen: 4 mg preinjury only (data taken from Figure 2K); multiple-dose tamoxifen: before injury (6 mg n = 3 or 12 mg n = 4) and after injury (all 4 mg). Sample size: homeostasis control marrow (12 mg tamoxifen, n = 4, data taken from experiments related to Supplemental Figure 4 as described in the text); injury control marrow and wounds (n = 10 combining all time points from Supplemental Figure 9); single-dose tamoxifen wounds (n = 5 from Figure 2K); multiple-dose tamoxifen wounds (total n = 7). One-way ANOVA, F = 227.0 (degrees of freedom between groups = 4 and within groups = 31). ****P < 0.0001.