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, 10 (1), 17-26

JAK/STAT-1 Signaling Is Required for Reserve Intestinal Stem Cell Activation During Intestinal Regeneration Following Acute Inflammation

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JAK/STAT-1 Signaling Is Required for Reserve Intestinal Stem Cell Activation During Intestinal Regeneration Following Acute Inflammation

Camilla A Richmond et al. Stem Cell Reports.

Abstract

The intestinal epithelium serves as an essential barrier to the outside world and is maintained by functionally distinct populations of rapidly cycling intestinal stem cells (CBC ISCs) and slowly cycling, reserve ISCs (r-ISCs). Because disruptions in the epithelial barrier can result from pathological activation of the immune system, we sought to investigate the impact of inflammation on ISC behavior during the regenerative response. In a murine model of αCD3 antibody-induced small-intestinal inflammation, r-ISCs proved highly resistant to injury, while CBC ISCs underwent apoptosis. Moreover, r-ISCs were induced to proliferate and functionally contribute to intestinal regeneration. Further analysis revealed that the inflammatory cytokines interferon gamma and tumor necrosis factor alpha led to r-ISC activation in enteroid culture, which could be blocked by the JAK/STAT inhibitor, tofacitinib. These results highlight an important role for r-ISCs in response to acute intestinal inflammation and show that JAK/STAT-1 signaling is required for the r-ISC regenerative response.

Keywords: CD3; JAK/STAT; STAT-1; dormant; inflammation; intestinal stem cells; quiescent; reserve.

Figures

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Figure 1
Figure 1
αCD3 Leads to Epithelial Injury and CBC Loss with Subsequent Recovery (A) Representative H&E images of small intestine at baseline, injury, early recovery, and late recovery. Scale bar, 100 μm. (B) Villus height and crypt depth (μm) following αCD3 compared with baseline. N = 2 mice/time point, 30/19, 83/83, 94/94, 147/147 total villus/crypt measurements counted. (C) Crypt cells expressing cleaved caspase-3 (CC3) at baseline (N = 20) and injury (N = 9). (D) Crypt cells expressing TUNEL at baseline (N = 4) and injury (N = 7). (E) Lgr5-GFP+CC3+CD45PI cells 48 hr after control IgG (N = 3) and αCD3 (N = 3). (F) mTert-GFP+CC3+CD45PI cells 48 hr after control IgG (N = 3) or αCD3 (N = 3). (G) Live Lgr5-GFPhi cells 48 hr after control IgG (N = 3) or αCD3 (N = 3). (H) Live mTert-GFP cells 48 hr after control IgG (N = 3) or αCD3 (N = 3). Error bars indicate the SEM. p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001, n.s., not significant. Representative flow cytometry plots shown.
Figure 2
Figure 2
R-ISC Numbers Increase and Enter the Cell Cycle following αCD3 (A) mTert-GFP+ crypt cells per 150 crypt sections at baseline, injury, early recovery, and late recovery. N = 2 mice/time point, 18, 21, 20, and 9 sections counted. (B) mTert-GFP+ crypt cells co-expressing Ki67 in control IgG and αCD3-treated mice at early recovery. Representative optical sections from control IgG- (N = 8) and αCD3-treated (N = 4) mice showing cycling (arrowhead) and quiescent (arrow) mTert-GFP+ cells in merged, DAPI, mTert-GFP, and Ki67 channels. (C) mTert-GFP+ cells co-expressing EdU in isolated crypts from control IgG- (N = 3) and αCD3-treated (N = 3) mice at early recovery. Representative optical sections showing quiescent mTert-GFP+ cells (arrow) in control IgG and cycling mTert-GFP+ cells in αCD3-treated (arrowhead) crypts in merged DAPI, mTert-GFP, and EdU (Click-iT) channels. Error bars indicate the SEM. p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. Scale bar, 10 μm.
Figure 3
Figure 3
R-ISCs Show Increased Lineage Contribution Following αCD3 while the CBC Contribution Is Reduced (A) Quantitative whole-mount analysis showing a decrease in the number of lineage-marked LacZ+ crypts from Lgr5-Cre::R26R(LacZ) IgG- (N = 4) and αCD3-treated (N = 4) mice at injury. Representative whole mounts of β-gal+ crypts. (B) Quantitative whole-mount analysis showing increase in the number of lineage-marked LacZ+ crypts from mTert-Cre::R26R(LacZ) IgG- (N = 9) and αCD3-treated (N = 10) mice during late recovery. Representative whole mounts of β-gal+ crypts. (C) Change in distribution of crypts with single and multiple mTert-LacZ+ cells 7 days after IgG (N = 9 mice) or αCD3 (N = 10 mice) indicating an increase in the number of crypts with multiple cells following αCD3 treatment. Error bars indicate the SEM. p < 0.05, ∗∗p < 0.01. Scale bar, 1 mm.
Figure 4
Figure 4
Cytokines Induce R-ISCs via JAK/STAT-1 (A) Live mTert-GFP+ cells from vehicle (PBS)- and IFN-γ-treated enteroid cultures after 48 hr. Representative images of mTert-GFP enteroid cultures 48 hr after vehicle or IFN-γ. Arrows identify mTert-GFP+ cells. Enteroid lines derived from N = 3 mice. Scale bar, 50 μm. (B) Live Lgr5-GFPhi cells from vehicle and IFN-γ-treated enteroid cultures after 48 hr. Representative images of Lgr5-GFP enteroid cultures 48 hr after vehicle or IFN-γ. Arrows identify Lgr5-GFP+ cells. Enteroid lines derived from N = 3 mice. Scale bar, 50 μm. (C) mTert-GFP+ crypt cells that co-express pSTAT-1 at baseline (N = 7 mice), early recovery (N = 4 mice), and late recovery (N = 3 mice). Representative optical sections showing an mTert-GFP+ pSTAT-1 cell (arrow) in IgG-treated and an mTert-GFP+ pSTAT-1+ cell (arrowhead) in αCD3-treated mice at early recovery in DAPI, mTert-GFP, and pSTAT-1 merged channels. Scale bar, 10 μm. (D) Fold change in mTert-GFP+ enteroid cells following treatment with vehicle, JAK/STAT-1 inhibitor tofacitinib (TOFA) only, IFN-γ only, and IFN-γ + TOFA. Enteroid lines derived from N = 3 mice. Error bars indicate the SEM. ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.

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References

    1. Barker N., van Es J.H., Kuipers J., Kujala P., van den Born M., Cozijnsen M., Haegebarth A., Korving J., Begthel H., Peters P.J. Identification of stem cells in small intestine and colon by marker gene Lgr5. Nature. 2007;449:1003–1007. - PubMed
    1. Barrios-Rodiles M., Chadee K. Novel regulation of cyclooxygenase-2 expression and prostaglandin E2 production by IFN-gamma in human macrophages. J. Immunol. 1998;161:2441–2448. - PubMed
    1. Beaurepaire C., Smyth D., McKay D.M. Interferon-gamma regulation of intestinal epithelial permeability. J. Interferon Cytokine Res. 2009;29:133–144. - PubMed
    1. Biteau B., Hochmuth C.E., Jasper H. Maintaining tissue homeostasis: dynamic control of somatic stem cell activity. Cell Stem Cell. 2011;9:402–411. - PMC - PubMed
    1. Breault D.T., Min I.M., Carlone D.L., Farilla L.G., Ambruzs D.M., Henderson D.E., Algra S., Montgomery R.K., Wagers A.J., Hole N. Generation of mTert-GFP mice as a model to identify and study tissue progenitor cells. Proc. Natl. Acad. Sci. USA. 2008;105:10420–10425. - PMC - PubMed

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