Smoothened is a master regulator of adult liver repair
Gregory A. Michelotti, … , Daniel Metzger, Anna Mae Diehl
Gregory A. Michelotti, … , Daniel Metzger, Anna Mae Diehl
Published April 8, 2013
Citation Information: J Clin Invest. 2013;123(6):2380-2394. https://doi.org/10.1172/JCI66904.
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Research Article Hepatology

Smoothened is a master regulator of adult liver repair

Abstract

When regenerative processes cannot keep pace with cell death, functional epithelia are replaced by scar. Scarring is characterized by both excessive accumulation of fibrous matrix and persistent outgrowth of cell types that accumulate transiently during successful wound healing, including myofibroblasts (MFs) and progenitors. This suggests that signaling that normally directs these cells to repair injured epithelia is deregulated. To evaluate this possibility, we examined liver repair during different types of liver injury after Smoothened (SMO), an obligate intermediate in the Hedgehog (Hh) signaling pathway, was conditionally deleted in cells expressing the MF-associated gene, αSMA. Surprisingly, blocking canonical Hh signaling in MFs not only inhibited liver fibrosis but also prevented accumulation of liver progenitors. Hh-sensitive, hepatic stellate cells (HSCs) were identified as the source of both MFs and progenitors by lineage-tracing studies in 3 other strains of mice, coupled with analysis of highly pure HSC preparations using flow cytometry, immunofluorescence confocal microscopy, RT-PCR, and in situ hybridization. The results identify SMO as a master regulator of hepatic epithelial regeneration based on its ability to promote mesenchymal-to-epithelial transitions in a subpopulation of HSC-derived MFs with features of multipotent progenitors.

Authors

Gregory A. Michelotti, Guanhua Xie, Marzena Swiderska, Steve S. Choi, Gamze Karaca, Leandi Krüger, Richard Premont, Liu Yang, Wing-Kin Syn, Daniel Metzger, Anna Mae Diehl

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Figure 4

Blocking Hh signaling in MFs inhibits accumulation of liver epithelial progenitors, causes liver atrophy, and blocks liver cell proliferation after BDL injury.

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Blocking Hh signaling in MFs inhibits accumulation of liver epithelial p...
(A) SOX9 protein expression in representative immunostained liver sections from vehicle-treated or TMX-treated DTG mice 14 days after BDL (original magnification, ×20). (B) Representative liver sections immunostained for AFP (original magnification, ×20). (C) qRT-PCR analysis of Sox9, Afp, CD133, and Nanog in whole liver RNA of DTG animals. Results are expressed as mean ± SEM relative to vehicle-treated BDL group. **P < 0.01; *P < 0.05. (D) Liver/body weight (LW/BW) ratios, serum levels of alanine aminotransferase (ALT), and TUNEL-positive cells in intact liver tissue 14 days after BDL in DTG mice treated with vehicle or TMX. Results are graphed as mean ± SEM. *P < 0.05. (E) Ki67 staining in representative vehicle-treated and TMX-treated mice 14 days after BDL. Ki67-positive cells were quantified and graphed as mean ± SEM relative to the vehicle-treated sham-operated group. *P < 0.05 vs. vehicle-treated BDL group (original magnification, ×20). (F) Representative cyclin D1 (CycD1) staining is similarly shown, with cyclin D1–positive cells graphed as mean ± SEM relative the vehicle-treated sham-operated group (original magnification, ×20). *P < 0.05 vs. vehicle-treated BDL group. (G) qRT-PCR analysis of cyclin D1 (Ccnd1) and Foxm1 expression in whole liver mRNA. Results in TMX-treated BDL group are shown relative to results in vehicle-treated BDL group (mean ± SEM). *P < 0.05.