R-spondin3-LGR4 signaling protects hepatocytes against DMOG-induced hypoxia/reoxygenation injury through activating β-catenin

Biochem Biophys Res Commun. 2018 Apr 30;499(1):59-65. doi: 10.1016/j.bbrc.2018.03.126. Epub 2018 Mar 19.

Abstract

Background & aims: Leucine-rich repeat G-protein-coupled receptor 4 (LGR4) and its ligands R-spondin1-4 (Rspos) have been vastly investigated in embryonic development. The biological functions of Rspos-LGR4 system in liver remains largely unknown. Here, we explored whether it protects hepatocytes against hypoxia/reoxygenation (H/R) induced damage.

Methods: H/R injury was induced by dimethyloxalylglycine (DMOG) in AML12 cells and the effects of Rspo3 on cell proliferation and apoptosis were assessed. Specific shRNAs were used to interfere LGR4 or β-catenin.

Results: DMOG caused hepatocytes damage evidenced by increase in HIF-1α, cell death and apoptosis genes p27 and Bax, with concurrent decrease of cell proliferation genes PCNA and CyclinD1. Of all the Rspos, Rspo3 is predominantly expressed in AML12 hepatocytes. Importantly, Rspo3 demonstrated an alteration in a manner similar to proliferation-related genes during H/R injury. Rspo3 pretreatment rendered hepatocytes less vulnerable to DMOG induced H/R injury. Ablation of LGR4 using shRNA attenuated the protective effects of Rspo3. Wnt3a also protected AML12 cells from damages caused by H/R, showing enhanced proliferation activity. Notably, knockdown of β-catenin in hepatocytes completely abolished the effect of Rspo3 pretreatment on the expression levels of PCNA and CyclinD1.

Conclusion: Rspo3-LGR4 axis protects hepatocytes from H/R injury via activating β-catenin.

Keywords: Hepatocytes; Hypoxia/reoxygenation injury; LGR4; R-spondin3; β-catenin.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acids, Dicarboxylic / pharmacology*
  • Animals
  • Apoptosis / drug effects
  • Cell Line
  • Cell Proliferation
  • Cyclin D1 / genetics
  • Cyclin D1 / metabolism
  • Cyclin-Dependent Kinase Inhibitor p27 / genetics
  • Cyclin-Dependent Kinase Inhibitor p27 / metabolism
  • Gene Expression Regulation
  • Hepatocytes / cytology
  • Hepatocytes / drug effects
  • Hepatocytes / metabolism*
  • Humans
  • Hypoxia-Inducible Factor 1, alpha Subunit / genetics
  • Hypoxia-Inducible Factor 1, alpha Subunit / metabolism
  • Mice
  • Mice, Transgenic
  • Proliferating Cell Nuclear Antigen / genetics
  • Proliferating Cell Nuclear Antigen / metabolism
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / metabolism
  • Receptors, G-Protein-Coupled / antagonists & inhibitors
  • Receptors, G-Protein-Coupled / genetics
  • Receptors, G-Protein-Coupled / metabolism*
  • Signal Transduction
  • Thrombospondins / genetics
  • Thrombospondins / metabolism*
  • Thrombospondins / pharmacology
  • Wnt3A Protein / genetics
  • Wnt3A Protein / metabolism
  • bcl-2-Associated X Protein / genetics
  • bcl-2-Associated X Protein / metabolism
  • beta Catenin / agonists
  • beta Catenin / antagonists & inhibitors
  • beta Catenin / genetics
  • beta Catenin / metabolism*

Substances

  • Amino Acids, Dicarboxylic
  • Bax protein, mouse
  • Ccnd1 protein, mouse
  • Hif1a protein, mouse
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • LGR4 protein, mouse
  • Proliferating Cell Nuclear Antigen
  • R-spondin3 protein, mouse
  • RNA, Small Interfering
  • Receptors, G-Protein-Coupled
  • Thrombospondins
  • Wnt3A Protein
  • Wnt3a protein, mouse
  • bcl-2-Associated X Protein
  • beta Catenin
  • Cyclin D1
  • Cyclin-Dependent Kinase Inhibitor p27
  • oxalylglycine