Altered chondrocyte differentiation, matrix mineralization and MEK-Erk1/2 signaling in an INPPL1 catalytic knock-out mouse model of opsismodysplasia

Adv Biol Regul. 2020 May:76:100651. doi: 10.1016/j.jbior.2019.100651. Epub 2019 Sep 5.

Abstract

Opsismodysplasia (OPS) is a rare but severe autosomal recessive skeletal chondrodysplasia caused by inactivating mutations in the Inppl1/Ship2 gene. The molecular mechanism leading from Ship2 gene inactivation to OPS is currently unknown. Here, we used our Ship2Δ/Δ mouse expressing reduced amount of a catalytically-inactive SHIP2 protein and a previously reported SHIP2 inhibitor to investigate growth plate development and mineralization in vivo, ex vivo and in vitro. First, as observed in OPS patients, catalytic inactivation of SHIP2 in mouse leads to reduced body length, shortening of long bones, craniofacial dysmorphism, reduced height of the hyperthrophic chondrocyte zone and to defects in growth plate mineralization. Second, intrinsic Ship2Δ/Δ bone defects were sufficient to induce the characteristic OPS alterations in bone growth, histology and mineralization ex vivo. Third, expression of osteocalcin was significantly increased in SHIP2-inactivated chondrocyte cultures whereas production of mineralized nodules was markedly decreased. Targeting osteocalcin mRNA with a specific shRNA increased the production of mineralized nodules. Fourth, levels of p-MEK and p-Erk1/2 were significantly increased in SHIP2-inactivated chondrocytes in response to serum and IGF-1, but not to FGF2, as compared to control chondrocytes. Treatment of chondrocytes and bones in culture with a MEK inhibitor partially rescued the production of mineralized nodules, the size of the hypertrophic chondrocyte zone and bone growth, raising the possibility of a treatment that could partially reduce the phenotype of this severe condition. Altogether, our results indicate that Ship2Δ/Δ mice represent a relevant model for human OPS. They also highlight the important role of SHIP2 in chondrocytes during endochondral ossification and its different differentiation steps. Finally, we identified a role of osteocalcin in mineralized nodules production and for the MEK-Erk1/2 signaling pathway in the OPS phenotype.

Keywords: Bone mineraliation; Chondrocytes; Endochondral ossification; Inppl1; Opsismodysplasia; Osteocalcin; Ship2; Skeletal dysplasia.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aminoacetonitrile / analogs & derivatives
  • Aminoacetonitrile / pharmacology
  • Animals
  • Calcification, Physiologic / genetics
  • Cell Differentiation
  • Chondrocytes / metabolism*
  • Chondrocytes / pathology
  • Disease Models, Animal
  • Fibroblast Growth Factor 2 / pharmacology
  • Gene Expression Regulation
  • Growth Plate / metabolism
  • Growth Plate / pathology
  • Humans
  • Insulin-Like Growth Factor I / pharmacology
  • MAP Kinase Kinase Kinases / antagonists & inhibitors
  • MAP Kinase Kinase Kinases / genetics*
  • MAP Kinase Kinase Kinases / metabolism
  • Mice
  • Mice, Knockout
  • Mitogen-Activated Protein Kinase 1 / genetics*
  • Mitogen-Activated Protein Kinase 1 / metabolism
  • Mitogen-Activated Protein Kinase 3 / genetics*
  • Mitogen-Activated Protein Kinase 3 / metabolism
  • Osteocalcin / antagonists & inhibitors
  • Osteocalcin / genetics*
  • Osteocalcin / metabolism
  • Osteochondrodysplasias / genetics*
  • Osteochondrodysplasias / metabolism
  • Osteochondrodysplasias / pathology
  • Osteogenesis / genetics
  • Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases / antagonists & inhibitors
  • Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases / deficiency
  • Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases / genetics*
  • Phosphorylation / drug effects
  • Primary Cell Culture
  • RNA, Messenger / antagonists & inhibitors
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / metabolism
  • Signal Transduction
  • Thiophenes / pharmacology

Substances

  • 3-((4-chlorobenzyl)oxy)-N-((1S)-1-phenylethyl)-2-thiophenecarboxamide
  • RNA, Messenger
  • RNA, Small Interfering
  • SL 327
  • Thiophenes
  • insulin-like growth factor-1, mouse
  • Fibroblast Growth Factor 2
  • Osteocalcin
  • Aminoacetonitrile
  • Insulin-Like Growth Factor I
  • Mapk1 protein, mouse
  • Mapk3 protein, mouse
  • Mitogen-Activated Protein Kinase 1
  • Mitogen-Activated Protein Kinase 3
  • MAP Kinase Kinase Kinases
  • Inppl1 protein, mouse
  • Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases

Supplementary concepts

  • Opsismodysplasia