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O005 SMOC2 has differential effects on cartilage and bone formation
  1. T Peeters1,
  2. R Lories1,
  3. F Cailotto2
  1. 1Skeletal Biology and Engineering Research Centre – Dept. Development and Regeneration, KU LEUVEN, Leuven, Belgium
  2. 2IMoPA, Université de Lorraine, Nancy, France


Introduction SMOC2, a secreted calcium-binding protein of the BM-40/SPARC family was identified from a chondrogenic extract of articular cartilage and is increased in osteoarthritic cartilage. Alterations in bone and cartilage resulting from dysregulated signalling pathways are a hallmark of osteoarthritis.

Objectives To evaluate the role of SMOC2 in in vitro osteogenesis and chondrogenesis and its interaction with Wnt signalling.

Methods Wild-type Smoc2 (Smoc2+) or Smoc2 lacking the calcium binding domain (?CaBD) were stably overexpressed and wild-type Smoc2 was silenced (Smoc2-). Alkaline phosphatase (ALP) activity assay and Alizarin Red staining were performed in MC3T3 cells and human periosteal derived cells (hPDCs). Proteoglycan content and mineralization were assessed in ATDC5 micromasses by Alcian blue and Alizarin Red staining. Mice femoral head caps (FHC) were stimulated with recombinant IL-1b and SMOC2. Cartilage loss was evaluated by DMMB assay and Safranin O staining. TCF1 levels were analysed by immunohistochemistry. Human articular chondrocytes (hACs) were stimulated with recombinant SMOC2. Gene expression of osteoblast markers (Opn, Osx, Col1a2), chondrogenic markers (Acan, Col2a1, Col10a1, Mmp13) and canonical Wnt target genes (Axin2, Lef1) were analysed by qPCR and Wnt signalling by Western blot.

Results Alizarin red staining and ALP activity were reduced in Smoc2 +MC3 T3 cells. Osteoblast gene expression levels were altered in Smoc2 +cells. ?CaBD cells and calcium supplementation to Smoc2 +showed a partial rescue of the effects of Smoc2 +cells. We validated these results in hPDCs. We could not observe an effect on osteogenesis when silencing Smoc2. Smoc2 +ATDC5 micromasses showed reduced chondrogenic gene expression and Alcian blue and Alizarin Red staining intensity. We observed a reverse pattern with an increased chondrogenic gene expression in Smoc2- ATDC5 micromasses. Smoc2 +and Smoc2- ATDC5 cells altered canonical Wnt signalling in opposite directions. SMOC2 also alters protein levels of CamKII. IL-1b triggered cartilage loss was increased and TCF1 levels were reduced in mice FHC stimulated with recombinant SMOC2. In hACs, we observed a negative effect of SMOC2 on chondrocyte markers and canonical Wnt target genes.

Conclusions Our results suggest an inhibitory role of Smoc2 on mineralization. The inhibitory effects on mineralization are at least partially regulated through interaction with calcium. Moreover, Smoc2 is a regulator of chondrogenic differentiation and cartilage homeostasis by regulating canonical and non-canonical Wnt signalling.

Disclosure of interest None declared

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