TY - JOUR
T1 - DKK1AND FRZB are necessary for chondrocyte (RE) differentiation and prevention of cell hypertrophy in 3D cultures of human chondrocytes and human mesenchymal stem cells
AU - Zhong, L
AU - Huang, X
AU - Rodrigues, E
AU - Leijten, J
AU - Verrips, T.
AU - El Khattabi, M.
AU - Karperien, M
AU - Post, J
N1 - part of special issue: Abstract from the 2016 OARSI World Congress on Osteoarthritis: Promoting Clinical and Basic Research in Osteoarthritis
PY - 2016/4
Y1 - 2016/4
N2 - and differentiation was induced by keeping the cells for 21 days in a-MEM supplemented with 10% FBS, 1% sodium pyruvate, 1% antibiotic-antimycotic, 10mM b-glycerophosphate (BGP) and 50mg/ml ascorbic acid-2-phosphate (AA). Alkaline phosphatase activity (ALP) was meas-ured and mineralization was analyzed by alizarin red staining. Gene expression of extracellular matrix genes, genes linked to mineralization and TGFb superfamily genes was analyzed by RT-qPCR. Intracellular signaling pathways (PKCa, ERK1/2, SMAD1/5/8, p38MAPK) were investigated by Western blot. HPDCS were cultured for 21 days in DMEM (þ the same supplements as for MC3T3 differentiation and 100nM dexamethasone). HUVECS were stimulated in EGM-2 medium with 200 ng/ml of BMP6, to trigger endothelial to mesenchymal tran-sition. Then cells were cultured as hPDCS. hPDCs and HUVECS were cultured with supernatants harvested from control, Smoc2þ and DCaBD MC3T3 cells. Results: Mineralization and ALP activity was reduced in Smoc2þ MC3T3 cells compared to controls. Gene expression analysis showed an overall altered differentiation when overexpressing Smoc2. The acti-vation status of PKCa, ERK1/2, p38MAPK and SMAD1/5/8 was sig-nificantly modified in Smoc2þ cells. On the other hand, we could not observe an effect on osteogenesis when silencing Smoc2. DCaBD cells
AB - and differentiation was induced by keeping the cells for 21 days in a-MEM supplemented with 10% FBS, 1% sodium pyruvate, 1% antibiotic-antimycotic, 10mM b-glycerophosphate (BGP) and 50mg/ml ascorbic acid-2-phosphate (AA). Alkaline phosphatase activity (ALP) was meas-ured and mineralization was analyzed by alizarin red staining. Gene expression of extracellular matrix genes, genes linked to mineralization and TGFb superfamily genes was analyzed by RT-qPCR. Intracellular signaling pathways (PKCa, ERK1/2, SMAD1/5/8, p38MAPK) were investigated by Western blot. HPDCS were cultured for 21 days in DMEM (þ the same supplements as for MC3T3 differentiation and 100nM dexamethasone). HUVECS were stimulated in EGM-2 medium with 200 ng/ml of BMP6, to trigger endothelial to mesenchymal tran-sition. Then cells were cultured as hPDCS. hPDCs and HUVECS were cultured with supernatants harvested from control, Smoc2þ and DCaBD MC3T3 cells. Results: Mineralization and ALP activity was reduced in Smoc2þ MC3T3 cells compared to controls. Gene expression analysis showed an overall altered differentiation when overexpressing Smoc2. The acti-vation status of PKCa, ERK1/2, p38MAPK and SMAD1/5/8 was sig-nificantly modified in Smoc2þ cells. On the other hand, we could not observe an effect on osteogenesis when silencing Smoc2. DCaBD cells
U2 - 10.1016/j.joca.2016.01.278
DO - 10.1016/j.joca.2016.01.278
M3 - Meeting Abstract
SN - 1063-4584
VL - 24
SP - S142
JO - Osteoarthritis and cartilage
JF - Osteoarthritis and cartilage
IS - supplement 1
M1 - 229
T2 - 2016 OARSI World Congress on Osteoarthrtis
Y2 - 31 March 2016 through 3 April 2016
ER -