DKK1AND FRZB are necessary for chondrocyte (RE) differentiation and prevention of cell hypertrophy in 3D cultures of human chondrocytes and human mesenchymal stem cells

L Zhong, X Huang, E Rodrigues, J Leijten, T. Verrips, M. El Khattabi, M Karperien, J Post

Research output: Contribution to journalMeeting AbstractOther research output

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Abstract

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
Original languageEnglish
Article number229
Pages (from-to)S142
JournalOsteoarthritis and cartilage
Volume24
Issue numbersupplement 1
DOIs
Publication statusPublished - Apr 2016
Event2016 OARSI World Congress on Osteoarthrtis - Amsterdam RAI, Amsterdam, Netherlands
Duration: 31 Mar 20163 Apr 2016

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Chondrocytes
Stem cells
Mesenchymal Stromal Cells
Cell culture
Hypertrophy
Cell Differentiation
Genes
Phosphatases
Gene expression
Alizarin
Ascorbic acid
Alkaline Phosphatase
Antibiotics
Phosphates
Glycerophosphates
Gene Expression
Sodium
MAP Kinase Signaling System
Pyruvic Acid
Osteogenesis

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@article{9faaf43f37b6421caba71a38a8dd5d62,
title = "DKK1AND FRZB are necessary for chondrocyte (RE) differentiation and prevention of cell hypertrophy in 3D cultures of human chondrocytes and human mesenchymal stem cells",
abstract = "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 ({\th} 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, Smoc2{\th} and DCaBD MC3T3 cells. Results: Mineralization and ALP activity was reduced in Smoc2{\th} 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 Smoc2{\th} cells. On the other hand, we could not observe an effect on osteogenesis when silencing Smoc2. DCaBD cells",
author = "L Zhong and X Huang and E Rodrigues and J Leijten and T. Verrips and {El Khattabi}, M. and M Karperien and J Post",
note = "part of special issue: Abstract from the 2016 OARSI World Congress on Osteoarthritis: Promoting Clinical and Basic Research in Osteoarthritis",
year = "2016",
month = "4",
doi = "10.1016/j.joca.2016.01.278",
language = "English",
volume = "24",
pages = "S142",
journal = "Osteoarthritis and cartilage",
issn = "1063-4584",
publisher = "W.B. Saunders Ltd",
number = "supplement 1",

}

DKK1AND FRZB are necessary for chondrocyte (RE) differentiation and prevention of cell hypertrophy in 3D cultures of human chondrocytes and human mesenchymal stem cells. / Zhong, L; Huang, X; Rodrigues, E; Leijten, J; Verrips, T.; El Khattabi, M.; Karperien, M; Post, J.

In: Osteoarthritis and cartilage, Vol. 24, No. supplement 1, 229, 04.2016, p. S142.

Research output: Contribution to journalMeeting AbstractOther research output

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

VL - 24

SP - S142

JO - Osteoarthritis and cartilage

JF - Osteoarthritis and cartilage

SN - 1063-4584

IS - supplement 1

M1 - 229

ER -