[Effect of transforming growth factor beta3, bone morphogenetic protein 2, and dexamethasone on chondrogenic differentiation of rabbit synovial mesenchymal stem cells].
OBJECTIVE
To study the effect of transforming growth factor beta3 (TGF-beta3), bone morphogenetic protein 2 (BMP-2), and dexamethasone (DEX) on the chondrogenic differentiation of rabbit synovial mesenchymal stem cells (SMSCs).


METHODS
SMSCs were isolated from the knee joints of 5 rabbits (weighing, 1.8-2.5 kg), and were identified by morphogenetic observation, flow cytometry detection for cell surface antigen, and adipogenic and osteogenic differentiations. The SMSCs were cultured in the PELLET system for chondrogenic differentiation. The cell pellets were divided into 8 groups: TGF-beta3 was added in group A, BMP-2 in group B, DEX in group C, TGF-beta3 + BMP-2 in group C, TGF-beta3 + DEX in group E, BMP-2+ DEX in group F, and TGF-beta3 + BMP-2 + DEX in group G; group H served as control group. The diameter, weight, collagen type II (immuohistochemistry staining), proteoglycan (toluidine blue staining), and expression of cartilage related genes [real time quantitative PCR (RT-qPCR) technique] were compared to evaluate the effect of cytokines on the chondrogenic differentiation of SMSCs. Meanwhile, the DNA content of cell pellets was tested to assess the relationship between the increase weight of cell pellets and the cell proliferation.


RESULTS
SMSCs were isolated from the knee joints of rabbits successfully and the findings indicated that the rabbit synovium-derived cells had characteristics of mesenchymal stem cells. The diameter, weight, collagen type II, proteoglycan, and expression of cartilage related genes of pellets in groups A-F were significantly lower than those of group G (P < 0.05). RT-qPCR detection results showed that the relative expressions of cartilage related genes (SOX-9, Aggrecan, collagen type II, collagen type X, and BMP receptor II) in group G were significantly higher than those in the other groups (P < 0.01). Meanwhile, with the increase of the volume of pellet, the DNA content reduced about 70% at 7 days, about 80% at 14 days, and about 88% at 21 days.


CONCLUSIONS
The combination of TGF-beta3, BMP-2, and DEX can make the capacity of chondrogenesis of SMSCs maximized. The increase of the pellet volume is caused by the extracellular matrix rather than by cell proliferation.
Song
S
Chen
Peiliang
P
Fu
Ruijun
R
Cong
Haishan
H
Wu
Zhenyu
Z
Xu
D000818:Animals; D055396:Bone Morphogenetic Protein 2 / Q000008:administration & dosage / Q000494:pharmacology*; D002454:Cell Differentiation / Q000187:drug effects*; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000187:drug effects; D024043:Collagen Type II / Q000378:metabolism; D003907:Dexamethasone / Q000008:administration & dosage / Q000494:pharmacology*; D005260:Female; D005434:Flow Cytometry; D005786:Gene Expression Regulation / Q000187:drug effects; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D011509:Proteoglycans / Q000378:metabolism; D011817:Rabbits; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013583:Synovial Membrane / Q000166:cytology*; D023822:Tissue Engineering; D053782:Transforming Growth Factor beta3 / Q000008:administration & dosage / Q000494:pharmacology*
D055396:Bone Morphogenetic Protein 2; D024043:Collagen Type II; D011509:Proteoglycans; D053782:Transforming Growth Factor beta3; D003907:Dexamethasone