Osteogenic protein-1 with transforming growth factor-beta1: potent inducer of chondrogenesis of synovial mesenchymal stem cells in vitro.
BACKGROUND
Recently, cells derived from synovial mesenchymal stem cells (MSCs) have been regarded as a potential source of cells to induce repair of articular cartilage. To investigate more effective methods for promoting chondrogenesis, we examined the effects of osteogenic protein (OP)-1 with or without transforming growth factor-beta (TGFbeta1) on chondrogenesis of human MSCs in vitro.


METHODS
MSCs were isolated from the synovial membrane of patients with rheumatoid arthritis undergoing knee replacement surgery. After expansion of the cells, pellet cultures were performed in chondrogenic medium with OP-1 100-200 ng/ml, TGFbeta1 10 ng/ml, or both agents for 3 or 6 weeks. Chondrogenesis was evaluated histologically with safranin O staining, reverse transcription polymerase chain reaction for aggrecan and type II collagen mRNA, and quantification of glycosaminoglycan (GAG) content using a dimethylmethylene blue dye-binding assay. GAG content was normalized by DNA content measured using Hoechst 33258 dye.


RESULTS
At 3 weeks of culture, mRNAs for type II collagen and aggrecan were expressed by MSCs treated with either TGFbeta1 or OP-1; however, substantial matrix production was not induced. At 6 weeks, OP-1 increased GAG accumulation dose-dependently in the presence or absence of TGFbeta1, and the GAG content was the highest after combined treatment with 200 ng OP-1 and TGFbeta1. Histological staining for safranin O was poor after treatment with OP-1 or TGFbeta1 alone and slightly increased after combined treatment with TGFbeta1 and OP-1 at 3 weeks. At 6 weeks, OP-1 increased the intensity of staining dose-dependently in the presence or absence of TGFbeta1. However, the histological appearance of the cells treated with OP-1 alone was similar to that of hypertrophic chondrocytes, which was different from that of cells with combined treatment with OP-1 and TGFbeta1.


CONCLUSIONS
A high dose of OP-1 was useful for enhancing chondrogenesis from synovium-derived MSCs in combined treatment with TGFbeta1.
Department of Orthopaedic Surgery, Nagasaki University, School of Medicine, 1-7-1 Sakamoto, Nagasaki, 852-8501, Japan.
Chikara
C
Miyamoto
Tomoko
T
Matsumoto
Koichiro
K
Sakimura
Hiroyuki
H
Shindo
D000328:Adult; D000368:Aged; D053674:Aggrecans / Q000096:biosynthesis / Q000235:genetics; D001172:Arthritis, Rheumatoid / Q000378:metabolism / Q000473:pathology / Q000628:therapy; D019645:Arthroplasty, Replacement, Knee; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins / Q000494:pharmacology*; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000187:drug effects* / Q000502:physiology; D024043:Collagen Type II / Q000096:biosynthesis / Q000235:genetics; D015870:Gene Expression; D006025:Glycosaminoglycans / Q000378:metabolism; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008875:Middle Aged; D018696:Neuroprotective Agents; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013583:Synovial Membrane / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D053781:Transforming Growth Factor beta2 / Q000494:pharmacology*
D053674:Aggrecans; C525819:BMP7 protein, human; D055419:Bone Morphogenetic Protein 7; D019485:Bone Morphogenetic Proteins; D024043:Collagen Type II; D006025:Glycosaminoglycans; D018696:Neuroprotective Agents; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D053781:Transforming Growth Factor beta2