J Korean Soc Spine Surg.  2002 Sep;9(3):165-171. 10.4184/jkss.2002.9.3.165.

Transforming Growth Factor-beta1 and Bone Morphogenetic Protein-2 Upregulates Matrix Synthesis and Chondrogenic Phenotype in Intervertebral Disc Cells

• Kim DJ
• Moon SH
• Kim H
• Kwon EH
• Hahn SB
• Cheon YM
• Kim HS
• Han KJ
• Park MS
• Lee HM

• KMID: 2003220
• DOI: http://doi.org/10.4184/jkss.2002.9.3.165

Abstract

OBJECTIVES
To determine effect of transforming growth factor-beta1 and bone morphogenetic protein-2 in matrix synthesis and expression of chondrogenic phenotype in human intervertebral disc cells.
MATERIALS AND METHODS
The intervertebral disc cells were harvested and cultured from the surgical patients for the degenerative disc disease. TGF-beta1 was purchased from R&D and BMP-2 was produced by transfection of pcDNA3.1/Hygro/BMP-2 to CHO cell using Lipofectamine 2000. rhBMP-2 was separated by Heparin-Sepharose A chromatography. TGF-bata1 and BMP-2 were administered to culture. Proteoglycan synthesis was assessed by 35S incorporation and expression of matrix mRNA was analyzed by RT-PCR for collagen I, collagen II, aggrecan, and osteocalcin.
RESULTS
TGF-bata1 and BMP-2 showed increased proteoglycan synthesis and expression of collagen I, collagen II and aggrecan mRNA in dose dependent manner respectively. There was no recognizable synergistic effect in matrix synthesis and matrix mRNA expression. Throughout dosage, expression of osteogenic phenotype (osteocalcin mRNA) was not noted.
CONCLUSION
TGF-beta1 and BMP-2 proved to be effective anabolic agent for maximizing matrix synthesis without evidence of osteogenesis.

Keyword

TGF-bata1; BMP-2; Aggrecan; Collagen I; Collagen II; Osteocalcin

MeSH Terms

Aggrecans
Animals
CHO Cells
Chromatography
Collagen
Cricetinae
Humans
Intervertebral Disc*
Osteocalcin
Osteogenesis
Phenotype*
Proteoglycans
RNA, Messenger
Transfection
Transforming Growth Factor beta1
Aggrecans
Collagen
Osteocalcin
Proteoglycans
RNA, Messenger
Transforming Growth Factor beta1

Figure

• 

  Fig. 1. Newly synthesized proteoglycan as measured by incorporation 35S-sulfate according to the dose of TGF-b1 and BMP-2. In given doses, proteoglycan synthesis was upregulated compared to control culture (p<0.05).

• 

  Fig. 2. Dose response of BMP-2 in mRNA expression of collagen type I, collagen type II, aggrecan, and osteoclacin as measured by reverse transcriptase-polymerase chain reaction. Collagen type I, collagen type II, and aggrecan mRNA expression were upregulated, while osteoclacin mRNA showed no recognizable upregulation.

• 

  Fig. 3. Effect of BMP-2 and TGF-b1 in mRNA expression of collagen type I, collagen type II, aggrecan, and osteoclacin as measured by reverse transcriptase-polymerase chain reaction. Even with combination of two growth factors, there was no synergistic effect in matrix mRNA expression. In given doses, there was no expression of osteocalcin mRNA.

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