title: [Altered expression of endogenous transforming growth factor β1 and early calcification related genes in rat endplate].
abstract: OBJECTIVE
  To explore the relationship between endogenous transforming growth factor (TGF)-β1 and calcification-related genes through an in vitro degeneration model by propagating rat endplate chondrocytes during a natural degeneration process.
  
  
  METHODS
  Endplate chondrocytes were extracted from rat lumbar vertebrae, isolated by enzyme digestion and P2 and P4 generations selected for a 6-day in vitro culture. The specimens were photographed microscopically to observe the cellular differences by alizarin red staining. Type II collagen marker gene, transcription factor SOX-9 gene and metabolism-related genes proteoglycan. matrix metalloproteinase (MMP)-13, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4 and ADAMTS-5 were detected by RT-PCR to verify the degeneration model. Based on this model, the changes of growth factor TGF-β1 and calcification-related genes ankyrin (ANK), ectonucleotide pyrophosphatase (ENPP), tissue-nonspecific alkaline phosphatase (TNAP) were continuously tested.
  
  
  RESULTS
  Compared with P2 cells, P4 cells tended to assume a spindle-shaped morphology. And alizarin red staining showed no change between them. The level of transcription factor SOX-9 of P4 cells (P4/P2 = 0.0690, P = 0.0489) was significantly lower than that of P2 cells. Type II collagen (P4/P2 = 0.0535, P = 0.009) and proteoglycan (P4/P2 = 0.2672, P = 0.0343) were also significantly lower than those of P2 cells. No significant changes were observed in other metabolism-related genes. TGF-β1 (P4/P2 = 0.5934, P = 0.0482) was significantly lower. The expressions of TNAP (P4/P2 = 0.0385, P = 0.0139) and ANK (P4/P2 = 0.2121, P = 0.0009) were significantly lower. But ENPP showed no significant change.
  
  
  CONCLUSIONS
  P4 endplate chondrocytes undergo natural degeneration in vitro with the rising passage number. Type II collagen, SOX-9 and proteoglycan are significantly reduced. Endogenous TGF-β1 gene and calcification-related genes are down-regulated. The decrease of ANK gene may be caused by the down-regulation of endogenous TGF-β1. Modulating the expression of endogenous TGF-β1 gene in endplate chondrocytes may become a new therapeutic approach for the degeneration of intervertebral disc.
authors.affiliation: Department of Orthopedic Surgery, Yijishan Hospital, Wannan Medical College, Wuhu 241001, China. xuhg@medmail.com.cn
authors.forename: Hong-Guang
authors.initials: HG
authors.lastname: Xu
authors.forename: Xiao-Ling
authors.initials: XL
authors.lastname: Zhang
authors.forename: Xiao-Hai
authors.initials: XH
authors.lastname: Zhang
authors.forename: Hong
authors.initials: H
authors.lastname: Wang
authors.forename: Rui-Ping
authors.initials: RP
authors.lastname: Jia
authors.forename: Wen-Xue
authors.initials: WX
authors.lastname: Tong
authors.forename: Hai-Jun
authors.initials: HJ
authors.lastname: Tong
mesh_terms: D000818:Animals; D017487:Ankyrins / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D024043:Collagen Type II / Q000378:metabolism*; D011509:Proteoglycans / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D055758:SOX9 Transcription Factor / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000235:genetics / Q000378:metabolism*
chemical_list: D017487:Ankyrins; D024043:Collagen Type II; D011509:Proteoglycans; D055758:SOX9 Transcription Factor; D053773:Transforming Growth Factor beta1