title: Transforming growth factor β controls CCN3 expression in nucleus pulposus cells of the intervertebral disc.
abstract: OBJECTIVE
  To investigate transforming growth factor β (TGFβ) regulation of CCN3 expression in cells of the nucleus pulposus.
  
  
  METHODS
  Real-time reverse transcription-polymerase chain reaction and Western blot analyses were used to measure CCN3 expression in the nucleus pulposus. Transfections were used to measure the effect of Smad3, MAPKs, and activator protein 1 (AP-1) on TGFβ-mediated CCN3 promoter activity. Lentiviral knockdown of Smad3 was performed to assess the role of Smad3 in CCN3 expression.
  
  
  RESULTS
  CCN3 was expressed in embryonic and adult intervertebral discs. TGFβ decreased the expression of CCN3 and suppressed its promoter activity in nucleus pulposus cells. DN-Smad3, Smad3 small interfering RNA, or DN-AP-1 had little effect on TGFβ suppression of CCN3 promoter activity. However, p38 and ERK inhibitors blocked suppression of CCN3 by TGFβ, suggesting involvement of these signaling pathways in the regulation of CCN3. Interestingly, overexpression of Smad3 in the absence of TGFβ increased CCN3 promoter activity. We validated the role of Smad3 in controlling CCN3 expression in Smad3-null mice and in nucleus pulposus cells transduced with lentiviral short hairpin Smad3. In terms of function, treatment with recombinant CCN3 showed a dose-dependent decrease in the proliferation of nucleus pulposus cells. Moreover, CCN3-treated cells showed a decrease in aggrecan, versican, CCN2, and type I collagen expression.
  
  
  CONCLUSIONS
  The opposing effect of TGFβ on CCN2 and CCN3 expression and the suppression of CCN2 by CCN3 in nucleus pulposus cells further the paradigm that these CCN proteins form an interacting triad, which is possibly important in maintaining extracellular matrix homeostasis and cell numbers.
authors: Department of Orthopaedic Surgery, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors: Cassie M
authors: CM
authors: Tran
authors: Harvey E
authors: HE
authors: Smith
authors: Aviva
authors: A
authors: Symes
authors: Laure
authors: L
authors: Rittié
authors: Bernard
authors: B
authors: Perbal
authors: Irving M
authors: IM
authors: Shapiro
authors: Makarand V
authors: MV
authors: Risbud
references: J Struct Biol. 2003 Sep;143(3):242-57
references: 14572479
references: J Biol Chem. 2004 Aug 27;279(35):36943-50
references: 15213231
references: Int J Biochem Cell Biol. 2009 Apr;41(4):771-83
references: 18775791
references: Angiogenesis. 2007;10(1):1-11
references: 17149534
references: J Cell Biochem. 2006 May 1;98(1):152-9
references: 16408279
references: J Cell Commun Signal. 2008 Jun;2(1-2):3-7
references: 18784988
references: J Bone Joint Surg Am. 2006 Apr;88 Suppl 2:10-4
references: 16595436
references: J Cell Biochem. 2007 Aug 15;101(6):1475-91
references: 17340618
references: J Bone Joint Surg Am. 2006 Apr;88 Suppl 2:52-7
references: 16595444
references: Biochem J. 2009 May 27;420(3):413-20
references: 19298220
references: J Orthop Surg Res. 2006 Oct 26;1:9
references: 17150129
references: BMC Dev Biol. 2008 Feb 20;8:18
references: 18289368
references: Cell Commun Signal. 2004 Sep 10;2(1):9
references: 15361251
references: Arterioscler Thromb Vasc Biol. 2010 Apr;30(4):675-82
references: 20139355
references: Arthritis Res Ther. 2003;5(3):120-30
references: 12723977
references: J Biol Chem. 2002 Oct 25;277(43):41220-9
references: 12149257
references: J Bone Joint Surg Am. 2006 Apr;88 Suppl 2:25-9
references: 16595439
references: Mol Cell Biol. 1992 Jan;12(1):10-21
references: 1309586
references: Am J Pathol. 2009 May;174(5):1725-34
references: 19359517
references: J Cell Commun Signal. 2009 Jun;3(2):135-45
references: 19626464
references: J Bone Miner Res. 2008 Nov;23(11):1751-64
references: 18597638
references: Arthritis Res Ther. 2008;10(6):R140
references: 19061498
references: Biofactors. 2009 Mar-Apr;35(2):200-8
references: 19449449
references: Arthritis Rheum. 2010 Jul;62(7):1983-92
references: 20222112
references: J Biol Chem. 2002 Aug 16;277(33):29399-405
references: 12050162
references: J Bone Miner Res. 2005 Dec;20(12):2213-23
references: 16294274
references: J Cell Commun Signal. 2007 Sep;1(2):91-102
references: 18481200
references: J Cell Commun Signal. 2009 Mar;3(1):25-41
references: 19424823
references: Trends Biochem Sci. 2008 Oct;33(10):461-73
references: 18789696
references: J Korean Neurosurg Soc. 2008 Mar;43(3):149-54
references: 19096623