Gene expression profiling of early intervertebral disc degeneration reveals a down-regulation of canonical Wnt signaling and caveolin-1 expression: implications for development of regenerative strategies.
BACKGROUND
Early degeneration of the intervertebral disc (IVD) involves a change in cellular differentiation from notochordal cells (NCs) in the nucleus pulposus (NP) to chondrocyte-like cells (CLCs). The purpose of this study was to investigate the gene expression profiles involved in this process using NP tissue from non-chondrodystrophic and chondrodystrophic dogs, a species with naturally occurring IVD degeneration.


METHODS
Dual channel DNA microarrays were used to compare 1) healthy NP tissue containing only NCs (NC-rich), 2) NP tissue with a mixed population of NCs and CLCs (Mixed), and 3) NP tissue containing solely CLCs (CLC-rich) in both non-chondrodystrophic and chondrodystrophic dogs. Based on previous reports and the findings of the microarray analyses, canonical Wnt signaling was further evaluated using qPCR of relevant Wnt target genes. We hypothesized that caveolin-1, a regulator of Wnt signaling that showed significant changes in gene expression in the microarray analyses, played a significant role in early IVD degeneration. Caveolin-1 expression was investigated in IVD tissue sections and in cultured NCs. To investigate the significance of Caveolin-1 in IVD health and degeneration, the NP of 3-month-old Caveolin-1 knock-out mice was histopathologically evaluated and compared with the NP of wild-type mice of the same age.


RESULTS
Early IVD degeneration involved significant changes in numerous pathways, including Wnt/β-catenin signaling. With regard to Wnt/β-catenin signaling, axin2 gene expression was significantly higher in chondrodystrophic dogs compared with non-chondrodystrophic dogs. IVD degeneration involved significant down-regulation of axin2 gene expression. IVD degeneration involved significant down-regulation in Caveolin-1 gene and protein expression. NCs showed abundant caveolin-1 expression in vivo and in vitro, whereas CLCs did not. The NP of wild-type mice was rich in viable NCs, whereas the NP of Caveolin-1 knock-out mice contained chondroid-like matrix with mainly apoptotic, small, rounded cells.


CONCLUSIONS
Early IVD degeneration involves down-regulation of canonical Wnt signaling and Caveolin-1 expression, which appears to be essential to the physiology and preservation of NCs. Therefore, Caveolin-1 may be regarded an exciting target for developing strategies for IVD regeneration.
Lucas A
LA
Smolders
Björn P
BP
Meij
David
D
Onis
Frank M
FM
Riemers
Niklas
N
Bergknut
Richard
R
Wubbolts
Guy C M
GC
Grinwis
Martin
M
Houweling
Marian J A
MJ
Groot Koerkamp
Dik
D
van Leenen
Frank C P
FC
Holstege
Herman A W
HA
Hazewinkel
Laura B
LB
Creemers
Louis C
LC
Penning
Marianna A
MA
Tryfonidou
D000818:Animals; D051242:Caveolin 1 / Q000096:biosynthesis*; D004285:Dogs; D015536:Down-Regulation; D020869:Gene Expression Profiling; D048091:Guided Tissue Regeneration / Q000379:methods; D055959:Intervertebral Disc Degeneration / Q000235:genetics* / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D020411:Oligonucleotide Array Sequence Analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D060449:Wnt Signaling Pathway / Q000502:physiology*
D051242:Caveolin 1