The biological basis of degenerative disc disease: proteomic and biomechanical analysis of the canine intervertebral disc.
BACKGROUND
In the present study, we sought to quantify and contrast the secretome and biomechanical properties of the non-chondrodystrophic (NCD) and chondrodystrophic (CD) canine intervertebral disc (IVD) nucleus pulposus (NP).


METHODS
We used iTRAQ proteomic methods to quantify the secretome of both CD and NCD NP. Differential levels of proteins detected were further verified using immunohistochemistry, Western blotting, and proteoglycan extraction in order to evaluate the integrity of the small leucine-rich proteoglycans (SLRPs) decorin and biglycan. Additionally, we used robotic biomechanical testing to evaluate the biomechanical properties of spinal motion segments from both CD and NCD canines.


RESULTS
We detected differential levels of decorin, biglycan, and fibronectin, as well as of other important extracellular matrix (ECM)-related proteins, such as fibromodulin and HAPLN1 in the IVD NP obtained from CD canines compared with NCD canines. The core proteins of the vital SLRPs decorin and biglycan were fragmented in CD NP but were intact in the NP of the NCD animals. CD and NCD vertebral motion segments demonstrated significant differences, with the CD segments having less stiffness and a more varied range of motion.


CONCLUSIONS
The CD NP recapitulates key elements of human degenerative disc disease. Our data suggest that at least some of the compromised biomechanical properties of the degenerative disc arise from fibrocartilaginous metaplasia of the NP secondary to fragmentation of SLRP core proteins and associated degenerative changes affecting the ECM. This study demonstrates that the degenerative changes that naturally occur within the CD NP make this animal a valuable animal model with which to study IVD degeneration and potential biological therapeutics.
Toronto Western Research Institute, Toronto Western Hospital, Toronto, ON, M5T 2S8, Canada. mark.erwin@utoronto.ca.|Divisions of Neurological and Orthopaedic Surgery, University of Toronto, Toronto, ON, Canada. mark.erwin@utoronto.ca.|Canadian Memorial Chiropractic College, North York, ON, Canada. mark.erwin@utoronto.ca.
William Mark
WM
Erwin
Department of Chemistry, York University, Toronto, ON, Canada. leroi.desouza@hotmail.com.
Leroi
L
DeSouza
Department of Physical Therapy, University of Alberta, 8205 114 Street, 2-50 Corbett Hall, Edmonton, AB, T6G 2G4, Canada. funabash@ualberta.ca.
Martha
M
Funabashi
Department of Physical Therapy, University of Alberta, 8205 114 Street, 2-50 Corbett Hall, Edmonton, AB, T6G 2G4, Canada. greg.kawchuk@ualberta.ca.
Greg
G
Kawchuk
Toronto Western Research Institute, Toronto Western Hospital, Toronto, ON, M5T 2S8, Canada. mzkarim@uhnresearch.ca.
Muhammad Zia
MZ
Karim
Toronto Western Research Institute, Toronto Western Hospital, Toronto, ON, M5T 2S8, Canada. sj3kim@uwaterloo.ca.
Sarah
S
Kim
Department of Chemistry, York University, Toronto, ON, Canada. stefanie.maedler@gmail.com.
Stefanie
S
Mӓdler
Toronto Western Research Institute, Toronto Western Hospital, Toronto, ON, M5T 2S8, Canada. amatta@uhnresearch.ca.
Ajay
A
Matta
Toronto Western Research Institute, Toronto Western Hospital, Toronto, ON, M5T 2S8, Canada. xiaomeiw@uhnresearch.ca.
Xiaomei
X
Wang
Spinal Surgery, University of Basel, Basel, Switzerland. Arne.Mehrkens@usb.ch.
K Arne
KA
Mehrkens
D000818:Animals; D058578:Biglycan / Q000032:analysis / Q000378:metabolism; D001696:Biomechanical Phenomena; D015153:Blotting, Western; D058575:Decorin / Q000032:analysis / Q000378:metabolism; D004195:Disease Models, Animal; D004285:Dogs; D016326:Extracellular Matrix Proteins / Q000032:analysis / Q000378:metabolism; D005260:Female; D000071235:Fibromodulin; D005353:Fibronectins / Q000032:analysis / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D007403:Intervertebral Disc / Q000378:metabolism* / Q000503:physiopathology; D055959:Intervertebral Disc Degeneration / Q000378:metabolism* / Q000503:physiopathology; D008297:Male; D011509:Proteoglycans / Q000032:analysis / Q000378:metabolism; D020543:Proteome / Q000032:analysis* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*
D058578:Biglycan; D058575:Decorin; D016326:Extracellular Matrix Proteins; C000604330:FMOD protein, human; D005353:Fibronectins; D011509:Proteoglycans; D020543:Proteome; D000071235:Fibromodulin