Potential use of Sox9 gene therapy for intervertebral degenerative disc disease.
METHODS
A new recombinant adenoviral vector expressing Sox9, a chondrocyte-specific transcription factor, was tested in a chondroblastic cell line and primary human intervertebral disc cells in vitro. Direct infection of intervertebral disc cells then was assessed in a rabbit model.


OBJECTIVE
To deliver a potentially therapeutic viral vector expressing Sox9 to degenerative human and rabbit intervertebral discs cells, and to assess the effect of Sox9 expression on Type 2 collagen production.


BACKGROUND
The concentration of competent Type 2 collagen, an essential constituent of the healthy nucleus pulposus, declines with intervertebral disc degeneration. Recent studies suggest that Sox9 upregulates Type 2 collagen production. Interventions that augment Type 2 collagen production by intervertebral disc cells may represent a novel therapeutic method for patients with degenerative disc disease.


METHODS
Adenoviral delivery vectors expressing Sox9 and green fluorescent protein were constructed using the AdEasy system. The chondroblastic cell line, HTB-94, and cultured human degenerated intervertebral disc cells were infected with the vectors. Reverse transcriptase-polymerase chain reaction and immunohistochemical analyses were performed to document increased Type 2 collagen expression. The AdSox9 virus then was injected directly into the intervertebral discs of three rabbits. After 5 weeks, the injected discs were evaluated histologically.


RESULTS
The AdSox9 virus efficiently transduced HTB-94 cells and degenerated human disc cells. Western blot analysis confirmed increased Sox9 production. Increased Type 2 collagen production was demonstrated in infected HTB-94 and human disc cells using both reverse transcriptase-polymerase chain reaction and immunohistochemical staining. In the rabbit model, cells infected with AdSox9 maintained a chondrocytic phenotype, and the architecture of the nucleus pulposus was preserved over a 5-week study period compared to control discs.


CONCLUSIONS
A novel adenoviral vector efficiently increased Sox9 and Type 2 collagen synthesis in cultured chondroblastic cells and human degenerated disc cells. In a rabbit model, sustained Sox9 production preserved the histologic appearance of the nucleus pulposus cells in vivo. These findings suggest a potential role for Sox9 gene therapy in the treatment of human degenerative disc disease.
Paul
Ronjon
R
Molecular Oncology Laboratory and the Spine Center, Department of Surgery, University of Chicago Medical Center, Chicago, Illinois 60640, USA.
Haydon
Rex C
RC
Cheng
Hongwei
H
Ishikawa
Akira
A
Nenadovich
Nikola
N
Jiang
Wei
W
Zhou
Lan
L
Breyer
Benjamin
B
Feng
Tao
T
Gupta
Purnendu
P
He
Tong-Chuan
TC
Phillips
Frank M
FM
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D024043:Collagen Type II / Q000096:biosynthesis; D004195:Disease Models, Animal; D004333:Drug Administration Routes; D015870:Gene Expression; D017930:Genes, Reporter; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage / Q000235:genetics; D049452:Green Fluorescent Proteins; D006609:High Mobility Group Proteins / Q000008:administration & dosage / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D007405:Intervertebral Disc Displacement / Q000378:metabolism / Q000473:pathology / Q000628:therapy*; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008164:Luminescent Proteins / Q000008:administration & dosage / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis; D011817:Rabbits; D055758:SOX9 Transcription Factor; D014157:Transcription Factors / Q000008:administration & dosage / Q000096:biosynthesis / Q000235:genetics*; D014161:Transduction, Genetic; D016896:Treatment Outcome
D024043:Collagen Type II; D006609:High Mobility Group Proteins; D008164:Luminescent Proteins; D012333:RNA, Messenger; D055758:SOX9 Transcription Factor; C527932:SOX9 protein, human; D014157:Transcription Factors; D049452:Green Fluorescent Proteins