GDF-5 Mice Disc Degeneration Model: The Spine Journal, epub 2009 Nov

[Slackwater]

Pubmed: 19926342

Spine Journal, Article in Press
© 2009 Elsevier Inc. All rights reserved.
doi:10.1016/j.spinee.2009.10.006


2009 Outstanding Paper: Runner-up

Therapeutic effects of adenovirus-mediated growth and differentiation factor-5 in a mice disc degeneration model induced by annulus needle puncture
Haixiang Liang, MD, MSa, Shen-Ying Ma, MDa, Gang Feng, MD, PhDa,b, Francis H. Shen, MDa,
Xudong Joshua Li, MD, PhDa,*
aDepartment of Orthopaedic Surgery, School of Medicine, University of Virginia, Charlottesville, VA 22908, USA
bResearch Institute of Tissue Engineering and Stem Cell, Nanchong Central Hospital, North Sichuan Medical College, Nanchong 637000, Sichuan Province, PR China

Received 5 February 2009; revised 28 September 2009; accepted 13 October 2009

Abstract

BACKGROUND CONTEXT:The therapeutic strategies that have thus far been used for the treatment
of intervertebral disc degeneration (IDD) have focused on relieving the symptoms, although
reversal of the degeneration remains an important challenge for the effective treatment of IDD.
Growth and differentiation factor-5 (GDF5), of which deficiency leads to early disc degeneration
changes, has the potential to increase proliferation of disc cells and expression of extracellular
matrix proteins.

PURPOSE:The purpose of the study was to develop a lumbar disc degeneration model in mice
and determine the effect of adenoviral GDF5 gene therapy.

STUDY DESIGN:The study design was to compare the degeneration changes of discs punctured
by different-size needles to develop a mice lumbar disc degeneration model and to evaluate the
effects of in vivo gene therapy for the mice disc degeneration model by an adenoviral vector
carrying GDF5 gene.

METHODS:A lumbar disc degeneration model was developed by needle punctures to the discs in
Balb/c mice. Afterward, a gene therapy treatment to disc degeneration was evaluated. Two of the
mice lumbar discs were randomly chosen to be punctured by a 30-gauge needle and then injected
with adenovirus that had been engineered to express either the luciferase gene (Ad-Luc) or the
GDF5 gene (Ad-GDF5). Animals were analyzed by bioluminescent imaging, radiographic, and
magnetic resonance imaging (MRI) scanning, then sacrificed at 1, 2, 4, or 8 weeks after operation,
and subjected to histological and biochemical assays.

RESULTS:By the detection of T2-weighted MRI scanning and histological study, the degeneration
was found in all of the discs punctured by different-size needles. But the development of the
degeneration in the discs injured by the 30-gauge needle was more reliable and moderate compared
with that in other groups. The detection of luciferase activity by bioluminescent imaging revealed
that adenovirus survived and the introduced genes were expressed over 6 weeks after injection.
There were no T2-weighted MRI signals in the mice injected with either Ad-Luc or Ad-GDF5
up to 4 weeks after operation. At 6 and 8 weeks, T2-weighted signals were detected in the
Ad-GDF5 group but none in the Ad-Luc control group. The percent disc height index (%DHI)
was significantly decreased (~20%) by 1 week after injury in both groups, indicating the development
of disc degeneration. At 2 weeks, the %DHI in the mice injected with Ad-GDF5 increased
significantly compared with that of the mice injected with Ad-Luc; the increase was sustained
for the rest of the experiment period. The disc histology treated with Ad-GDF5 was improved compared
with that in the control group. Glycosaminoglycan (GAG) levels were significantly decreased
in the Ad-Luc injection group since 2 weeks after injury, and the DNA content had diminished by 4
weeks after the operation. In contrast, in the discs injected with Ad-GDF5, there was no decrease in
the GAG and DNA levels after injury throughout the 8-week treatment period.

CONCLUSIONS:Disc degeneration animal model can be developed by using needle puncture to
the discs in mice. The adenovirus is an effective vehicle for gene delivery with rapid and prolonged
expression of target protein and resulting improvement in markers of disc degeneration. Ad-GDF5
gene therapy could restore the functions of injured discs and has the potential to be an effective
treatment.


Keywords: Intervertebral disc; Degeneration; Gene therapy; Growth factor


Article Outline
Introduction
Materials and methods

Preparation of the adenoviral vector
Study animals
Surgical procedures and in vivo transduction
In vivo bioluminescence imaging
Radiographic analyses
Magnetic resonance imaging
Biochemical assay
Histological evaluations
Statistical analysis

Results

Mice IDD model evaluation
Magnetic resonance imaging
Histology images of IDD model
Evaluation of gene therapeutic effect of Ad-GDF5
In vivo bioluminescence imaging
Radiographic analyses
Magnetic resonance imaging
Biochemical assay
Histology image

Discussion
References


Discussion
...
"Our research reports the curative effects of gene therapy

for the treatment of IDD in a novel mice model by adenoviral
mediation of human GDF5 gene. The results of this
study not only confirmed the long-term expression of the
target protein in the disc in vivo but also highlighted the
physiological improvements that occurred to the disc. Our
research also confirmed that this IDD model developed in
mice will be suitable for the study of the physiopathologic
changes and the effect of therapeutic intervention."

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