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Bovine explant model of degeneration of the intervertebral disc.

Roberts S, Menage J, Sivan S, Urban JP - BMC Musculoskelet Disord (2008)

Bottom Line: The central region of both papain and trypsin treated discs was macro- and microscopically fragmented, with severe loss of metachromasia.The integrity of the surrounding tissue was mostly in tact with cells in the outer annulus appearing viable.Biochemical analysis demonstrated greatly reduced glycosaminoglycan content in these compared to untreated discs.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Spinal Studies, Robert Jones and Agnes Hunt Orthopaedic Hospital, Oswestry, Shropshire SY10 7AG, UK. sally.roberts@rjah.nhs.uk

ABSTRACT

Background: Many new treatments for degeneration of the intervertebral disc are being developed which can be delivered through a needle. These require testing in model systems before being used in human patients. Unfortunately, because of differences in anatomy, there are no ideal animal models of disc degeneration. Bovine explant model systems have many advantages but it is not possible to inject any significant volume into an intact disc. Therefore we have attempted to mimic disc degeneration in an explant bovine model via enzymatic digestion.

Methods: Bovine coccygeal discs were incubated with different concentrations of the proteolytic enzymes, trypsin and papain, and maintained in culture for up to 3 weeks. A radio-opaque solution was injected to visualise cavities generated. Degenerative features were monitored histologically and biochemically (water and glycosaminoglycan content, via dimethylmethylene blue).

Results and conclusion: The central region of both papain and trypsin treated discs was macro- and microscopically fragmented, with severe loss of metachromasia. The integrity of the surrounding tissue was mostly in tact with cells in the outer annulus appearing viable. Biochemical analysis demonstrated greatly reduced glycosaminoglycan content in these compared to untreated discs. We have shown that bovine coccygeal discs, treated with proteolytic enzymes can provide a useful in vitro model system for developing and testing potential new treatments of disc degeneration, such as injectable implants or biological therapies.

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The profile from outer annulus to the central nucleus pulposus of (a) water and (b) GAG contents of fresh normal, bovine disc (i.e. not injected with enzyme or cultured).
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Figure 5: The profile from outer annulus to the central nucleus pulposus of (a) water and (b) GAG contents of fresh normal, bovine disc (i.e. not injected with enzyme or cultured).

Mentions: The water and GAG contents of untreated, fresh (normal) bovine discs was similar to the control discs and, as reported many times previously, higher in the central nucleus pulposus than in the annulus fibrosus. Profiles of water content showed no significant difference from those in any of the discs which had been enzymatically digested (at any time points; Figure 5). The profiles of GAG contents, in contrast, were much lower for both enzymes used, but were particularly reduced for papain treated discs, which all had a GAG content lower than 10% CS/dry weight (compared to 15–30% of normal discs in the nucleus). Trypsin treated discs reached this level 2 weeks after the enzyme had been introduced.


Bovine explant model of degeneration of the intervertebral disc.

Roberts S, Menage J, Sivan S, Urban JP - BMC Musculoskelet Disord (2008)

The profile from outer annulus to the central nucleus pulposus of (a) water and (b) GAG contents of fresh normal, bovine disc (i.e. not injected with enzyme or cultured).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2266744&req=5

Figure 5: The profile from outer annulus to the central nucleus pulposus of (a) water and (b) GAG contents of fresh normal, bovine disc (i.e. not injected with enzyme or cultured).
Mentions: The water and GAG contents of untreated, fresh (normal) bovine discs was similar to the control discs and, as reported many times previously, higher in the central nucleus pulposus than in the annulus fibrosus. Profiles of water content showed no significant difference from those in any of the discs which had been enzymatically digested (at any time points; Figure 5). The profiles of GAG contents, in contrast, were much lower for both enzymes used, but were particularly reduced for papain treated discs, which all had a GAG content lower than 10% CS/dry weight (compared to 15–30% of normal discs in the nucleus). Trypsin treated discs reached this level 2 weeks after the enzyme had been introduced.

Bottom Line: The central region of both papain and trypsin treated discs was macro- and microscopically fragmented, with severe loss of metachromasia.The integrity of the surrounding tissue was mostly in tact with cells in the outer annulus appearing viable.Biochemical analysis demonstrated greatly reduced glycosaminoglycan content in these compared to untreated discs.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre for Spinal Studies, Robert Jones and Agnes Hunt Orthopaedic Hospital, Oswestry, Shropshire SY10 7AG, UK. sally.roberts@rjah.nhs.uk

ABSTRACT

Background: Many new treatments for degeneration of the intervertebral disc are being developed which can be delivered through a needle. These require testing in model systems before being used in human patients. Unfortunately, because of differences in anatomy, there are no ideal animal models of disc degeneration. Bovine explant model systems have many advantages but it is not possible to inject any significant volume into an intact disc. Therefore we have attempted to mimic disc degeneration in an explant bovine model via enzymatic digestion.

Methods: Bovine coccygeal discs were incubated with different concentrations of the proteolytic enzymes, trypsin and papain, and maintained in culture for up to 3 weeks. A radio-opaque solution was injected to visualise cavities generated. Degenerative features were monitored histologically and biochemically (water and glycosaminoglycan content, via dimethylmethylene blue).

Results and conclusion: The central region of both papain and trypsin treated discs was macro- and microscopically fragmented, with severe loss of metachromasia. The integrity of the surrounding tissue was mostly in tact with cells in the outer annulus appearing viable. Biochemical analysis demonstrated greatly reduced glycosaminoglycan content in these compared to untreated discs. We have shown that bovine coccygeal discs, treated with proteolytic enzymes can provide a useful in vitro model system for developing and testing potential new treatments of disc degeneration, such as injectable implants or biological therapies.

Show MeSH
Related in: MedlinePlus