Clin Orthop Surg.  2009 Jun;1(2):81-89. 10.4055/cios.2009.1.2.81.

The Increased Expression of Matrix Metalloproteinases Associated with Elastin Degradation and Fibrosis of the Ligamentum Flavum in Patients with Lumbar Spinal Stenosis

• Park JB
• Kong CG
• Suhl KH
• Chang ED
• Riew KD

• KMID: 1122637
• DOI: http://doi.org/10.4055/cios.2009.1.2.81

Abstract

BACKGROUND: One of the characteristics of spinal stenosis is elastin degradation and fibrosis of the extracellular matrix of the ligamentum flavum. However, there have been no investigations to determine which biochemical factors cause these histologic changes. So we performed the current study to investigate the hypothesis that matrix metalloproteinases (MMPs), which possess the ability to cause extracellular matrix remodeling, may play a role as a mediator for this malady in the ligamentum flavum.
METHODS
The ligamentum flavum specimens were surgically obtained from thirty patients with spinal stenosis, as well as from 30 control patients with a disc herniation. The extents of ligamentum flavum elastin degradation and fibrosis were graded (grade 0-4) with performing hematoxylin-eosin staining and Masson's trichrome staining, respectively. The localization of MMP-2 (gelatinase), MMP-3 (stromelysin) and MMP-13 (collagenase) within the ligamentum flavum tissue was determined by immunohistochemistry. The expressions of the active forms of MMP-2, MMP-3 and MMP-13 were determined by western blot analysis, and the blots were quantified using an imaging densitometer. The histologic and biochemical results were compared between the two conditions.
RESULTS
Elastin degradation and fibrosis of the ligamentum flavum were significantly more severe in the spinal stenosis samples than that in the disc herniation samples (3.14 +/- 0.50 vs. 0.55 +/- 0.60, p < 0.001; 3.10 +/- 0.57 vs. 0.76 +/- 0.52, p < 0.001, respectively). The expressions of the active form of MMPs were identified in all the ligamentum flavums of the spinal stenosis and disc herniation patients. The expressions of active MMP-2 and MMP-13 were significantly higher in the spinal stenosis samples than that in the disc herniation samples (both p < 0.05). The expression of active MMP-3 was slightly higher in the spinal stenosis samples than that in the disc herniation samples, but the difference was not statistically significant (p = 0.131). MMP-2, -3, and -13 were positively stained on the ligamentum flavum fibroblasts.
CONCLUSIONS
The current results suggest that the increased expression of active MMPs by the ligamentum flavum fibroblasts might be related to the elastin degradation and fibrosis of the ligamentum flavum in the patients who suffer with lumbar spinal stenosis.

Keyword

Spinal stenosis; Ligamentum flavum; Elastin degradation; Fibrosis; Matrix metalloproteinases

MeSH Terms

Aged
Blotting, Western
Elastin/*metabolism
Extracellular Matrix/metabolism/pathology
Female
Fibrosis
Humans
Immunohistochemistry
Ligamentum Flavum/*metabolism/pathology
*Lumbar Vertebrae
Male
Matrix Metalloproteinase 13/metabolism
Matrix Metalloproteinase 2/metabolism
Matrix Metalloproteinase 3/metabolism
Matrix Metalloproteinases/*metabolism
Middle Aged
Spinal Stenosis/*metabolism/pathology

Figure

• 

  Fig. 1 Grading of the ligamentum flavum fibrosis by Masson's trichrome staining. In the grade 0 samples (A and C), all the area of the ligamentum flavum was stained a pink color, indicating a normal, non-fibrotic state. However, in the grade 4 samples (B and D), most of the area was stained a blue color, indicating that most of the area was fibrotic.

• 

  Fig. 2 Grading of the ligamentum flavum elastin degradation by hematoxylin-eosin staining. In the grade 0 samples (A and C), rich normal elastin fibers were organized in a strictly parallel order. However, in the grade 4 samples (B and D), the residual scanty elastin fivers were fragmented and disorderly.

• 

  Fig. 3 Immunohistochemical analysis demonstrates that MMP-2 (A), MMP-3 (B), and MMP-13 (C) were positively stained on the ligamentum flavum fibroblasts of the patients with spinal stenosis (original magnifi cation, × 400).

• 

  Fig. 4 Western blot analysis (A) demonstrated the expressions of active MMP-2, MMP-3 and MMP-13 in the ligamentum flavum specimens of the spinal stenosis and disc herniation patients, and the blots were quantified using an Imaging Densitometer. (B) Density is presented as the mean ± standard deviation (arbitrary units). The mean density of active MMP-2 and MMP-13 was statistically higher in the spinal stenosis samples than that in the disc herniation samples (both p < 0.05). Although the mean density of active MMP-3 was slightly higher in the spinal stenosis samples than that in the disc herniation samples, the difference was not statistically significant (p = 0.131). * p < 0.05.

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