Phenotypic Characterization of Ligamentum Flavum Cells from Patients with Ossification of Ligamentum Flavum

PURPOSE: The objective of this study was to determine the phenotypic characterization of ligamentum flavum cells from patients with ossification of the ligamentum flavum (OLF). MATERIALS AND METHODS: Ligamentum flavum tissues were harvested from OLF and non-OLF patients during surgery. OLF and non-OLF cells were isolated from explant cultures. Cultured cells were analyzed using immunofluorescence staining and reverse transcription-polymerase chain reaction. RESULTS: OLF cells exhibited various appearances compared with the typical fibroblast-like morphology of non-OLF cells. Expressions of collagen type I and collagen type III were observed in OLF and non-OLF cells. OLF cells uniquely expressed osteocalcin, which is a marker for osteoblasts, and collagen type II which is a marker for chondrocytes, whereas they were negative in non-OLF cells. CONCLUSION: These findings indicate that OLF cells have phenotypic characterization of osteoblasts and chondrocytes which could play a role in the pathophysiology of OLF.

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

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.