Herniated Lumbar Disc Tissue Research Articles

Analysis and prevalence of inflammatory cells in subtypes of lumbar disc herniations under cyclooxygenase-2 inhibitor therapy

Objective: The aim of the present study was to analyse inflammatory cells in lumbar disc tissue under cyclooxygenase-2 (COX-2) inhibitor therapy, to detect their prevalence in different subtypes of lumbar disc herniations and to assess the influence of inflammatory reactions in herniated disc tissue on postoperative outcome.Methods: In this prospective study, intervertebral disc specimens were obtained from 50 patients. All the patients were given COX-2 inhibitor therapy (Rofecoxib) 25 mg/day for 10 days before surgery. The herniated disc specimens were routinely fixed in a 10% buffered formaline solution and paraffin-embedded; 5 &mum thin sections were stained with monoclonal antibodies CD-68 for macrophages, CD-45 RO for T cells and CD-20 for B cells. The specimens were microscopically examined and classified by two independent examiners in a blinded manner.Results: CD-68 macrophages were evident in herniated lumbar disc tissue in 40% of the cases, but abundant inflammation was observed in only 18% of the cases. Macrophages were significantly common in sequesters with a prevalence of 80%, and no macrophages were observed in the protrusions. We obtained the best outcome scores in the patients with a sequestrated lumbar disc herniation.Conclusion: Although most of the specimens from all the patients showed no significant inflammatory reaction, maybe due to COX-2 inhibitor therapy, they all needed surgery with no relief of the symptoms despite conservative therapy. Nerve root compression still seems to be the leading pathomechanism.

Mitochondrial Involvement in Fas-Mediated Apoptosis of Human Lumbar Disc Cells

Two main pathways of Fas-mediated apoptosis have been identified: the Type-I (death-inducing signaling complex) pathway and the Type-II (mitochondrial) pathway. While apoptotic cell death has been implicated in lumbar degenerative disc disease, we are not aware of any studies in which surgically removed discs from live humans have been examined to determine which of the two pathways is involved in the apoptosis of disc cells. As an initial step in the development of therapies to inhibit inappropriate or premature apoptosis of disc cells, our objective was to determine which pathway is involved. We examined thirty-two samples of herniated lumbar disc tissue with use of immunohistochemical staining and Western blot analysis to determine the presence of several proteins, including caspase-8 (associated with the Type-I pathway); BID (BH3 interacting domain death agonist), cytochrome-c, and caspase-9 (associated with the Type-II pathway); and caspase-3 (an executioner of apoptosis). The TUNEL (terminal deoxynucleotidyl transferase [TDT]-mediated dUTP nick end labeling) assay was performed to confirm the occurrence of apoptosis of the disc cells. The proteins associated with the Type-II pathway (BID, cytochrome-c, and caspase-9) stained positively in all samples. Although the protein associated with the Type-I pathway (caspase-8) was not detected on immunohistochemical analysis, a small amount of caspase-8 was detected on Western blot analysis. However, the expression of Type-II proteins was still higher than the expression of caspase-8 on Western blot analysis. The expression of caspase-3 was identified in all samples with immunohistochemical and Western blot analysis. TUNEL-positive disc cells were identified in all samples. The results of the present study suggest that human disc cells are Type-II cells, which undergo apoptotic cell death through mitochondrial involvement.

Occurrence and Regional Distribution of Apoptosis in Scoliotic Discs

Seventy surgically obtained intervertebral discs from 9 patients with idiopathic and 7 patients with neuromuscular scoliosis were analyzed for the regional distribution of apoptosis. To investigate the role of apoptotic mechanisms in scoliotic discs. The reasons for the development of scoliosis are complex and yet not completely understood. In herniated lumbar disc tissue, increased apoptosis and a high expression of Fas and Fas ligand and caspase-3 activity have already been reported, suggesting a pivotal role of apoptotic mechanisms in intervertebral disc degeneration. In scoliotic discs, cell death was correlated with disc deformity and changes in nutrient supply and metabolic levels. The role of apoptosis in scoliotic discs, however, is still unclear. Apoptosis was determined by terminal deoxyribonucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay and poly(ADP-ribose) polymerase (PARP) p85 immunohistochemistry. Expression of Fas and Fas-ligand was analyzed by immunohistochemistry and reverse transcription polymerase chain reaction analysis. In all samples analyzed, apoptotic cells could be detected in the nucleus, anulus, and endplate. The number of apoptotic cells was significantly higher in the nucleus compared to the other areas and in the apex versus the nonapex discs. There was no difference between the discs of idiopathic and neuromuscular scoliosis and between the 2 age groups studied (10-17 years and 17-48 years, respectively). A strong expression of Fas and Fas-ligand could be detected in all samples. Increased numbers of apoptotic cells in the nucleus of scoliotic discs and the apex disc suppose a pivotal role of programmed cell death for the progression of this common disorder. The simultaneous increase of Fas and Fas-ligand expression in areas with increased cell death point to an activation of the apoptotic process via the Fas/Fas-L system.

The pattern of interleukin-12 and T-helper types 1 and 2 cytokine expression in herniated lumbar disc tissue.

A study was conducted to investigate the expression of cytokines related to the immune reaction in herniated lumbar disc tissues. To investigate the immunologic status of lumbar disc tissue and the type of immune reaction that occurs in response to lumbar disc herniation. It has been proposed that herniated lumbar disc tissue causes an immune reaction. Various inflammatory cells, proinflammatory cytokines, antibodies, and immunoglobulins have been identified in and around herniated lumbar disc tissue. Recently, it has been reported that lumbar disc tissue may be another potential immune-privileged site in the human body. This study included 40 herniated lumbar disc tissues: 20 contained and 20 noncontained discs. The concentrations of interleukin-12, T-helper Type 1, interferon-gamma (IFN-gamma), and Th2 (IL-4) cytokines were determined by enzyme-linked immunosorbent assay, respectively. The results were compared between the two groups. The concentrations of interleukin-12 and interferon-gamma were higher in the noncontained discs than in the contained discs: 28.3 +/- 10.7 pg/mL vs 9.2 +/- 4.2 pg/mL (P = 0.001) and 4.7 +/- 5.0 pg/mL vs 2.3 +/- 3.8 pg/mL (P = 0.029), respectively. On the contrary, the concentration of IL-4 was higher in the contained discs than in the noncontained discs: 24.3 +/- 20.1 pg/mL vs 1.9 +/- 4.5 pg/mL; P= 0.001. The degrees of interleukin-12 and interferon-gamma expression were negatively correlated with that of IL-4 (n = 40): correlation coefficient, -0.671 (P = 0.001) and correlation coefficient, -0.344 (P = 0.03), respectively. The findings suggest that preferential expression of Th2 cytokines by disc cells, as shown in contained discs, is another factor contributing to the immune privileged status of lumbar disc tissue. The exposure of lumbar disc tissue to the epidural space may increase the concentration of interleukin-12 in herniated lumbar disc tissue, changing the pattern of T-helper Types 1 and 2 cytokine expression.

Expression of Fas ligand and apoptosis of disc cells in herniated lumbar disc tissue.

An examination of surgically obtained herniated lumbar disc tissues performed by using immunohistochemical staining and the DNA nick end labeling method. To investigate the cell type that expresses Fas ligand (FasL) and any evidence of apoptosis of the disc cells in herniated lumbar disc tissues. The Fas/FasL system is involved in delivering a death signal that rapidly commits the cells to apoptosis. In the authors' previous study, the expression of Fas on disc cells was identified in herniated lumbar disc tissue. Twenty-three herniated lumbar disc tissues (contained disc, n = 9; noncontained disc, n = 14) were examined to investigate the cell type that expresses FasL and any evidence of apoptosis of the disc cells by using immunohistochemical staining and the DNA nick end labeling method, respectively. The percentage of FasL-positive disc cells was calculated and compared with clinical and radiologic data. FasL was expressed in the cytoplasm of the disc cells, and nuclear DNA fragmentation in a few disc cells was identified. A higher degree of FasL expression in disc cells was found in noncontained discs than in contained discs (P < 0.05). The percentage of FasL-positive disc cells significantly increased with the patient's age (P < 0.05), but not with the degree of disc degeneration (P > 0.05). The current results indicate that disc cells, after herniation, undergo apoptotic cell death via autocrine or paracrine FasL mechanisms by the disc cells themselves.

Expression of Fas Receptor on Disc Cells in Herniated Lumbar Disc Tissue

The expression of Fas receptor, an apoptosis-related protein, on disc cells was examined in surgically obtained disc specimens. To assess the fate of disc cells in herniated disc tissue and the difference in the degree of expression of the Fas receptor between contained and noncontained discs. Little is known about the fate of disc cells after herniation. Twenty-three herniated lumbar disc specimens were classified into contained discs (protrusion or subligamentous extrusion; n = 9) and noncontained discs (transligamentous extrusion or sequestration; n = 14). All specimens were stained using the avidin-biotin-peroxidase complex method. The percentage of disc cells positive for Fas receptor was calculated and compared with clinical and radiologic data. There was a significant difference in the percentage of Fas-positive disc cells between the contained and noncontained discs (8.44 vs.- 14.29;P = 0.044). The percentage of Fas-positive disc cells correlated significantly with the patient's age (r = 0.455, P = 0.029), but not with the degree of disc degeneration on magnetic resonance imaging (r = 0.252, P = 0.214). This is the first study to identify the expression of Fas receptor on disc cells in herniated disc tissue. The results show that the disc cells after herniation may undergo Fas-mediated apoptosis and that the degree of expression of Fas receptor differs depending on the type of herniation.

Influence of macrophage infiltration of herniated lumbar disc tissue on outcome after lumbar disc surgery.

An immunohistochemical examination of the presence of inflammatory cells in routinely processed resection specimens of the lumbar disc, and a comparison of the histologic results with clinical data collected prospectively before and after surgery. To assess the influence of inflammatory reactions in herniated lumbar disc specimens on the outcome after lumbar disc surgery. Histologic and biochemical studies on herniated lumbar disc tissue led to the notion of inflammation-induced sciatic pain. At this writing, no investigations have sought to discover how outcome after lumbar disc surgery is influenced by histologically described inflammation. Disc specimens from 79 patients who underwent surgery for lumbar disc herniation were studied immunohistologically with regard to the presence of inflammatory reactions. Of these, 92% were followed up approximately 7 months after surgery. The histologic results were compared with the outcome at follow-up evaluation. A statistically significant correlation was found between the histologically proven inflammation and the outcome, as shown by the pain grading scale. The results from this study seem to support the theory of a foreign body reaction to the herniated material. This reaction may result in inflammation-induced sciatic pain.

The role of mast cells in disc herniation inflammation.

A study of herniated lumbar disc tissue samples and control disc material to determine the presence of mast cells in disc herniations. To analyze whether mast cells have any involvement in disc herniation pathophysiology and lumbar pain, because mast cells may have an important role in acute and chronic inflammatory responses. Studies of inflammatory cells, biochemical mediators of inflammation, and tissue degrading enzymes have suggested that these factors may be involved--and perhaps play an important role--in the pathophysiology of lumbar pain and radiculopathy. Mast cells are known to play an important role in acute and chronic inflammatory responses. It was therefore of interest to clarify their possible role in intervertebral disc herniation inflammation. Fifty herniated lumbar discs from 50 patients who had undergone disc surgery and three normal control discs were obtained. Sections from every disc then were examined histologically and immunocytochemically for mast cells by using monoclonal antibodies to either of two types of specific proteases of mast cells, tryptase and chymase. By none of the methods could any mast cells be observed in any of the control disc samples. With toluidine blue staining, mast cells were observed in 9 of 50 (18%) of discs. Mast cells immunoreactive to either tryptase or chymase were observed in 10 of 50 disc samples (20%) and immunoreactive for tryptase and chymase simultaneously in 4 of 50 disc samples (8%). However, the majority of the samples studied (80%) demonstrated immunoreactivity to neither tryptase nor chymase. Among the samples studied were five disc protrusions that totally lacked mast cells. A minority of disc herniations exhibited mast cells, as verified by toluidine blue staining and immunocytochemistry. The results may suggest a role of mast cells in intervertebral disc herniation inflammation, but only in a subset of these cases. Massive infiltration by mast cells never was observed.

Point of View: Is There a Clinical Correlate to the Histologic Evidence of Inflammation in Herniated Lumbar Disc Tissue?

Point of View: Is There a Clinical Correlate to the Histologic Evidence of Inflammation in Herniated Lumbar Disc Tissue?

Immunohistochemical demonstration of sensory and autonomic nerve terminals in herniated lumbar disc tissue.

Thirty-five lumbar disc herniations removed at surgery were studied by indirect immunocytochemistry. To localize immunohistochemically both sensory and autonomic nerve terminals in disc herniations. Using various more or less specific histologic and histochemical methods, investigators have reported the presence of free nerve terminals in disc tissue. However, very few studies have, to date, convincingly demonstrated nerve terminals in disc tissue that morphologically resemble the tiny nerve terminals of sensory and autonomic nerve fibers. Amplification of the peroxidase reaction product in avidin-biotin-peroxidase complex immunostaining by the glucose oxidase-diaminobenzidine-nickel sulfate method was used to visualize small punctate nerve terminals at high magnification. Thin frozen sections from disc herniation tissue prefixed in Zamboni fixative were incubated with antibodies to synaptophysin to visualize nerve terminals in general, and with antibodies to substance P and C-flanking peptide of neuropeptide Y to further characterize nerve terminals as either sensory or sympathetic. Nerve terminals could be demonstrated in 29 (83%) of the 35 disc herniations. They were observed with the synaptophysin antibody in 17 of 35 (49%) disc herniations, with substance P in 16 of 35 (46%) disc herniations, and with C-flanking peptide of neuropeptide Y in 13 of 35 (37%) disc herniations. Morphologically, the nerve terminals were seen as tiny immunoreactive dots. Some of the nerve terminals were observed close to disc cells, possibly suggesting direct interaction. Small nerve terminals in disc herniations, both sensory substance P endings and sympathetic C-flanking peptide of neuropeptide Y endings, could be involved in mechanisms of discogenic pain, disc tissue neurogenic inflammation, tissue repair processes after injury, and control of local blood circulation in the newly formed blood vessels. Disc cells may be directly affected by the neuropeptides released from nearby nerve terminals.