| 1. |
- Finskas, Oscar, et al.
(author)
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New, clinically more relevant model for nerve root injury in the rat
- 2013
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In: Spine. - 0362-2436 .- 1528-1159. ; 38:20, s. 1744-8
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Journal article (peer-reviewed)abstract
- STUDY DESIGN: Exposure to nucleus pulposus and displacement of intraspinal nervous structures with assessment of spontaneous behavioral changes in rats. OBJECTIVE: To develop a controlled, experimental model for nerve root injury. SUMMARY OF BACKGROUND DATA: There are a number of experimental models presented for studies on radiculopathies. One frequently used model is based on exposure to nucleus pulposus and displacement of the dorsal root ganglion (DRG). However, it is clinically more common that the nerve roots are displaced/compressed than the DRG. In this study, we developed a model for displacement of the nerve root by modifying the DRG model. METHODS: After removing the left L3-L4 facet joint, the underlying disc was punctured, and the L4 nerve root was displaced laterally by an injection needle (n = 10). In sham experiments, the same procedure was performed without disc puncture and displacement (n = 10). In 10 rats, the left L4-L5 facet joint was removed. The underlying disc was punctured and the L4 DRG was displaced medially by an injection needle. Assessment of spontaneous behavioral changes was performed on days 1, 3, 7, 14, and 21, postsurgery. RESULTS: There was a clear increase in duration of the behavior "unloading of the paw" after displacement of the DRG that was most pronounced on day 1 and then gradually declined. There was a similar pattern for this behavior induced by nerve root displacement, although the duration was higher than that for the DRG displacement. No apparent trends in behavioral changes were observed for the other behaviors studied. CONCLUSION: Displacement of the nerve root induced more changes in the pain behavior than displacement of the DRG, but only for the behavior unloading of the paw. Because nerve root injury is more common than DRG injury, this model may be more clinically relevant than the DRG model. LEVEL OF EVIDENCE: N/A.
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| 2. |
- Geiss, A., et al.
(author)
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Autoimmune properties of nucleus pulposus: an experimental study in pigs
- 2007
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In: Spine. - 1528-1159. ; 32:2, s. 168-73
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Journal article (peer-reviewed)abstract
- STUDY DESIGN: Assessment of activated T and B cells in a subcutaneous chamber filled with autologous nucleus pulposus using flow cytometry and immunohistochemistry. OBJECTIVES: To examine if subcutaneously placed autologous nucleus pulposus may attract activated T and B cells in an animal model. SUMMARY OF BACKGROUND DATA: Nucleus pulposus has been suggested to trigger an autoimmune response if exposed to the immune system, for example, in association with disc herniation. T-cell activation represents a hallmark in the generation of an autoimmune response, subsequently leading to the differentiation of B cells, but a causal association between the exposure of nucleus pulposus to the systemic circulation and T and B cell activation is still lacking. METHODS: Autologous nucleus pulposus was harvested from the intervertebral disc of 9 pigs and placed subcutaneously in perforated titanium chambers. In order to control for the effect of the titanium chamber, an additional empty chamber was placed subcutaneously in each pig. After 7 days, the pigs were killed and the chambers were harvested. Flow cytometry and immunohistochemistry were used for analysis of T-helper cells (CD4+), cytotoxic T cells (CD8+), and B cells (Igkappa) in the chamber exudates and T cells (CD45RC) in the remaining blood clot tissue of the chamber. RESULTS: As compared with the empty chambers, the proportion of activated T cells (CD4+ and CD8+) was significantly higher in the exudate of the nucleus pulposus filled chamber. The proportion of activated B cells expressing immunoglobulin kappa (Igkappa) was also significantly elevated in the exudate of the nucleus pulposus chambers. The analysis of the remaining chamber tissue revealed a significantly higher amount of T cells (CD45RC) in the nucleus pulposus chambers than in the empty chambers. CONCLUSIONS: The present findings indicate that nucleus pulposus attracts activated T and B cells. However, since the cell population in the nucleus pulposus of young pigs may differ from that of adult humans, the obtained data may not be directly transferred to the human situation of a disc herniation. The observations in the present study may nevertheless explain some of the local tissue reactions occurring in association with disc herniation and nerve root involvement, thereby providing further insight into the pathophysiology of sciatica.
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| 3. |
- Igarashi, A., et al.
(author)
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Inflammatory cytokines released from the facet joint tissue in degenerative lumbar spinal disorders
- 2004
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In: Spine. - 1528-1159. ; 29:19, s. 2091-5
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Journal article (peer-reviewed)abstract
- STUDY DESIGN: A prospective study of surgical cases of degenerative lumbar spinal disorders demonstrated inflammatory cytokines in the facet joint tissue. OBJECTIVE: To quantify various inflammatory cytokines released from the facet joint tissue in surgical cases of degenerative lumbar spinal disorders. SUMMARY OF BACKGROUND DATA: In degenerative lumbar spinal disorders, pain is often caused by osteoarthritis of a facet joint. There are inflammatory mediators such as prostaglandins and leukotrienes in facet joint tissue in lumbar spinal degeneration. However, no reports have studied if there are also inflammatory cytokines in the facet joint, which generated arthropathic changes in degenerative lumbar spinal disorder and if pain is caused by chemical factors associated with inflammation such as inflammatory cytokines. METHODS: Forty patients with degenerative lumbar disorders who had undergone operative treatment were included in this study. Fifty-five joint cartilages and 67 synovia were harvested from the lumbar facet joints in responsible intervertebral levels of patients. There were 24 male and 16 female subjects with average ages of 50 and 67 years, respectively, in 11 cases of lumbar disc herniation and 29 cases of lumbar spinal canal stenosis. Using ELISA and CLEIA methods, joint cartilage and synovial tissues were harvested during surgery from the facet joint at the responsible upper levels to measure IL-1beta, TNF-alpha, and IL-6 in individual tissues. RESULTS: IL-1beta was detected in joint cartilage and synovium in both groups and its positive reaction rate was higher in LSCS than in LDH. There was no difference in IL-1beta concentration in cartilage tissue between the two groups. There was TNF-alpha in the synovium of LSCS. IL-6 was high in joint cartilage and synovium in both groups. The concentration was significantly higher in LSCS than in LDH. CONCLUSIONS: There are inflammatory cytokines in facet joint tissue at high levels in degenerative lumbar spinal disorders. Inflammatory cytokines have a higher concentration rate in lumbar spinal canal stenosis than in lumbar disc herniation. This finding suggests that inflammatory cytokines in degenerated facet joints may have some relation to the cause of pain in degenerative lumbar disorders.
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| 4. |
- Larsson, Karin, 1955, et al.
(author)
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Disc related cytokines inhibit axonal outgrowth from dorsal root ganglion cells in vitro
- 2005
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In: Spine. - 1528-1159. ; 30:6, s. 621-4
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Journal article (peer-reviewed)abstract
- STUDY DESIGN: Application of nucleus pulposus and disc related cytokines in vitro on cultured dorsal root ganglion (DRG) cells. OBJECTIVES: To study if tumor necrosis factor (TNF) and interleukin-1beta (IL-1beta) may induce similar inhibition of axonal outgrowth from cultured DRG cells as application of nucleus pulposus and to compare a new assessment method to previous data. SUMMARY OF BACKGROUND DATA: Pro-inflammatory cytokines related to the intervertebral disc have been suggested to affect adversely neurons following local application, with implications for the nucleus pulposus-induced nerve injury seen in various studies. Nucleus pulposus is known to inhibit axonal outgrowth from cultured DRG cells, thereby indicating a neurotoxic potential. The mechanisms were not understood, but it was suspected that the effect was mediated by pro-inflammatory cytokines produced by the nucleus pulposus. METHODS: DRG were harvested from newborn rats and put in culture. The axonal outgrowth was determined 24 hours after starting the culture. Twenty-four hours after exposing the cultured cells to nucleus pulposus, frozen nucleus pulposus, TNF, or IL-1beta, the axonal outgrowth was reassessed, and the outgrowth during the exposure time was calculated. RESULTS: Nucleus pulposus clearly reduced the axonal outgrowth. Also, application of TNF and IL-1beta reduced the outgrowth but not as pronounced as the nucleus pulposus. Frozen nucleus pulposus had no effects on the outgrowth. Overall, the data were similar regarding frozen and nonfrozen nucleus pulposus compared to a previous study. CONCLUSIONS: It was evident that the 2 studied cytokines inhibited the outgrowth of axons from cultured DRG cells, thus suggesting a neurotoxic potential. However, the inhibition was not as pronounced as for nucleus pulposus. These data may increase our understanding for cytokine induced nerve injury, with implications for future treatment strategies for such conditions.
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| 5. |
- Murata, Y., et al.
(author)
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Changes in pain behavior and histologic changes caused by application of tumor necrosis factor-alpha to the dorsal root ganglion in rats
- 2006
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In: Spine. - 1528-1159. ; 31:5, s. 530-5
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Journal article (peer-reviewed)abstract
- STUDY DESIGN: Histologic changes in the dorsal root ganglion (DRG) and the nociceptive stimulation thresholds were studied in rats. OBJECTIVE: To examine the effects of tumor necrosis factor-alpha (TNF) with special reference to pain behavior and histology of the DRG. SUMMARY OF BACKGROUND DATA: Recently, it was reported that local application of nucleus pulposus induces a characteristic tissue reaction at the surface of the DRG. However, to our knowledge, there have been no previous reports about the relationship between the histologic changes and pain behavior caused by cytokines. METHODS: Recombinant TNF was applied to the L4 DRG. Mechanical and thermal nociceptive thresholds were tested. The L4 DRG was sectioned and observed by light microscopy. RESULTS: After the application of 5 ng/microL TNF, significant differences were observed in mechanical and thermal stimulation thresholds. At the site of application of TNF, a characteristic a semilunar-shaped enlargement was observed. The average width of the part was significantly larger in the 5 ng/microL TNF application, as compared to the 0.5-ng/microL TNF application. CONCLUSIONS: The higher concentration of TNF used induced allodynia and hyperalgesia responses. Because the region showing the histologic changes was significantly larger after application of the higher concentration of TNF, the reaction of the DRG may be related to pain.
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| 6. |
- Murata, Y., et al.
(author)
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Distribution and appearance of tumor necrosis factor-alpha in the dorsal root ganglion exposed to experimental disc herniation in rats
- 2004
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In: Spine. - 1528-1159. ; 29:20, s. 2235-41
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Journal article (peer-reviewed)abstract
- STUDY DESIGN: Distribution and appearance of tumor necrosis factor-alpha (TNF-alpha) in the dorsal root ganglion (DRG) exposed to experimental disc herniation were investigated using an immunohistochemical method in rats. OBJECTIVES: To study the distribution and appearance of TNF-alpha in the DRG following experimental disc herniation in rats. SUMMARY OF BACKGROUND DATA: Nucleus pulposus in the epidural space induces spinal nerve root injury not only by mechanical but also chemical mechanisms. Cytokines may play a key role in the chemical damage. There is, however, no report on the distribution and appearance of TNF-alpha in the DRG exposed to nucleus pulposus. METHODS: Nucleus pulposus from the discs was smeared on the glass slides and processed for immunohistochemistry by the avidin-biotinylated peroxidase complex technique using rabbit antisera to TNF-alpha in rats. A herniation of the nucleus pulposus was made by incision of the L4-L5 disc in rats. The L4 and L5 DRGs were resected 1, 3, 7, 14, and 21 days after surgery. The specimens were processed for immunohistochemistry using rabbit antisera to TNF-alpha. The TNF-alpha-positive cells were observed and counted using light microscopy. Distribution of the TNF-alpha products was compared on each day after surgery. RESULTS: A positive staining was seen in the cell bodies and in the matrix between the cells in the smeared nucleus pulposus. In the L4 DRG sections, the number of positive cells was significantly higher in the disc incision group than in the sham group at 1, 3, 7, and 14 days after surgery (P < 0.05). The positive cells showed a decrease in number day by day after surgery. On the contrary, in the L5 DRG, only a few positive cells were observed in the disc incision group after surgery. There was no statistically significant difference between disc incision and the sham groups at each day after surgery for the L5 DRGs. CONCLUSIONS: The immunoreactivity of TNF-alpha in the DRG directly exposed to nucleus pulposus increases during 2 weeks. A collapse of the positive cells was seen in the DRG directly exposed to the nucleus pulposus.
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| 7. |
- Murata, Y., et al.
(author)
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Incision of the intervertebral disc induces disintegration and increases permeability of the dorsal root ganglion capsule
- 2005
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In: Spine. - 1528-1159. ; 30:15, s. 1712-6
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Journal article (peer-reviewed)abstract
- STUDY DESIGN: The origin and the barrier properties of the characteristic reaction at the surface of the dorsal root ganglion (DRG) exposed to the nucleus pulposus was studied using Alcian-Blue staining, van Gieson staining, and the application of Evans Blue Albumin (EBA) complex in rats. OBJECTIVE: To study the origin and the barrier properties of the capsule, including the characteristic reaction, at the surface of the DRG exposed to the nucleus pulposus. SUMMARY OF BACKGROUND DATA: Local application of nucleus pulposus may induce a characteristic reaction at the surface of the DRG. This reaction histologically resembles an acute inflammatory reaction. However, it is not evident if this is a swelling of the DRG capsule, if it is located between the capsule and neurons of the DRG, or if it is only an attached nucleus pulposus. METHODS: Nucleus pulposus from the discs was obtained. The nucleus pulposus was smeared on glass slides. Alcian-Blue with hematoxylin and eosin staining was performed for each smear. Herniation of the nucleus pulposus was made in the L4-L5 disc in rats. The L4 DRGs were resected 3, 24, and 72 hours after surgery, and sectioned. The sections were processed for Alcian-Blue staining, van Gieson staining, and EBA complex infiltration. The sections were observed using light or fluorescent microscopy. RESULTS: Smear of nucleus pulposus was stained bright blue indicating mucins. A characteristic reaction, "inflammatory crescent," was confirmed at the surface of the DRG exposed to the nucleus pulposus. No mucins were observed in the crescent using Alcian-Blue. The results of van Gieson staining showed that the reaction started both inside and outside the elastic fiber layer, the DRG capsule, within 3 hours. The EBA complex was capable of infiltrating into the DRG capsule 24 hours after disc incision. CONCLUSIONS: The disintegrated capsule showed an increased permeability even for a large molecule as albumin, which indicates a possible entrance route for various substances induced by locally applied nucleus pulposus.
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| 8. |
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| 9. |
- Murata, Y., et al.
(author)
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Nucleus pulposus-induced apoptosis in dorsal root ganglion following experimental disc herniation in rats
- 2006
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In: Spine. - 1528-1159. ; 31:4, s. 382-90
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Journal article (peer-reviewed)abstract
- STUDY DESIGN: The mechanisms behind the formation of a characteristic tissue reaction at the surface of the dorsal root ganglion (DRG) exposed to nucleus pulposus was studied with special reference to apoptosis using electron microscopy and immunohistochemistry in rats. OBJECTIVES: To study the mechanism of the characteristic tissue reaction at the surface of the DRG exposed to nucleus pulposus. SUMMARY OF BACKGROUND DATA: Recently, it was observed that local application of nucleus pulposus may induce a characteristic tissue reaction at the surface of the DRG. This change occurred as early as 1 day after the application of nucleus pulposus. METHODS.: Herniation of nucleus pulposus was created in the L4-L5 disc in rats. The L4 DRG were resected 3 and 24 hours after surgery. The sections of the specimens were observed using light and electron microscopy. The sections were processed for immunohistochemistry using antibodies to single-stranded DNA (ssDNA), caspase 3, and tumor necrosis factor alpha (TNF). RESULTS: There were typical changes of the cell nuclei observed by light and electron microscopy, especially those of the small-sized cells, in the DRG 24 hours after application of nucleus pulposus, indicating the presence of apoptosis. The presence of ssDNA, caspase 3, and TNF further enhanced the impression that there was apoptosis in the DRG. Nucleus pulposus induced apoptosis in the DRG at the site of application within as little as 24 hours. CONCLUSIONS: Nucleus pulposus herniated from the disc induced apoptosis in at the surface of the DRG exposed to nucleus pulpous as early as 24 hours after exposure.
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| 10. |
- Murata, Y., et al.
(author)
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Selective inhibition of tumor necrosis factor-alpha prevents nucleus pulposus-induced histologic changes in the dorsal root ganglion
- 2004
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In: Spine. - 1528-1159. ; 29:22, s. 2477-84
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Journal article (peer-reviewed)abstract
- STUDY DESIGN: The possibility to prevent nucleus pulposus-induced structural changes of the dorsal root ganglion (DRG) by selective tumor necrosis factor-alpha (TNF-alpha) inhibition was assessed in an experimental model in the rat spine. OBJECTIVES: To evaluate the role of TNF-alpha in the mediation of nucleus pulposus-induced structural changes by using selective inhibition and to confirm the effect of TNF-alpha inhibitor at the point of histologic findings. SUMMARY OF BACKGROUND DATA: TNF-alpha is known to be released from the nucleus pulposus, and has been suggested to play a key role in chemical damage of the adjacent nerve tissue. The TNF-alpha inhibitor prevents the reduction of nerve conduction velocity and may limit the nerve fiber injury, intracapillary thrombus formation, and intraneural edema formation caused by nucleus pulposus. However, there is no report on the effect of the inhibitor regarding histologic findings and the appearance of the TNF-alpha in the DRG exposed to nucleus pulposus. METHODS: 1) Rats were treated with an intraperitoneal injection of infliximab. Nucleus pulposus from the disc was obtained 1, 3, 7, 14, and 21 days after the injection. The TNF-alpha-positive cells were observed using immunohistochemistry. 2) Disc herniation of the nucleus pulposus was made on the L4-L5 disc in rats. Two groups were treated with selective TNF-alpha inhibitor 1 day before or 3 hours after surgery. The other group received no TNF-alpha inhibitor. The L4 DRG was resected 1, 3, 7, 14, and 21 days after surgery. The specimens were processed for hematoxylin and eosin staining and immunohistochemistry using rabbit antisera to TNF-alpha. The histologic findings and TNF-alpha-positive cells were observed by light microscopy. RESULTS: 1) While positively stained immunoreactive TNF-alpha appeared between 7 and 21 days, no immunoreactive TNF-alpha was observed 1 and 3 days after injection in the nucleus pulposus. 2) The histologic changes of the DRG caused by nucleus pulposus were smaller in the infliximab treatment group than those in the nontreatment group. The number of immunoreactive TNF-alpha cells was high 1 and 3 days after surgery in the DRGs of disc herniation rats that were treated without an injection of the inhibitor, low on day 7 and 14, and very low on day 21 after surgery. No immunoreactive TNF-alpha was observed in the DRGs of the TNF-alpha inhibitor treatment groups on day 1, 3, and 21 after surgery. Weakly stained cells were sometimes observed in rats at day 7 and 14 after surgery. CONCLUSIONS: Infliximab may prevent the histologic damage induced by nucleus pulposus. When rats were given a single intraperitoneal injection of infliximab at the beginning of disc herniation, the histologic damage seemed to be reduced in comparison with the nontreated rats.
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