Nucleus Pulposus Group Research Articles

Involvement between social defeat stress and pain-related behavior in a rat lumbar disk herniation model.

Psychological and social factors are involved in the disability and chronicity of pain. Our study aim was to investigate whether social defeat stress (SDS) as a psychophysical stress affected mechanical withdrawal thresholds in the lumbar disk herniation (LDH) rat model. Changes in microglia and astrocytes, which play important roles in neuropathic pain states, were also investigated. For the LDH model, nucleus pulposus (NP) was applied to the L5 dorsal root ganglion (DRG) in adult female Sprague-Dawley rats. SDS was performed 15min daily for 8days. Mechanical withdrawal thresholds were measured, and immunoreactive cells of glial fibrillary acidic protein (GFAP) and ionized calcium-binding adaptor molecule-1 (Iba-1), which were used as markers of microglia, satellite glial cells, and astrocytes, were assessed in the DRG, spinal cord (SC), and ventrolateral periaqueductal gray matter (VLPAG). Mechanical withdrawal thresholds decreased in the NP group for 21days and for 35days in the NP + SDS group. Expression of GFAP and Iba-1 in the DRG and SC increased up to day 21 in the NP and NP + SDS groups. In the sham + SDS and NP + SDS groups, expression of GFAP in the VLPAG decreased until day 35. SDS prolongs mechanical allodynia induced by NP. Changes of GFAP expression in the VLPAG were associated with mechanical allodynia of the NP + SDS group during the late phase. These results suggest that psychological chronic stress might delay recovery from mechanical allodynia induced by the LDH model.

Recurrent disc herniation following percutaneous endoscopic lumbar discectomy preferentially occurs when Modic changes are present

ObjectiveTo investigate the relationship between Modic changes (MCs) and recurrent lumbar disc herniation (rLDH) and that between the herniated disc component and rLDH following percutaneous endoscopic lumbar discectomy (PELD).MethodsWe included 102 (65 males, 37 females, aged 20–66 years) inpatients who underwent PELD from August 2013 to August 2016. All patients underwent CT and MRI preoperatively. The presence and type of Modic changes were assessed. During surgery, the herniated disc component of each patient was classified into two groups: nucleus pulposus group and hyaline cartilage group. The association of herniated disc component with Modic changes was investigated. The incidence of rLDH was assessed based on a more than 2-year follow-up.ResultsIn total, 11 patients were lost to follow-up; the other 91 were followed up during 24–60 months. Of the 91 patients, 99 discs underwent PELD; 28/99 (28.3%) had MCs. Type I and II MCs were seen in 9 (9.1%) and 19 (19.2%), respectively; no type III MCs were found. Among 28 endplates with MCs, according to the herniated disc component, 18/28 (64.3%) showed evidence of hyaline cartilage in the intraoperative specimens, including 6/9 and 12/19 endplates with type I and II MCs, respectively. Among 71 endplates without MCs, 14/71 (19.7%) showed evidence of hyaline cartilage in the intraoperative specimens. Hyaline cartilage was more common in patients with MCs (P < 0.05). We found 2 cases of rLDH in the non-MC group (n = 71); 6 cases of rLDH were found in the MC group (n = 28), including 2 and 4 cases for types I and II, respectively. There was no significant difference between types I and II (P > 0.05). rLDH was more common in patients with MCs (P < 0.05). We found 5 rLDH cases in the hyaline cartilage group (n = 32); 3 rLDH cases were found in the nucleus pulposus group (n = 67). rLDH was more common in the hyaline cartilage group (P < 0.05).ConclusionsrLDH following PELD preferentially occurs when MCs or herniated cartilage are present.

Association Between Neurotrophic Factor Expression and Pain-Related Behavior Induced by Nucleus Pulposus Applied to Rat Nerve Root.

An experimental animal study. To investigate the relationship between pain-related behavior and the expression of neurotrophic factors in the dorsal root ganglion (DRG) and spinal cord (SC) using a nucleus pulposus (NP) rat model. Neurotrophic factors are released from activated glial cells and are associated with pain-related behavior. Nerve growth factor (NGF) is a neurotrophic factor that is induced by inflammation. Rats were divided into an NP group (n = 94) and a sham-operated group (n = 46). NP harvested from the tail was applied to the left L5 DRG. Rats in the NP group were then divided into five subgroups: one non-treatment and four treatment groups. In the treatment groups, a dose of anti-NGF antibody or phosphate-buffered saline was administered into the DRG. Behavioral testing was performed to investigate the mechanical withdrawal threshold of the left hind paw for all groups. Immunohistochemical localization of NGF, phosphorylated p38 (p38), and brain-derived neurotrophic factor (BDNF) in the DRGs and SCs was performed, and the numbers of immunoreactive (IR) cells were counted. The withdrawal threshold in the nontreatment NP group was significantly decreased for 35 days, and that of the middle- and high-dose treatment rats was significantly higher than the phosphate-buffered saline group values. In the DRG, NGF-IR, p38-IR, and BDNF-IR cells were increased for days 21. In the SC, BDNF-IR, and p38-IR cells were increased from days 7 to 21. In the DRG, NGF expression increased, mechanical thresholds were reduced, and p38 and BDNF expression was increased in the NP group. p38 and BDNF expression was increased in SC neurons during the same period. Inhibition of NGF may be a potential treatment for neuropathic pain due to lumbar disc herniation. 5.

Effects of Degenerative Autologous Nucleus Pulposus on the Lumbar Sympathetic Trunk: Sympathetic Inflammation in Rats

Objective We have reported the presence of discogenic visceral pain secondary to anterior herniation of the lumbar disc. The aim of this study was to observe the inflammatory response of the sympathetic trunk to an autologous degenerative nucleus pulposus (NP) injection under fluoroscopy. Methods A total of 72 rats were used. In 24 rats, the autologous NP suspension was injected into the right sympathetic trunk. Next, food intake and body weight of each rat were monitored for 14 days. Fourteen days after the injection, the right lumbar sympathetic trunk was harvested for histological assessment, and protein levels of IL-1 β, IL-6, and TNF-α were quantified via ELISA. Results In the NP-treated group, endoneural hyperemia and intensive infiltration of inflammatory cells could be identified in sections of the sympathetic trunk by the hematoxylin and eosin (H&amp;E) stain. Meanwhile, elevated concentrations of IL-1 β, IL-6, and TNF-α were determined in the sympathetic trunk of the NP group as compared to that of the naïve and control groups, which indicated the development of an inflammatory response. Furthermore, food intake and body weight of rats in the NP group decreased significantly. Conclusions The results indicated that an inflammatory response in the sympathetic trunk can be caused by anterior herniation of the lumbar disc, which can generate a sympathetic inflammatory pain response.

Measurement of mechanical withdrawal thresholds and gait analysis using the CatWalk method in a nucleus pulposus-applied rodent model

BackgroundThere are some previous reports of gait analysis using a rodent pain model. Applying the CatWalk method, objective measurements of pain-related behavior could be evaluated, but this method has not been investigated using the nucleus pulposus (NP) applied model, which was developed as a model of lumber disc herniation. We aimed to measure mechanical withdrawal thresholds and analyze gait patterns using the CatWalk method for the evaluation of the pain-related behavior caused by NP application.MethodsTwenty-four nine-week-old female Sprague-Dawley rats were randomly divided into two experimental groups, the NP group (n = 12), in which autologous NP from the tail was applied to the left L5 dorsal root ganglion, and the sham-operated group (n = 12). Measurements of mechanical withdrawal thresholds were performed using von Frey filaments touching the left footpads, and gait analysis was performed using the CatWalk method. These experiments were conducted 1 day before surgery and 7, 14, 21, and 28 days after surgery. Data were statistically analyzed using the Wilcoxon rank-sum test.ResultsThe NP group showed significantly lower withdrawal thresholds than the sham group at days 14 and 21. Stand (duration of contact of a paw with the glass plate) was significantly higher in the NP group at days 7 and 14, whereas step cycle (duration between two consecutive initial contacts of the same paw) and duty cycle (stand as a percentage of step cycle) were the same at day 7. Long initial dual stance (duration of ground contact for both hind paws simultaneously, but the first one in a step cycle of a target hind paw) of the right hind paw was measured at days 7 and 14. The left hind paw per right hind paw ratio of the stand index (speed at which the paw loses contact with the glass plate) and mean intensity (mean intensity of the complete paw) changed at day 7 or 14. Phase dispersion (parameter describing the temporal relationship between placement of two paws) of the hind paws decreased at day 7.ConclusionsRats with applied NP showed a decreased withdrawal threshold and abnormal gait. The differences in gait parameters between the NP and sham groups were observed at an earlier time point than the withdrawal thresholds. Gait analysis could be an effective method for understanding pain caused by applied NP.

Effects of degenerative autologous nucleus pulposus on the lumbar sympathetic trunk: Sympathetic inflammation in rats

Objective We have reported the presence of discogenic visceral pain secondary to anterior herniation of the lumbar disc. The aim of this study was to observe the inflammatory response of the sympathetic trunk to an autologous degenerative nucleus pulposus (NP) injection under fluoroscopy. Methods A total of 72 rats were used. In 24 rats, the autologous NP suspension was injected into the right sympathetic trunk. Next, food intake and body weight of each rat were monitored for 14 days. Fourteen days after the injection, the right lumbar sympathetic trunk was harvested for histological assessment, and protein levels of IL-1 β, IL-6, and TNF- α were quantified via ELISA. Results In the NP-treated group, endoneural hyperemia and intensive infiltration of inflammatory cells could be identified in sections of the sympathetic trunk by the hematoxylin and eosin (H & E) stain. Meanwhile, elevated concentrations of IL-1 β, IL-6, and TNF- α were determined in the sympathetic trunk of the NP group as compared to that of the naive and control groups, which indicated the development of an inflammatory response. Furthermore, food intake and body weight of rats in the NP group decreased significantly. Conclusions The results indicated that an inflammatory response in the sympathetic trunk can be caused by anterior herniation of the lumbar disc, which can generate a sympathetic inflammatory pain response.

Low Affinity NGF Receptor (p75 Neurotrophin Receptor) Inhibit Sensory Nerve Growth in Human Degenerative Nucleus Pulposus Cells

Introduction Degeneration of the lumbar intervertebral disc is a cause of low back pain. The pathological mechanism is thought to be sensory nerve ingrowth into the inner layers of the degenerated intervertebral disc. NGF is also important for mediating inflammatory pain from intervertebral discs via the high-affinity receptor, TrkA. Recent research has also revealed that the low-affinity NGF receptor, p75 neurotrophic receptor (p75NTR), plays an important role in inflammatory pain. In most DRG neurons, innervating rat intervertebral discs were positive for two types of NGF receptors. However, the participative two types of NGF receptors have not been clarified in human degenerative discs. To evaluate the axonal growth and induction of a painful neuropeptide, CGRP, using neutralize antibody and extracted medium from human degenerative disc cells in vitro. Materials and Methods The nucleus pulposus (NP) of human intervertebral discs were harvested from patients with discogenic low back pain. Extracted medium from human degenerative intervertebral discs and rat DRGs were cultured with TrkA or p75NTR neutralize antibody. We evaluated the promotion of axonal growth and CGRP induction of DRG neurons in extracted medium from the NP using immunocytochemistry. Results The average length of growing axons in the NP and positive control group with TrkA or p75NTR neutralize antibody were significantly longer than that in the control group ( p &lt; 0.001). The average length of growing axons in the NP group and positive control group was significantly shortened after TrkA and p75NTR neutralize antibody treatment ( p &lt; 0.001). The percentage of CGRP-immunoreactive cells with growing axons was significantly shorter than in the NP and positive control group compared with the control groups with p75NTR neutralize antibody ( p &lt; 0.05). The expression of CGRP in all group with TrkA neutralize antibody was not decreased. Conclusion TrkA and p75NTR were associated with the growing axons in extracted medium from the NP of human intervertebral discs. p75NTR were associated with the percentage of CGRP-immunoreactive cells with growing axons more than TrkA. These in vitro results may suggest that p75NTR is related with discogenic low back pain. Disclosure of Interest None declared References Yamauchi K, Inoue G, Koshi T, et al. Nerve growth factor of cultured medium extracted from human degenerative nucleus pulposus promotes sensory nerve growth and induces substance p in vitro. Spine 2009;34(21):2263–2269 Sugiura A, Ohtori S, Yamashita M, et al. Effect of applying p75NTR saporin to a punctured intervertebral disc on calcitonin gene-related peptide expression in rat dorsal root ganglion neurons. J Orthop Sci 2010;15(3):407–413 Fukui Y, Ohtori S, Yamashita M, et al. Low affinity NGF receptor (p75 neurotrophin receptor) inhibitory antibody reduces pain behavior and CGRP expression in DRG in the mouse sciatic nerve crush model. J Orthop Res 2010;28(3):279–283

Sciatic Nerve Regeneration in Mice Induced by Transplantation of in Vitro Differentiated Nucleus Pulposus Progenitor Cells

Sciatic Nerve Regeneration in Mice Induced by Transplantation of in Vitro Differentiated Nucleus Pulposus Progenitor Cells

Experimental Noncompressive Disc Herniation Neuropathy Elicits Sustained Peripheral Hypersensitivity to Subthreshold Stimuli Mediated by Heightened TRPV1 Receptor Activity

Experimental Noncompressive Disc Herniation Neuropathy Elicits Sustained Peripheral Hypersensitivity to Subthreshold Stimuli Mediated by Heightened TRPV1 Receptor Activity

Inhibition of Nitric Oxide Mediated Protein Nitration

Experimental animal study. This study investigated whether nitric oxide (NO) mediated protein nitration is involved in the pathogenesis of radiculopathy and whether the symptoms can be relieved by its suppression. It has been reported that nitration of protein mediated by NO is involved in the degenerative neurological disorders, but its involvement is not clear in the radiculopathy. Two kinds of rat models of radiculopathy were used. Radiculopathy was induced either by ligation of spinal nerve roots or transplantation of autologous nucleus pulposus. In separate groups of rats, aminoguanidine, a potent nitric oxide synthetase inhibitor, was administered just before induction of radiculopathy, to suppress NO production and resultant nitration of protein. Sensation of the hind limb was evaluated by plantar stimulation test, and motor weakness was assessed by observation of gait pattern. Nitrotyrosine, product of protein nitration, was assayed quantitatively by Western immunoblotting. Mechanical allodynia was observed in both compression and nucleus pulposus groups, but motor weakness was observed only in the compression group. Preoperative administration of aminoguanidine attenuated mechanical allodynia and motor weakness. Optical densities of nitrotyrosine bands increased significantly in radiculopathy groups, but they were lowered by administration of aminoguanidine. NO mediated protein nitration contributes to the development of both types of radiculopathies. Suppression of NO production can decrease protein nitration and relieve neural dysfunctions of radiculopathy. N/A.

Cytokine Expression in the Epidural Space

Animal study. Development of an animal model for the study of biochemical changes that occur in the epidural space after intervertebral disc herniation. Although strong evidence for an inflammatory component exists, the biochemical processes underlying pain after disc herniation remain unknown. Epidural lavage was performed in 48 rats after L5 dorsal root ganglion exposure at baseline and 3, 6, or 24 hours after placement of autologous nucleus pulposus (NP) (N = 15), saline (N = 15), or NP + an interferon-γ antibody (anti-IFN-γ; N = 18) directly onto the dorsal root ganglion. Multiplex assays quantifying interleukin (IL)-1α, IL-1β, IL-2, IL-4, IL-6, IL-10, tumor necrosis factor α (TNF-α), IFN-γ, and granulocyte-macrophage colony-stimulating factor (GM-CSF) were performed. NP (N = 7) was also analyzed for these cytokines by placing NP into saline and measuring the relative concentration. Cytokines measured low at baseline (0-100 pg/mL) in all groups. Compared with saline, NP application caused IL-6 elevation, peaking at T = 3 hours, that was prevented by anti-IFN-γ. NP induced elevation of TNF-α, peaking at T = 24 hours and was prevented by anti-IFN-γ. IFN-γ was elevated after NP at T = 3 hours and T = 24 hours. IL-1α was similar after saline versus NP. The concentrations of IL-1β and IL-10 were elevated at T = 3 hours, 6 hours, and 24 hours in all groups without between-groups difference. The level of IL-4 peaked at T = 3 hours in the NP group and was different than saline and NP + anti-IFN-γ groups, but the time effect was insignificant. There was no change for GM-CSF. The concentration of cytokines measured in normal NP was less than 2 pg/mL for all cytokines except TNF-α. In this model of acute noncompressive disc herniation, NP caused the elevation of epidural IL-6, TNF-α, and IFN-γ--all attenuated by IFN-γ blockade. IL-1β and IL-10 were both significantly elevated by saline alone and their response was not prevented by IFN-γ blockade. This model may prove useful for the study of the biochemical processes by which NP induces inflammation-induced nerve root irritation and radiculopathic pain.

Clinical evaluation of interspinous process device Coflex for degenerative disk diseases

To study indications and complications of interspinous process device Coflex for degenerative disk diseases. One hundred and eight patients with degenerative lumbar disc diseases were underwent procedures of surgical decompression and additional fixation of Coflex between November 2007 and October 2010. Sixty-eight patients were male and the other fourty were female, and their average age was 53.5 years (range from 37 to 75 years). Fifty-nine patients were underwent surgery of excision of nucleus pulposus and Coflex fixation, 41 patients were underwent surgery of decompression by fenestration and Coflex fixation, 6 patients were underwent surgery of topping-off, and 2 patients were underwent surgery of Coflex fixation for two level. Preoperative and postoperative visual analogue scales (VAS) and Oswestry disability index (ODI) were recorded, as well as height of ventral intervertebral space (HV), height of dorsal intervertebral space (HD), height of intervertebral foramen (HIF) and segmental range of motion (ROM). One-way ANOVA was used for statistical analysis. Surgical complications were also recorded. The average follow-up time was 28.8 months. All groups had apparent improvement of VAS and ODI, and maintained well to last follow-up (F = 6.16-25.92, P = 0.00). Statistical analysis showed that HD and HIF increased significantly in group with excision of nucleus pulposus and Coflex fixation and group with decompression by fenestration and Coflex fixation (F = 7.37 - 11.68, P < 0.05). Although both HD and HIF decreased one-year after surgery, they were still higher than those preoperatively (F = 6.31 and 7.05, P = 0.00). Preoperative segmental ROM was respectively 6.3° ± 1.8° and 6.2° ± 1.7° in group with excision of nucleus pulposus and Coflex fixation and group with decompression by fenestration and Coflex fixation, and 3.1° ± 0.6° and 3.0° ± 0.8° at last follow-up. Three cases were found with device-related complications and five with non-device-related complications, and all five cased were cured after appropriate treatment. Surgical method assisted with Coflex has significant clinical efficacy for degenerative disc disease, it can maintain segmental stability, simultaneously, partly reserve movement. It's key to strictly master indications and precisely choose patients.

Analgesic Effects of Prostaglandin E2 Receptor Subtype EP1 Receptor Antagonist

The effect of an EP1 receptor antagonist on pain-related behavior induced by nucleus pulposus (NP) applied to the dorsal root ganglion (DRG) in rats was investigated. We investigated pain-related behavior, the amount of prostaglandin E2 (PGE2), and neural damage to the DRG after application of NP to the DRG after administration of an EP1 receptor antagonist. PGE2 induces mechanical allodynia and hyperalgesia, which are mediated by PGE2 receptors. EP1 is one of the PGE2 receptor subtypes. An EP1 antagonist reduces hyperalgesia, allodynia, and c-fos expression in the rat chronic nerve constriction model. Sprague-Dawley rats (n = 103) were used. Animals receiving NP were divided into three experimental groups (n = 12 in each group): saline, high-dose (5 mg/kg) EP1 receptor antagonist (RA), and low-dose (2.5 mg/kg) EP1-RA (orally once daily for 5 days). Animals in the sham group did not receive NP. Von Frey tests were used for pain-behavior testing. The amount of PGE2 in DRG and the number of activating transcription factor-3 (ATF3) immunoreactive positive cells were compared among groups. The mechanical thresholds in the three groups decreased 7 days after surgery (just before treatment). The threshold in both the high- and low-dose EP1-RA groups increased at 11 days (5 days after treatment) and continued for 14 days. The thresholds in both the low- and high-dose EP1-RA groups increased significantly compared with the saline group (P < 0.05). The amount of PGE2 was significantly increased in the NP group compared with the sham and naïve animals after application of NP. ATF3 expression was increased by NP but was not increased after administration of the EP1-RA. An EP1 receptor antagonist improves pain-related behavior in the rat model and might be a potential agent to improve pain-related behavior in patients with lumbar disc herniation.

Activation of satellite cells in the dorsal root ganglia in a disc-punctured rat model

BackgroundThe neural mechanisms underlying discogenic low back pain caused by disc degeneration remain unclear. Previous studies demonstrated that satellite cells (SC) play an important role in neuropathic pain. MethodsTwenty adult female Sprague-Dawley rats were used. The rats were divided into two groups: a nucleus pulposus (NP) group whose discs were punctured to expose the NP (n = 10) and a sham-operated group whose annulus fibrosus surface was scratched superficially (n = 10). In this study, we investigated the expression and cellular distribution of glial fibrillary acidic protein (GFAP, a marker of SC activation) in the dorsal root ganglia (DRG) innervating the intervertebral discs using a retrograde tracing method and immunohistochemistry in a discpunctured rat model. ResultsIn the sham-operated group, GFAP-immunoreactive (IR) SCs were not detected. In the NP group, GFAP-IR SC became evident, and 49 ± 13% of neurons innervating the punctured discs were surrounded by GFAP-positive SCs. ConclusionsOur results were the first to provide evidence for a potential role of SCs in the neural mechanisms of discogenic low back pain caused by disc degeneration.

The effect of repeated restraint stress in pain-related behavior induced by nucleus pulposus applied on the nerve root in rats

Chronic pain has an impact on psychological and social factors. It is known that stress influences physiological and behavioral changes and affects several neurotransmitter and hormonal systems. It is also known that corticosterone is increased by stress. The role of chronic stress in sciatica in lumbar disc herniation (LDH) in rats has not been investigated. The aim of this study was to investigate the effect of the restraint stress (RS) on pain-related behavior induced by application of nucleus pulposus (NP) in rats. Adult female Sprague-Dawley rats were divided into six experimental groups (naive group; naive+RS; sham group; sham+RS; autologous nucleus pulposus [NP] applied on the left L5 nerve root [NP group]; and NP+RS group). Von Frey tests were used to test pain-related behavior. Concentrations of plasma corticosterone were measured to assess changes in levels of endogenous corticosterone caused by RS. Expression of ATF-3 in the left L5 DRG was examined by immunohistochemical analyses in each group. Mechanical withdrawal thresholds of the NP and NP+RS groups were significantly decreased after surgery compared with the naive group. Although the thresholds in the NP group recovered after 28days, the thresholds in the NP+RS group were significantly decreased during the 42days after surgery. RS increased the concentration of plasma corticosterone at 21 and 42days after surgery. In the NP and the NP+RS groups, the expression of ATF-3 was significantly increased at 7days after surgery. The expression of ATF-3 was sustained for 21days by RS. Concentrations of plasma corticosterone were increased in three groups that underwent RS. The pain-related behavior persisted for the long term in the LDH model. The expression of ATF-3 in DRG neurons increased for 21days by RS. These results suggest that RS plays a role in the chronicity of pain-related behavior in the LDH rats.

Interaction of 5-Hydroxytryptamine and Tumor Necrosis Factor-α to Pain-Related Behavior by Nucleus Pulposus Applied on the Nerve Root in Rats

The effects of exogenous 5-hydroxytryptamine (5-HT), tumor necrosis factor(TNF)-α, 5-HT + TNF in combination, and autologous nucleus pulposus (NP) at dorsal root ganglion (DRG) were examined using rat models. To examine the interaction of 5-HT with TNF for pain-related behavior in a rat lumbar disc herniation (LDH) model. 5-HT and TNF have been shown to play roles in sciatica in lumbar disc herniation as chemical factors. Adult female Sprague-Dawley rats were divided into six groups: 5-HT group, TNF group, 5-HT + TNF (combination) group, NP group, control group, and naive group. Von Frey tests were used for pain-related behavior testing. Expressions of activating transcription factor 3 and calcitonin gene-related peptide (CGRP) were evaluated immunohistochemically. Expressions of TNF, TNF receptor 1 (TNFR1), and 5-HT2A receptors in the left L5 DRG were examined using western blotting. Plasma levels of 5-hydroxyindole acetic acid, a metabolite of 5-HT, were measured. Mechanical withdrawal thresholds were significantly decreased in the 5-HT, TNF, combination, and NP groups compared with controls. Thresholds recovered after 14 days in the 5-HT and TNF groups, and after 28 days in the combination group. Exogenous 5-HT and TNF to the nerve root induced pain-related behavior and lasted for a shorter period compared with combination and NP groups. Activating transcription factor 3- and calcitonin gene-related peptide-immunoreactive DRG neurons were significantly increased only in the early phase in 5-HT, TNF, combination, and NP groups. TNF induced 5-HT2A receptor expressions in the DRG, while 5-HT induced TNF and TNF receptor 1 expressions. The present findings suggest that both 5-HT and TNF induce pain-related behavior and interact with each other to prolong pain-related behavior in a rat LDH model.

A COMPARISON OF PAIN-RELATED BEHAVIOR FOLLOWING LOCAL APPLICATION OF NUCLEUS PULPOSUS AND/OR MECHANICAL COMPRESSION ON THE DORSAL ROOT GANGLION

Symptomatic induction of disc herniation involves both mechanical compression and chemical factors. Inhibitors of tumor necrosis factor-alpha (TNF-α) are known to reduce pain-related behavior in experimental models. Animals were divided into mechanical compression (MC) group; a stainless steel rod was inserted on the dorsal root ganglion, nucleus pulposus (NP) group: NP was harvested from the coccygeal vertebral disc, MC and NP group; rats were received stainless rod and NP, and sham group; rats were received neither rod nor NP. Rats in the MC group received a TNF-α antibody (10 mg/kg) (antibody group) or were not treatment (untreated group). The withdrawal thresholds of the MC, NP and MC+NP groups decreased significantly compared with the sham group. In the antibody group, the threshold was significantly higher than that of the untreated group. An anti-TNF-α antibody reduced allodynia caused by DRG compression.

Increased Expression of Vascular Endothelial Growth Factor Protein in Dorsal Root Ganglion Exposed to Nucleus Pulposus on the Nerve Root in Rats

An experimental rat study. To assess pain-related behavior and vascular endothelial growth factor (VEGF) expression induced by nucleus pulposus (NP) applied to nerve roots in rats. Radiculopathy from disc herniation is caused by nerve compression and chemical inflammation. In experimental studies, a decrease in mechanical withdrawal threshold, blood flow, and nerve root dysfunction was reported when the NP was applied to nerve roots. However, neuron reproduction and changes in nerve root blood vessels have not been clearly understood. NP harvested from the tail was applied to the left L5 dorsal root ganglion (NP group; n = 77). As a control, sham-operated animals were used (n = 77). Behavioral testing with von Frey hairs was performed for 35 days. Immunoreactivity (IR) for VEGF, activating transcription factor-3, growth-associated protein-43 (GAP-43), factor VIII, and hypoxia-inducible factor-1[alpha] (HIF-1 a) were studied by immunohistochemistry. Western blot analyses of VEGF and GAP-43 were also performed. The mechanical withdrawal threshold significantly decreased from 7 to 28 days in the NP group versus the sham group (P < 0.05). In the NP group, activating transcription factor-3-IR cells increased from 3 to 14 days (P < 0.05), hypoxia-inducible factor-1[alpha]-IR cells increased at 14 days (P < 0.05), and blood vessels with Factor VII-IR cells increased at 28 days (P < 0.05) compared with the sham group. The expression levels of VEGF and GAP-43 in the NP group significantly increased at 14 and 28 days (P < 0.05). Neuron damage induced by NP applied to the nerve root at the early stage, and axon extension occurred from 14 days. VEGF increased at 14 and 28 days, and the numbers of blood vessels increased 28 days after surgery. The mechanical withdrawal threshold improved at 35 days. Regeneration and vascularization by VEGF might be associated with pain-related behavior.

Nerve Growth Factor of Cultured Medium Extracted From Human Degenerative Nucleus Pulposus Promotes Sensory Nerve Growth and Induces Substance P In Vitro

We investigated the mechanism of discogenic low back pain using an in vitro model. To evaluate the axonal growth and induction of a painful neuropeptide, substance P (SP), using rat dorsal root ganglion (DRG) neurons and degenerated human disc cells in vitro. Degeneration of the lumbar intervertebral disc is a cause of low back pain. The pathologic mechanism is thought to be sensory nerve ingrowth into the inner layers of the degenerated intervertebral disc; however, the precise patho-mechanism has not been clarified. The nucleus pulposus (NP) and annulus fibrosus (AF) of human intervertebral discs were harvested from patients with discogenic low back pain. Extracted medium from human degenerative intervertebral discs was cultured with neurons of rat DRGs. We evaluated the promotion of axonal growth and SP induction of DRG neurons in extracted medium from the NP and AF using immunocytochemistry. The average length of growing axons in the NP and AF was significantly longer than that in the control (P < 0.005). That in the NP was significantly longer than that in the AF. The average length of growing axons in the NP was significantly shortened after anti-nerve growth factor (NGF)beta treatment (P < 0.005); however, that in the AF was not (P > 0.05). The percentage of SP-immunoreactive cells with growing axons was significantly higher only in the NP group compared with the control and AF groups (P < 0.005), and anti-NGFbeta treatment decreased the expression of SP in the NP group (P < 0.05). Extracted medium from the NP and AF promoted axonal growth. Furthermore, NGF from the NP promoted axonal growth and induced SP. These in vitro results may suggest that NGF from the NP promotes the growth of sensory nerve fibers innervating the degenerated intervertebral disc and may induce SP related with pain transmission.

The effects of punctured nucleus pulposus on lumbar radicular pain in rats: a behavioral and immunohistochemical study

Application of the nucleus pulposus (NP) to the nerve root has been developed as a model of lumbar radicular pain. The relationship between disc degeneration and the induction of radicular pain, however, has not yet been fully explored. The authors of this study investigated pain-related behaviors and expression of tumor necrosis factor-alpha (TNF-alpha) in the dorsal root ganglion (DRG) to evaluate the effects of punctured NP on lumbar radiculopathy. An anular needle puncture model of intervertebral disc degeneration in a rat tail was established. Normal and previously punctured NP tissues were obtained and placed on the L-5 nerve root following a hemilaminectomy. Behavioral tests including assessment of motor function, mechanical threshold, and thermal withdrawal latency were performed before and after surgery. The TNF-alpha immunoreactivity in L-5 DRG specimens was examined through immunohistochemical study. The punctured discs showed significant degeneration 2 weeks after intervention. Application of both normal and punctured NP induced mechanical hyperalgesia in the ipsilateral paw for 10 days after surgery, but hyperalgesia was more severe in the punctured NP group. No statistically significant within-group changes in thermal withdrawal latency over time were found. A significant increase in the expression of TNF-alpha-positive neurons in DRG specimens was observed in both NP graft groups. Needle puncture led to degenerative changes in the rat tail disc, and the degenerated NP enhanced mechanical hyperalgesia induced by application of the NP to the lumbar nerve root. This model of disc degeneration and lumbar radicular pain is appropriate for evaluating the efficacy of biological treatments for degenerative disc diseases.