AO Spine Reference Centre & Institute of Health & Biomedical Innovation, Queensland University of Technology, Brisbane, Australia. Traumatic spinal cord injury (SCI) is a devastating condition with no curative therapy. Pro-inflammatory therapy has been suggested recently to try and reduce the inhibitory glial scar and promote neural regeneration and healing. The aim of this study is to investigate the potential of sustained delivery of angiogenic/pro-inflammatory growth factors to reduce the secondary degeneration after spinal cord injury. Adult male Wistar Kyoto rats (200-300g; 12-16weeks old) were subjected to cord hemisections via a T10 laminectomy. Animals were randomised to treatment or control groups after the spinal cord injury had been induced. Treatment consisted of implantation of a mini-osmotic pump capable of delivering 5 micrograms vascular endothelial growth factor (VEGF) and 5 micrograms platelet-derived growth factor (PDGF), via a catheter, to the site of the lesion, over 7 days(n=6). Control animals were subjected to either cord lesion only (n=6) or lesion plus mini-pump delivering PBS (phosphate-buffered saline) solution (n=6). Rats were sacrificed at one month and the spinal cords were harvested and examined by immunohistology, using anti-neurofilament-200 and anti-Glial Acidic Fibrillary Acidic Protein (GFAP) antibodies. RESULTS: Active treatment spinal cords showed a higher level with aboration of the axonal filament through the defect and more dense neurofilament-200 staining at the lesion site compared to both control groups. The treatment also showed the elevated presence of activated microglia in the lesion, whilst distal to the lesion the microglia and astrocytes retained an unreactive phenotype. Pro-inflammatory therapy in the rat spinal cord-injury model showed favourable histological findings after sustained delivery of PDGF and VEGF

Aims. In this investigation, we administered oxidative stress to nucleus pulposus cells (NPCs), recognized DNA-damage-inducible transcript 4 (DDIT4) as a component in intervertebral disc degeneration (IVDD), and devised a hydrogel capable of conveying small interfering RNA (siRNA) to IVDD. Methods. An in vitro model for oxidative stress-induced injury in NPCs was developed to elucidate the mechanisms underlying the upregulation of DDIT4 expression, activation of the reactive oxygen species (ROS)-thioredoxin-interacting protein (TXNIP)-NLRP3 signalling pathway, and nucleus pulposus pyroptosis. Furthermore, the mechanism of action of small interfering DDIT4 (siDDIT4) on NPCs in vitro was validated. A triplex hydrogel named siDDIT4@G5-P-HA was created by adsorbing siDDIT4 onto fifth-generation polyamidoamine (PAMAM) dendrimer using van der Waals interactions, and then coating it with hyaluronic acid (HA). In addition, we established a rat puncture IVDD model to decipher the hydrogel’s mechanism in IVDD. Results. A correlation between DDIT4 expression levels and disc degeneration was shown with human nucleus pulposus and needle-punctured rat disc specimens. We confirmed that DDIT4 was responsible for activating the ROS-TXNIP-NLRP3 axis during oxidative stress-induced pyroptosis in rat nucleus pulposus in vitro. Mitochondria were damaged during oxidative stress, and DDIT4 contributed to mitochondrial damage and ROS production. In addition, siDDIT4@G5-P-HA hydrogels showed good delivery activity of siDDIT4 to NPCs. In vitro studies illustrated the potential of the siDDIT4@G5-P-HA hydrogel for alleviating IVDD in rats. Conclusion. DDIT4 is a key player in mediating pyroptosis and IVDD in NPCs through the ROS-TXNIP-NLRP3 axis. Additionally, siDDIT4@G5-P-HA hydrogel has been found to relieve IVDD in rats. Our research offers an innovative treatment option for IVDD. Cite this article: Bone Joint Res 2024;13(5):247–260

Aims. Inflammatory response plays a pivotal role in the pathophysiological process of intervertebral disc degeneration (IDD). A20 (also known as tumour necrosis factor alpha-induced protein 3 (TNFAIP3)) is a ubiquitin-editing enzyme that restricts nuclear factor-kappa B (NF-κB) signalling. A20 prevents the occurrence of multiple inflammatory diseases. However, the role of A20 in the initiation of IDD has not been elucidated. The aim of the study was to investigate the effect of A20 in senescence of TNF alpha (TNF-α)-induced nucleus pulposus cells (NPCs). Methods. Immunohistochemical staining was performed to observe the expression of A20 in normal and degenerated human intervertebral discs. The NPCs were dissected from the tail vertebrae of healthy male Sprague-Dawley rats and were cultured in the incubator. In the experiment, TNF-α was used to mimic the inflammatory environment of IDD. The cell viability and senescence were examined to investigate the effect of A20 on TNF-α-treated NPCs. The expression of messenger RNA (mRNA)-encoding proteins related to matrix macromolecules (collagen II, aggrecan) and senescence markers (p53, p16). Additionally, NF-κB/p65 activity of NPCs was detected within different test compounds. Results. The expression of A20 was upregulated in degenerate human intervertebral discs. The A20 levels of NPCs in TNF-α inflammatory microenvironments were dramatically higher than those of the control group. TNF-α significantly decreased cell proliferation potency but increased senescence-associated beta-galactosidase (SA-β-Gal) activity, the expression of senescence-associated proteins, the synthesis of extracellular matrix, and G1 cycle arrest. The senescence indicators and NF-κB/p65 expression of A20 downregulated group treated with TNF-α were significantly upregulated compared to TNF-α-treated normal NPCs. Conclusion. A20 has a self-protective effect on the senescence of NPCs induced by TNF-α. The downregulation of A20 in NPCs exacerbated the senescence of NPCs induced by TNF-α. Cite this article:Bone Joint Res. 2020;9(5):225–235

Introduction. Melatonin-deficient rats are known to develop scoliosis when rendered bipedal. In a previous study we have shown that melatonin-deficient bipedal mice with scoliosis had lower bone density than did mice without scoliosis. Published work suggests that children with AIS have lower bone density than do healthy children. The aim of this study is to establish whether osteoporosis causes scoliosis. We hypothesised that bipedal rats with lower bone density would have increased spinal malalignment compared with the control group. Methods. 50 female Sprague-Dawley rats were rendered bipedal at 3 weeks of age by amputation of the forelimbs and tails. Two groups were formed: control group (n=25), in which rats received no drug; and the experiment group (n=25), in which rats received daily subcutaneous 1 U/g heparin injections. Animals were kept in standard cages, and food and water was provided at the top of the cages to encourage more time standing erect. DEXA scans were done on week 4 to assess bone density. Radiographs were taken on week 40 to assess spinal alignment in both control and experiment groups. Results. 19 rats in the heparin group and 23 rats in the control group were available for evaluation at the end of the study. At week 4, DEXA scans showed significant difference between the bone densities of the control and heparin groups (p<0·05), with the heparin group having lower bone density. The incidence of curves between the heparin and control groups were not statistically significant (p>0·01) (table). The magnitude of curves in scoliotic rats for the heparin group was 11·8° (SD 3·75) and for the control group 10° (4·3). The difference between the groups was not significant (p>0·05). Conclusions. This study involved rats with normal melatonin levels and both groups showed a high frequency of scoliosis incidence. Although no significant differences were recorded between groups, the results suggest that bipedality is a cause for scoliosis, and low bone mineral density may further increase this tendency

This review provides a concise outline of the advances made in the care of patients and to the quality of life after a traumatic spinal cord injury (SCI) over the last century. Despite these improvements reversal of the neurological injury is not yet possible. Instead, current treatment is limited to providing symptomatic relief, avoiding secondary insults and preventing additional sequelae. However, with an ever-advancing technology and deeper understanding of the damaged spinal cord, this appears increasingly conceivable. A brief synopsis of the most prominent challenges facing both clinicians and research scientists in developing functional treatments for a progressively complex injury are presented. Moreover, the multiple mechanisms by which damage propagates many months after the original injury requires a multifaceted approach to ameliorate the human spinal cord. We discuss potential methods to protect the spinal cord from damage, and to manipulate the inherent inhibition of the spinal cord to regeneration and repair. Although acute and chronic SCI share common final pathways resulting in cell death and neurological deficits, the underlying putative mechanisms of chronic SCI and the treatments are not covered in this review.

Objectives. In order to elucidate the influence of sympathetic nerves on lumbar radiculopathy, we investigated whether sympathectomy attenuated pain behaviour and altered the electrical properties of the dorsal root ganglion (DRG) neurons in a rat model of lumbar root constriction. Methods. Sprague-Dawley rats were divided into three experimental groups. In the root constriction group, the left L5 spinal nerve root was ligated proximal to the DRG as a lumbar radiculopathy model. In the root constriction + sympathectomy group, sympathectomy was performed after the root constriction procedure. In the control group, no procedures were performed. In order to evaluate the pain relief effect of sympathectomy, behavioural analysis using mechanical and thermal stimulation was performed. In order to evaluate the excitability of the DRG neurons, we recorded action potentials of the isolated single DRG neuron by the whole-cell patch-clamp method. Results. In behavioural analysis, sympathectomy attenuated the mechanical allodynia and thermal hyperalgesia caused by lumbar root constriction. In electrophysiological analysis, single isolated DRG neurons with root constriction exhibited lower threshold current, more depolarised resting membrane potential, prolonged action potential duration, and more depolarisation frequency. These hyperexcitable alterations caused by root constriction were significantly attenuated in rats treated with surgical sympathectomy. Conclusion. The present results suggest that sympathectomy attenuates lumbar radicular pain resulting from root constriction by altering the electrical property of the DRG neuron itself. Thus, the sympathetic nervous system was closely associated with lumbar radicular pain, and suppressing the activity of the sympathetic nervous system may therefore lead to pain relief

Aims

Non-coding microRNA (miRNA) in extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) may promote neuronal repair after spinal cord injury (SCI). In this paper we report on the effects of MSC-EV-microRNA-381 (miR-381) in a rodent model of SCI.

Introduction. The response of the intervertebral disc to asymmetric forces may accelerate degeneration through changes in the matrix. Macroscopically, the disc sustains structural changes that may play a part in the progression of a scoliotic curve. Molecularly, disc degeneration is the outcome of the action of matrix metalloproteases (MMPs), members of a family of enzymes that bring about the degradation of extracellular matrix components. In this study we measured in vivo the expression of MMPs in a rat scoliotic intervertebral disc and studied the effect of the degree of the deformity on their production. Methods. Asymmetric forces were applied in the intervertebral disc between the ninth and tenth vertebrae at the base of a rat tail with the use of a mini Ilizarov external fixator, under anaesthesia. Animals were categorised into three groups according to the degree of the deformity. In group I, the deformity that was applied on the intervertebral disc was 10°, in group II 30°, and in group III 50°. All the animals used were female Wistar rats before adulthood, to take into account the effect of growth for the study of intervertebral disc changes. The intact intervertebral discs outside the fixator were used as controls. After the rats' death on day 35, the tails were prepared and analysed with an immunohistochemical protocol for chromogenic detection and location of MMPs 1 and 12 in tissue sections of the intervertebral discs. Results. We recorded an increase of the concentration of the MMPs in all groups compared with controls. The quantity of the MMPs increased as the degree of the deformity progressed. MMPs were detected mainly in fibrocartilage cells of the degenerative part, which were formed as result of the compression forces. We detected a differentiation of a large number of disc cells into chondrocytes at the transitional zone of the intervertebral disc adjacent to the vertebral end plates. Conclusions. The application of asymmetric forces on the intervertebral discs of a rat tail results in an increase of MMP expression in the disc cells. The amount of MMPs produced is proportional to the degree of the deformity and has an asymmetrical pattern of distribution into the intervertebral disc

Background. We have reported an injectable L-pNIPAM-co-DMAc hydrogel with hydroxyaptite nanoparticles (HAPna) which promotes mesenchymal stem cell (MSC) differentiation to bone cells without the need for growth factors. This hydrogel could potentially be used as an osteogenic and osteoconductive bone filler of spinal cages to improve vertebral body fusion. Here we investigated the biocompatibility and efficacy of the hydrogel in vivo using a proof of concept femur defect model. Methods. Rat sub-cut analysis was performed to investigate safety in vivo. A rat femur defect model was performed to evaluate efficacy. Four groups were investigated: sham operated controls; acellular L-pNIPAM-co-DMAc hydrogel; acellular L-pNIPAM-co-DMAc hydrogel with HAPna; L-pNIPAM-co-DMAc hydrogel with rat MSCs and HAPna. Following 4 weeks, defect site and organs were histologically examined to determine integration, repair and inflammatory response, as well as Micro-CT to assess mineralisation. Results. No inflammatory reactions or toxicity were seen in any animal. Enhanced bone healing was observed in aged exbreeder female rats where hydrogel was injected with increased deposition of collagen type I. Integration of the hydrogel with surrounding bone was observed without the need for delivered MSCs; native cell infiltration was also seen and bone formation was observed within all hydrogel systems investigated. Conclusion. This novel hydrogel is biocompatible, facilitates migration of cells, promotes increased bone formation and integrates with surrounding bone. This system could be injected to fill spaces within and surrounding spinal cages to aid in cage fixation and spinal fusion without the need for harvesting of bone autografts, thus reducing operative risk and surgical cost. Conflicts of Interest: None. Source of Funding: BMRC, MERI Sheffield Hallam University

Mesenchymal stem-cell based therapies have been proposed as novel treatments for intervertebral disc degeneration, a prevalent and disabling condition associated with back pain. The development of these treatment strategies, however, has been hindered by the incomplete understanding of the human nucleus pulposus phenotype and by an inaccurate interpretation and translation of animal to human research. This review summarises recent work characterising the nucleus pulposus phenotype in different animal models and in humans and integrates their findings with the anatomical and physiological differences between these species. Understanding this phenotype is paramount to guarantee that implanted cells restore the native functions of the intervertebral disc.

Cite this article: Bone Joint Res 2013;2:169–78.