Ankle sprains are common athletic injuries, with a peak lifetime incidence between the ages of 15 and 19 years, especially in young males. However, an unclear history, an imprecise physical exam, and unhelpful radiographies lead to frequent misdiagnosis of paediatric ankle traumas, and subsequently, inappropriate treatment. Improper management may lead to residual pain, instability, slower return to physical activity, and long-term degenerative changes. The purpose of this study was to evaluate the initial management and treatment of acute paediatric ankle sprains at our center, a tertiary care paediatric hospital. Our hypothesis was that the initial diagnosis is often incorrect, and treatment varies considerably amongst orthopaedic surgeons.

We conducted a retrospective study of all cases of ankle sprains and Salter-Harris one (SH1) fractures referred to our orthopaedic surgery service between May and August 2014. Exclusion criteria included ankle fractures other than SH1 types, and cases where treatment was initially undertaken elsewhere before referral to our service. Patients were evaluated on a clinical and radiographic basis. Primary outcome was the difference between initial and final diagnosis. Secondary outcome was variation in immobilisation duration for each diagnosis. The main variables we considered were age, sex, mechanism of trauma, referral delay, patient symptoms, physical exam findings, radiographic findings, type and duration of immobilisation, prescription of any medication, and referral to physical therapy.

A total of 3047 patients were reviewed and 31 cases matched our inclusion criteria, comprised of 17 girls and 14 boys, with a mean age of 10.4 years. Patients were seen at a mean of 10.3 days after injury. Initial diagnosis was SH1 fracture in 20 cases, acute ankle sprain in 8 cases, and uncertain in 3 cases. Final diagnosis was SH1 fracture in 11 cases, acute ankle sprain in 13 cases, uncertain in 5 cases, and other in 3 cases. During follow up, 48.5% of cases saw a change in diagnosis. Forty five percent (9/20) of cases initially diagnosed as SH1 fractures proved to be incorrect, with 55.5% (5/9) of these being ultimately diagnosed as acute ankle sprains. Amongst cases initially diagnosed as acute ankle sprains, 37.5% (3/8) received a different final diagnosis. Duration of immobilisation was significantly different between acute ankle sprain and SH1 fracture groups, with an average of 17.3 days and 26.1 days, respectively. Physical therapy was prescribed to 33.3% of acute ankle sprains and 9.1% of SH1 fractures.

Initial distinction between acute ankle sprains and SH1 fractures can be difficult in paediatric ankle trauma. Case management and specific treatments vary considerably, as there is neither an evaluation algorithm nor consensus on treatment of these paediatric pathologies. This study reinforces the need to develop a systematic diagnostic and treatment protocol for paediatric ankle sprains.

Anterior lumbar inter-body fusion (ALIF) is a surgical procedure that is available to chronic lower back pain patients who fail to respond to conservative treatments. Failure to achieve fusion may result in persistence of pain. Fusion of the lumber vertebral segment is more accurately assessed using fine-cut helical Computed tomography (CT) scans (0.25 mm thickness slices). Unfortunately this technique exposes the body to high radiation dose with hazard of increase risk of late malignancy. An alternative imaging tool is radiostereometry (RSA) which developed as a means to determine the magnitude of relative motion between two rigid bodies. In this study we used RSA to detect movement at the fused lumbar segment (ALIF site) during flexion and extension and compare the results obtained with fine-cut helical CT scan using histopathology as final gold standard assessment tool.

ALIF of three levels of lumbar spine (L1-L2, L3-L4, and L5-L6) was done in 9 sheep. The sheep divided into three groups (3sheep each). The first group had RSA assessment immediately, 3, and 6 months after surgery. The second group had RSA immediately, 3, 6, 9 months after surgery. The third group had an RSA immediately, 3, 6, 9, 12 months after surgery All the animals were humanly killed immediately after having the last scheduled RSA (group1, group2, and group 3 sheep were killed 6 month, 9month and 12 months after surgery respectively). This followed by in vitro fine cut CT and histopathology after the animals are scarified. Micro CT scan has been also used to identify the area where histopathology slide should be made to pick up fusion. Fine cut CT scan assessment for all sheep were done. The CT scan has been reported by two independent radiologists. Histopathology has been started and will finish in 2 weeks

RSA showed there was significant increasing stiffness of the spine though the fused segments as the time pass on compare to immediate postoperative assessment. CT scan were done and showed variable fusion though out the spinal segments. Histopathology of all sheep has been started and the results will be available in 2 weeks which will be followed by statistical assessment to decide how accurate RSA compare to CT scan in assessment of fusion.

Introduction: Thickened ligamentum flavum (LF) is a major contributor to the clinical syndrome of lumbar canal stenosis (LCS). The patho-mechanisms responsible for this phenomenon remain unclear. Cysts adjacent to facet joints (FJ) in the spine are regarded as rare entities that may uncommonly contribute to LCS. Inaccurate pathological interpretation and unawareness of a key anatomical feature has generated erratic terminology and confusion about their origin.

Methods: Twenty-seven consecutive patients with radiologically confirmed central canal or lateral recess stenosis underwent lumbar laminectomy for neurogenic symptoms. Surgical specimens comprising en bloc excision of LF and medial inferior facet (to retain LF and FJ relationships) were examined microscopically following staining with haematoxylin-eosin and Miller’s elastic stain. Controls were facet/LF specimens from 89 cadaver lumbar spines.

Results: Mean LF thickness was 8.9 mm (+/− 0.3 mm SEM) at the operated levels and 2.9 mm (+/− 0.3 mm) at the non-operated, adjacent levels (p < 0.01). Twenty-eight synovial cysts (8 bilateral, 12 unilateral) were present at a single level in 20 (74%) patients. Synovial cysts per spine level were: L1/2 = 0; L2/3 = 3; L3/4 = 7; L4/5 = 16; L5/S1 = 2. The cyst levels all showed advanced osteoarthritis and LF degeneration. Ten patients (50 %) with cysts had pre-existing degenerative spondylolisthesis (DS). Only 5 patients had pre-operative radiological apperances of unilateral facet cysts. Therefore 82 % of our observed synovial cysts were microscopic or occult. The synovial cysts communicated with the FJ via a bursa-like cleft within the LF, and their linings of synoviocytes and other cells contained fragments shed from the articular surface. The control cadaver specimens revealed that a synovial bursa or intra-ligamentous out-pouching from the synovial cavity was present in 90% of normal LF at L4/5 and was up to 12 mm in length. This intra-ligamentous synovial recess, either wholly or partially lined by synoviocytes, was only present in 55% of specimens at L1/2 with a maximum length of 5 mm. Several other juxtafacet cyst types were observed in the experimental group and a novel classification based upon pathological findings is presented.

Discussion: Para-facetal intraspinal cysts are common in degenerative lumbar spinal stenosis. DS is also a frequent finding but is statistically unrelated to cyst formation (Chi-square: p=0.187). We have found that debris from osteoarthritic facet joints enters a bursa-like cleft within the LF where it becomes incorporated into the wall where it excites a granulomatous reaction leading to blockage and synovial cyst formation. The existence of this channel has not been reported previously. We suggest that microscopic synovial cysts contribute significantly to the ligamentous thickening seen in LCS. We also present a novel classification of juxtafacet cysts based on our pathological findings.

Introduction: The age-related reduction of water-trapping proteoglycans needed to maintain optimal disc hydration may be caused by reduced synthetic ability or fewer chondrocytes but there is a paucity of objective quantitative studies of disc cellularity.

Methods: Sagittal sections of L4–5 discs were subdivided into a nucleus zone and 18 annulus zones prior to determining cell density (cells/mm2) in

the mid-sagittal plane of 10 male and 10 female discs aged 13–79 years;

Results:

Most chondrocytes were unicellular but bicellular and multicellular chondrons were common in the margins of large tears and the nucleus in degenerate discs.

Discussion: The findings that cell density is higher in the posterior annulus and in the right half of the disc, tends to be increased if the cartilage end-plate is thicker, and is not uniformly diminished by large tears, indicates that disc cellularity is influenced by a complex interplay of factors which needs to be understood before attempts are made to restore the structural and functional integrity of degenerate discs.

Introduction: Current treatments for osteoporosis do not completely eliminate the risk of fracture and bone loss may continue even at a low level. Enhanced bone formation and mineralization could minimize the risk of fracture in osteoporosis and prevent the pain and associated morbidity in these patients. Bone morphogenetic protein-type 2 has been successfully used to promote bone formation and to augment fracture repair in general and in the spine in particular [1]. The aim of this study was to increase local bone formation and mineralization in osteopenic vertebrae by administration of recombinant human morphogenetic proteins (rhBMP-2) in an ovine model.

Methods: Osteoporosis was induced in ten skeletally mature sheep with ovariectomy, low calcium diet and weekly steroid injection. Bone mineral density (BMD) of the lumbar spine was assessed monthly by DXA. When the BMD of the lumbar spine was reduced by at least 25% the induction treatment was stopped and pellets containing inert carrier alone (control) or rhBMP-2 in either slow or fast release formulation were implanted directly into three adjacent lumbar vertebrae of each animal in a random order. BMD was assessed at regular intervals and two and three months later five animals were euthanized and the lumbar spines were collected for histomorphometric analysis using the SkyScan 1076 Micro CT (SkyScan, Belgium). Significant differences between BMD and bone morphometric data (including trabecular bone volume, separation and number) were examined using ANOVA and Tukey’s test with significance set at P< 0.05.

Results: After five months of induction treatment BMD in the lumbar spines of all animals was reduced by at least 25% (p< 0.05). BMD increased insignificantly after cessation of the induction treatment but remained lower than the initial values. As there were no significant differences the histomorphometric data after two and three months were pooled. The trabecular bone volume in the vicinity of both the slow and fast release BMP implants increased by over 15% compared with the control (p< 0.05). Trabecular separation was reduced over 13% and trabecular number around both types of pellets increased by over 12% compared to the control (NS).

Discussion: This animal model provides an opportunity to evaluate systemic and local treatments for osteoporosis. The significant increase in bone formation adjacent to the implants as early as two months suggests that rhBMP-2 in either formulation improves bone quality at sites with high risk of fracture. The impact of the fast and slow release BMPs implants were not significantly different

Introduction: Interspinous implants have recently been proposed as an alternative in the treatment of lumbar spinal stenosis and foraminal stenosis. Interspinous implants are thought to unload the facet joints, restore foraminal height and provide improved spinal stability especially in extension with a minimally invasive approach. It has been proposed that, combined with a tension band, stabilisation could also be obtained in flexion, thus avoiding the need for pedicle screw fixation. Little biomechanical data exists to support these notions. The aim of this in vivo study was to investigate the effect of a novel, minimally invasive, unilaterally inserted interspinous implant on flexion-extension range of motion of the lumbar spine.

Methods: Following the induction of general anesthesia, ten adolescent Merino lambs (24–30 kg) underwent a destabilisation procedure at the level of L1–L2, thus simulating a stenotic degenerative spondylolisthesis, as described previously. All animals were placed in a sidelying posture and lateral radiographs were taken in the neutral posture and in end stage flexion and extension trunk positions with the central ray at the level of L1–L2. The flexion manouvre was reproduced in each subject by securing a rope above the carpus (forelimb) and the tarsus (hindlimb). This same radiographic protocol was repeated following the insertion of an 8 mm InSwing interspinous device at L1–L2, and again with the implant secured by means of a tension band tightened to 1 N/m around the L1 and L2 spinous processes. Care was given to respect the integrity of the supra-spinous ligament. The insertion technique requires a minimally invasive unilateral approach, therefore leaving the attachment of the erector spinae muscle on the contralateral side intact. Using Cobb’s method, intersegmental range of motion (ROM) was assessed in each of the conditions and compared. A paired t-test compared ROM for each of the experimental conditions (P< .05).

Results: The addition of the InSwing interspinous implant reduced mean total flexion-extension ROM from 6.3 degrees to 5.3 degrees which was further reduced to 3.6 degrees with the device secured by means of a tension band. These differences were not statistically significant, but the addition of the tension band to the interspinous device resulted in a significant reduction of lumbar flexion (p< .05).

Discussion: The interspinous device tended to reduce the total flexion-extension ROM at the level of the implant, however the results were not significant. The addition of a tension band was found to significantly stabilize the spine in flexion. To our knowledge, this is the first in vivo study radiographically showing the advantage of using an interspinous device, specifically InSwing, to stabilize the spine in flexion. These results are important findings particularly for patients with clinical symptoms of instable degenerative spondylolisthesis.

Introduction: Spondylolysis in skeletally immature individuals is common but only a small proportion of cases develop pain and spondylolisthesis. The pattern of instability and pathologic consequences of surgically created spondylolytic defects have not previously been described. An animal model of lumbar spondylolysis was created using skeletally immature sheep with the aim of observing the intervertebral mobility and pathologic consequences of creating a spondylolytic segment.

Methods: Bilateral spondylolytic defects were created in the fifth lumbar vertebrae of fourteen sheep aged approximately 16 weeks using a posterior surgical approach. Using tantalum markers that were placed in the lumbar spine segments in the vicinity of the lesion, three dimensional translations and rotations in flexion and extension were recorded under general anaesthetic using Radiostereometric Analysis (RSA) at the time of surgery. A novel method of ovine spine manipulation was developed. Briefly, with the animal in a lateral decubitus position spinal extension was achieved by fixing the shoulders and hips in full extension. Similarly, the limbs were brought into apposition to achieve spinal flexion. A control group of seven sheep had tantalum markers only in the same lumbar spine segments. The animals were re-tested under general anaesthetic at six weeks and six months following surgery. After six months the animals were sacrificed and their spines examined using CT and microscopic analysis.

Results: Bilateral spondylolytic defects did not result in significant transverse (x-axis) translation. RSA showed significant differences between the spondylolytic group (mean 1.22°, range 0.30–3.74) and controls (mean 0.28°, range 0.03–0.77)) for rotation in the axial plane (y-axis, p< 0.0025) immediately after surgery. The spines radiologically stabilised over six months.

Discussion: Surgically induced bilateral spondylolytic defects in this immature ovine model result in increased rotation in the axial plane but do not create transverse translation (spondylolisthesis).

Introduction Sheep are being used increasingly for spinal and other skeletal-related research. However, there is still limited information about the molecular pathways of bone remodelling in this species compared to rats or mice. It has been demonstrated in other animal models and in the human that the receptor activator of nuclear factor kappaB ligand (RANKL) and osteoprotegerin (OPG) play major regulatory roles in controlling osteoclast activity and their differentiation. We investigated the expression of RANKL and OPG in trabecular bone of an ovariectomised steroid-treated osteopaenic sheep model.

Methods Trabecular bone from the lumbar spine (LS) and proximal femur (PF) of ten osteopaenic ewes and four normal ewes were collected [1]. Total RNA was isolated and complementary DNA (cDNA) was synthesised. DNA encoding RANKL and OPG were sequenced and ovine specific primers were designed to amplify the cDNA by real time RT-PCR to generate products corresponding to mRNA encoding RANKL and OPG. The results were normalised to 18S RNA.

Results Total OPG expression (in trabecular bone) from the PF region was over two fold higher than the LS (P< 0.0001). The relative expression of OPG in the both LS and PF regions were significantly higher in the treated animals (steroid & oophorectomy) compared to controls (p< 0.05). The relative expression of RANK-L in the PF was significantly higher than in the LS (P< 0.0001). However, the relative RANK-L expression in the treated animals was not significantly different from the control animals in either region. The ratio of RANK-L:OPG in the PF and the LS was not significantly different but it was significantly reduced in the osteopaenic animals.

Discussion Based on this gene expression study and previous histomorphological data, it appears that trabecular bone loss is not due to increased osteoclastic activity but may rather due to lack of osteoblastic activity and function. Higher expression of OPG and RANK-L and greater bone loss compared to LS suggest that the rate of bone turnover is greater in the PF. Further investigation of the molecular pathways of bone loss in this animal model will increase its utility for osteoporosis research.

Introduction Infection is a risk following surgical procedures that violate the disc. Bacteria, often from normal skin flora, can be introduced into the disc space causing inflammation with destruction of the adjacent vertebrae. The incidence of iatrogenic discitis is thought to be reduced with the use of prophylactic antibiotics. Although there is substantial evidence from animal studies that prophylactic antibiotics reduce the incidence of discitis, evidence in humans is not strong. The aim of this study was to determine if levels of cephazolin in human disc and serum reached the stated minimum inhibitory concentration (MIC > 1 mg/L) against Staphylococcus aureus following intravenous administration.

Methods Thirty patients (15 female and 15 male) with a mean age 42 years, (range 21–63 years) received 1g cephazolin prior to one or two level lumbar spinal fusion surgery. Venous blood was collected prior to intravenous administration of cephazolin and again at the time of disc removal to measure its concentration in serum and disc tissue.

Results The interval between cephazolin administration and tissue sampling ranged from 7 to 137 minutes. Cephazolin concentration in the serum (31.1 – 148 mg/L) was greater than in the disc (0 – 9.5 mg/L). The concentration of cephazolin peaked in the serum at 7 minutes and in the disc between 37 and 53 minutes. Over 70% of the disc samples had detectable levels of cephazolin at the time the disc was removed, although only half had cephazolin levels above 1 mg/L.

Discussion For a given time period the antibiotic concentration in the disc varies between patients. Only one measurement was taken from the disc using the method described when in fact if multiple measurements were taken over time there may be an antibiotic concentration peak in the disc followed by a steady decline. Variability between peak antibiotic concentration in the disc may be attributed to patient size, weight, gender, co-morbidities (diabetes, rheumatoid arthritis) and degree of disc degeneration. However the most critical variable for all patients is timing of administration of antibiotic to achieve optimal concentration in the disc when it is at greatest risk of inadvertent inoculation.

Introduction Bone loss and increased bone fragility may occur following long-term steroid therapy resulting in painful vertebral fractures. To fully understand the mechanisms underlying steroid-induced osteoporosis and to test potential spinal implants a large animal model is needed. The sheep is appropriate since bone remodelling is similar to humans and the vertebral body dimensions permit easy surgical manipulation. This study was undertaken to fully characterise the sheep as a model of steroid-induced vertebral osteoporosis and to evaluate its suitability for testing balloon kyphoplasty.

Methods Osteoporosis was induced in ten lactating ewes (mean age 8 years) by ovariectomy, weekly injection of 54mg dexamethasone (Dexafort, Intervet, Australia) and a diet containing 0.2% calcium for up to 6 months. Baseline and endpoint iliac crest biopsies were taken to measure static histomorphometric indices of bone formation and resorption. Control animals were not ovariectomised and not treated with steroids. Dual energy X-ray absorptiometry (DXA, Hologic QDR 1000+, USA) was used to monitor bone mineral density (BMD) in the lumbar spine (L2-L5) after 0, 3 and 6 months of steroid treatment. At each time interval sheep were killed by barbiturate injection and the entire lumbar spine (L1-L6) was processed for histology, quantitative histomorphometry, mechanical testing, micro-CT (computed tomography) and ex-vivo trials to inflate kyphoplasty balloon tamps.

Results After six months of treatment, BMD in the lumbar spine decreased by 29.5% from baseline. Trabecular bone volume of L2, L3 and L4 vertebrae (pooled) decreased by 31.4% (p< 0.05) and trabecular thickness decreased by 33.9%. Cortical bone thickness decreased by 43.9% (p< 0.05). The average load at which L1 yielded decreased by 67.4%. Static measurements of bone formation decreased by 68.3% and bone resorption increased 10 fold. Kyphoplasty balloon tamps were successfully inflated ex-vivo in vertebral bodies from treated animals.

Discussion Using DXA, cancellous bone histomorphometry and mechanical testing, this study has demonstrated significant trabecular and cortical bone loss in the sheep lumbar spine up to six months after ovariectomy and continuous steroid treatment. These changes are the result of increased resorption and decreased formation of bone. The successful inflation of bone tamps in osteoporotic vertebrae ex-vivo is a prerequisite for in-vivo studies to assess the safety and efficacy of the kyphoplasty procedure.

Introduction Magnetic resonance imaging (MRI) is a valid investigation for the diagnosis of intervertebral disc disease, including infection, but it is expensive and difficult to access conveniently for research studies on live animals under anaesthesia. The aim of this study was to compare the MRI signal changes in spines from living and recently deceased sheep.

Methods MRI was conducted on the lumbar spines of six adult sheep from research studies investigating bacterial discitis, chemical discitis and disc degeneration resulting from annular incision. The sheep was anaesthetised and the lumbar spines were imaged with a Siemens Magneton Vision MRI (Numaris VB33G software) using T1 SGE, T1 FSE, STIR and T2 FSE sequences. The sheep were killed with an intravenous overdose of barbiturate and identical images were obtained commencing five minutes post mortem.

Results For each of the disease processes under consideration the MRI contrast relationships were maintained between all of the anatomical structures of interest. The post-mortem images provided improved clarity, particularly in the STIR and T2-weighted sequences, due to the absence of pulsation from the CSF and aorta, as well as the absence of respiratory artefact.

Discussion The MRI appearances of spinal tissues following death correlate well with those in the live animal confirming the validity of this method of investigation. This is particularly relevant for optimisation of a busy clinical resource for research purposes.

Objective: To investigate the effects of intra-discal electro-thermal therapy (IDET) on an experimentally induced posterolateral annular inter vertebral disclesion in sheep.

Summary of Background Data: IDET is being used increasingly as a minimally-invasive treatment for chronic discogenic low back pain, with success reported in up to 70% of cases. The mechanism of action however is poorly understood. Proposed mechanisms include the contraction of collagen and the coagulation of annular nociceptors. An ovine model was used to assess the innervation of peripheral posterolateral annular lesions and the potential for IDET to denervate this region.

Methods: Posterolateral annular incisions were made in 36 lumbar discs of 18 sheep. After twelve weeks the sheep underwent IDET at one level and a sham treatment at the other level. IDET was performed using a modified intradiscal catheter (SpineCATH™, Oratec Interventions Inc., Menlo Park, CA). Temperatures were recorded in the nucleus (T_N) and the posterior annulus (T_Pa). The spines were harvested at intervals up to eighteen months. Histological sections of the discs were stained with haematoxylin and eosin and an antibody to the general neuronal marker PGP 9.5.

Results: The target temperature of 90°C at the catheter was tip was reached in all cases. The mean maximum T_Pa was 63.6°C and the mean maximum T_N 67.8°C. Vascular granulation tissue consistent with a healing response was observed in the region of the posterior annulus tear of all incised discs from 12 weeks. PGP 9.5 positive nerve fibres were clearly identified in the adjacent periannular tissue, but were scarce within the outer few lamellae of the annulus. There were no fewer nerve fibres identified in those specimens that had undergone IDET. From six weeks after IDET there was evidence of thermal necrosis in the inner annulus, sparing the periphery of the disc.

Conclusions: IDET delivered at 90°C in the sheep consistently heats the posterior annulus and the nucleus to a temperature range associated with coagulation of nociceptors and collagen contraction. Thermal necrosis was observed within the inner annulus from six weeks after IDET. In this model IDET did not produce denervation of the experimentally induced posterior annular lesion.

Objectives: Posterolateral annular lesions were experimentally induced and allowed to mature for 12 weeks in the intervertebral discs of sheep. IDET was performed in an attempt to denervate and repair the annular lesion. The histological and immunohistochemical effects of IDET were studied.

Summary of Background Data: IDET continues to be used as a minimally-invasive treatment for chronic discogenic low back pain, with success rates reported in up to 70% of cases. The mechanism of action by which IDET exerts its effect is poorly understood. Proposed mechanisms include the contraction of collagen and the coagulation of annular nociceptors. An ovine model was used firstly to induce a posterolateral annular lesion, secondly to assess the innervation of such a lesion, and thirdly to assess the effect of IDET on this innervation.

Methods: Posterolateral annular incisions were made in 40 lumbar discs of 20 sheep. Twelve weeks were allowed for each annular lesion to mature. IDET was then performed in the disc with the posterolateral annular tear and in another control level. IDET was performed using a modified intradiscal catheter. Temperatures were recorded in the nucleus (T_N) and the posterior annulus (T_PA). The spines were harvested at predetermined intervals up to eighteen months. Histological sections of the discs were graded for disc morphology to assess degeneration and immunohistochemical staining to assess potential denervation.

Results: Vascular granulation tissue consistent with a healing response was observed in the posterior annular tear of all incised discs from 12 weeks. PGP 9.5 positive nerve fibres were clearly identified in the adjacent periannular tissue and the outer few lamellae of the posterior annulus. During the IDET procedure the mean maximaximum T_PA was 63.6°C and the mean maximaximum T_N was 67.8°C. At sacrifice the number of nerve fibres identified in the posterior annular tear was the same for those specimens that had undergone IDET and those that had not. From six weeks after IDET there was evidence of thermal necrosis in the inner annulus and adjacent nucleus but sparing the periphery of the disc.

Conclusions: IDET delivered at 90°C in the sheep consistently heated the posterior annulus and the nucleus to a temperature normally associated with coagulation of nociceptors and collagen contraction. IDET did not denervate the posterior annular lesion. Thermal necrosis was observed within the inner annulus and adjacent nucleus from six weeks after IDET. The reported benefits from IDET appear to be related to factors other than denervation and repair.

Introduction The annulus fibrosus of the intervertebral disc is composed of a series of concentric collagenous lamellae that constrain the highly pressurised fluid of the nucleus pulposus. With advancing age and even after physical injury in youth the disc almost invariably becomes progressively degenerate due to the combined effects of dehydration of the nucleus and disruption of the annulus. There is conjecture however, about which of the two compartments shows degenerate changes first.

Methods A histological and biochemical review is presented, based on a review of the literature and work carried out in our laboratories.

Results Three distinct types of annulus tears are seen histologically. Rim tears are formed by detachment of the peripheral annular fibres from the vertebral rim. Autopsy studies show that these lesions are rare in subjects younger than 30 years but the incidence increases significantly with advancing age. Although granulation tissue grows into the outer layers of the annulus in a normal healing response, these lesions frequently extend deeper into the disc. Radiating tears course radially across several lamellae, most often extending from the vertebral rim across the nucleus to involve the posterior side of the disc. These lesions are seen mostly between the ages of 30 and 50 years. Concentric tears are characterised by separation of adjacent lamellae and may appear as early as the second decade of life. In advanced degeneration nuclear changes may be seen with any combination of annular lesions. Biochemical changes, including dehydration and reduced proteoglycan content of the nucleus, parallel the morphologic changes to the disc. Altered biochemistry is further reflected by MRI studies in which some scanning sequences can detect even modest loss of fluid from the nucleus as early as three months after experimental annular incision. Annular lesions also compromise the biomechanical properties of the disc. While internal fixation that aims to immobilise the injured disc may promote some recovery of the mechanical integrity, degeneration nevertheless advances in the long term. Attempts to seal peripheral annular defects in experimental studies using biocompatible glue have also failed to promote healing and to prevent progression of even minor structural defects.

Conclusions Technological developments such as gene transfer into disc cells and direct implantation of either stem cells or more mature cells are emerging as potential candidates for the treatment of disc degeneration.

Introduction: Disc degeneration is consistent with advancing age and in many cases is associated with back pain and restricted mobility. The traditional surgical treatment for chronic back pain has been spinal fusion to immobilize the painful level. Long-term studies, however, suggest that fusion actually promotes degeneration at adjacent levels. One of the hallmarks of disc degeneration is aggregation of chondrocytes in the nucleus of chondrones, and more recently apoptosis has been implicated as a factor controlling the longevity of the cells. Recent research suggests that it may be possible to restore normal function to degenerate discs by introducing a fresh population of cells. This study investigated the potential for autologous costal chondrocyte implantation to prevent lumbar disc degeneration after annular injury in the sheep.

Methods: the lumbar spines of eight adult sheep were exposed. In four animals, full thickness annular incisions were made in three alternate discs. No annular incisions were made in the other four sheep. A minimum of 500 mg of cartilaginous tissue was harvested from the twelfth rib of all animals. Tissue was cultured in vitro and the chondrocytes were labelled with a fluorescent marker for retrospective identification. After six weeks the chondrocytes were injected into the lower two alternate discs of all animals, leaving the uppermost discs and those untouched as internal controls. The animals were killed at intervals from three to twenty-four weeks and MRI, plain x-ray, histology and immunocytochemistry were evaluated.

Results: MRI at twelve and twenty-four weeks showed apparent preservation of all incised discs that had been transplanted with autologous chondrocytes. Histology revealed clusters of viable chondrocytes of normal appearance within the nucleus. These cells stained positive for the fluorescent label. The same cells and the surrounding matrix were also positive for collagen type II. Serial X-ray measurements suggested that progressive disc degeneration was arrested in the discs that received autologous costal chondrocytes.

Discussion: This pilot study showed evidence that cultured autologous costal chondrocytes remained viable and produced extracellular matrix following transplantation into normal and degenerate discs. In contrast to other studies that have used mesenchymal stem cells or chondrocytes harvested directly from discs, this study demonstrated success with cells from a source other than the disc. Costal cartilage is a convenient source of cells for transplantation and this technique warrants further investigation as a potential treatment for degenerative disc disease.

Introduction: Although prophylactic antibiotic administration is common in spinal surgery, the choice of drug, dose, and timing of administration often varies. Little is known about the activity of antibiotics in the spine and indeed if they are distributed throughout the disc and if the time intervals are optimal. Because infections that produce iatrogenic discitis generally arise within the disc, the antibiotic concentration of the disc is more relevant than serum concentrations. The aims of the study were to determine if a 2g dose of cephazolin was effective at preventing discitis over a four-hour period in immature ovine discs that were both non-degenerate and degenerate; and also to determine the concentration of cephazolin in serum and disc tissue.

Methods: In 10 Merino wethers aged 12 weeks, three lumbar discs were “degenerated” by incising the posterolateral annulus with a scalpel blade and using ronguers, removing the bulk of the nucleus pulposus. After 12 weeks nine animals were anaesthetised and given a 2g dose of cephazolin (David Bull Laboratories, Australia) at predetermined time intervals over a four-hour period. The antibiotic was chosen for effectiveness against Staphylococcus aureus a common discitis-causing organism. One sheep (control) did not receive any antibiotics to follow the natural progression of infection. All animals had discography with radiographic contrast that contained S. aureus at two incised levels and at two non-incised levels. Lateral radiographs of the lumbar spine were taken at two, six and 12 weeks to monitor the bony changes. At 12 weeks all sheep were given a 2g intravenous dose of cephazolin at time intervals before being killed. The spines were removed and prepared for light microscopy to assess pathology of the discs and for biochemical analysis of antibiotic concentration. Success of treatment was judged using histologic and radiographic features.

Results: The control sheep that did not receive any antibiotics developed discitis at four levels. Histology at 12 weeks confirmed discitis in 10/36 “prophylactic discs”. Of these “prophylactic discs” 7/10 had previously been “degenerated”. Discitis only developed in immature discs that were administered cephazolin two hours prior to inoculation. When antibiotic was administered after inoculation discitis was prevented. Biochemistry results confirmed that antibiotic diffused throughout the disc but was concentrated in the annulus more than the nucleus. Antibiotic levels in the disc peaked at 15 minutes (annulus mean concentration 15.5 mg/L, nucleus mean concentration 3.2 mg/L). Serum levels at 15 minutes were up to 50 times greater at this time (serum mean concentration 178 mg/L).

Discussion: The discs that were “degenerate” had a higher incidence of discitis compared to “non-degenerate” discs. However the concentration of antibiotic in degenerate discs was not significantly different than in non-degenerate discs. A 2 gram dose of cephazolin is reasonably effective (approx 70% success rate) at preventing discitis over a four-hour period.

Introduction: Vertebral compression fractures are common in osteoporosis, resulting in spinal deformities, severe back pain and decreased mobility. Vertebroplasty and kyphoplasty procedures aim to restore the integrity of the deformed vertebral body by injection of biocompatible cement. To date, there have been no long-term studies of the bone-cement interaction in this setting. A reliable large animal model of vertebral osteoporosis would be useful to fully characterise the disease process, to assess potential treatment regimens and to investigate the biocompatibility of bone cements used in kyphoplasty and vertebroplasty. The aim of this pilot study was to develop such a model with ovariectomy, low calcium diet and continuous steroid treatment.

Methods: To induce osteoporosis, ten lactating ewes (mean age 8 years) were ovariectomised, injected weekly with 9 mg dexamethasone (Dexafort, Intervet, Australia) and fed low calcium diet. Weekly serum samples were taken to quantify generalised bone resorption (Type 1 collagen C-telopeptide [CTX], ‚-Cross Laps assay, Roche Diagnostics, Australia). Dual-energy X-ray absorptiometry (DEXA, Hologic QDR 1000+, USA) was used to monitor bone mineral density (BMD) in the lumbar spine (L3-L6) after 0, 2, 4, 6 and 9 months of treatment. At each time interval two sheep were killed by barbiturate injection. The entire lumbar spine (L1-L6) was processed for histology, quantitative histomorphometry, mechanical testing and micro-CT (computed tomography).

Results: CTX levels increased rapidly after two months (p< 0.05). Baseline BMD in the lumbar spine (0.87±0.06 g/cm2) decreased by 16.9±3.8% or 2.72 standard deviations (p< 0.001) after nine months of treatment. Structural parameters of cancellous bone also showed osteoporotic change. Trabecular bone volume of L2, L3 and L6 vertebrae (pooled) progressively decreased from 24.9±1.2% at two months to 16.5±0.47% at nine months (p< 0.05). Trabecular thickness decreased from 0.14±0.01mm to 0.09±0.01mm, (p< 0.05) and trabecular spacing increased from 0.42±0.03mm to 0.47±0.02mm in the same period. The compressive load at which the L1 vertebrae failed decreased by 39.4% after 9 months.

Discussion: This pilot study has demonstrated by DEXA, cancellous bone histomorphometry and mechanical testing, significant bone loss in the sheep lumbar spine up to nine months after ovariectomy and continuous steroid treatment. Assuming that the baseline BMD is representative of mature sheep, the changes in the lumbar spine could be interpreted as osteoporotic. Vertebral bone loss did not reach levels that would result in fracture. However, further work is underway using higher steroid doses to accelerate bone loss. This experimental model will be used to assess aspects of osteoporosis in general and vertebral augmentation procedures in particular.

Introduction: Infection can occur after any spinal procedure that involves entry into the disc and although it is not common, the potential consequences are serious. Treatment usually requires identification of the bacteria followed by a course of antibiotics. This treatment remains controversial since it is not clear whether antibiotics actually penetrate the disc and if so, whether they are effective, or even if the outcome would be the same without antibiotics.

For an antibiotic to be effective against the infecting organism it must diffuse through the disc matrix. Blood vessels that surround the disc facilitate the diffusion process, but with age this vascularity decreases and may impede diffusion.

The aims of the pilot study were to assess the effectiveness of antibiotic in treating infection in both normal and degenerate sheep discs and to measure the concentration of antibiotic in non-operated discs at varying ages.

Methods: In each of six Merino wethers aged 12 weeks (n=3) and 24 months (n=3), two lumbar discs were “degenerated” by incising the posterolateral annulus with a scalpel blade. After four weeks all animals had discography with radiographic contrast that contained Staphylococcus aureus at the incised levels and at two non-incised levels. Seven days after infection four animals began IV antibiotic treatment with cephazolin sodium (David Bull Laboratories, Australia) for 21 days at a dose of 50mg/kg/day. The antibiotic was chosen for effectiveness against S. aureus. One control animal from each age group did not receive any antibiotics, to follow the natural progression of infection. Lateral radiographs of the lumbar spine were taken at two, six and 12 weeks. At 12 weeks all sheep were given a single intravenous dose of cephazolin sodium as either a 1, 2 or 3g dose. The sheep were then killed after 30 minutes. The spines were removed and prepared for light microscopy to assess pathology of the discs and for biochemical analysis of antibiotic concentration. Success of treatment was judged using histologic and radiographic features.

Results: Discitis was evident radilogically as early as two weeks after inoculation in all animals. Histology at 12 weeks confirmed discitis in all discs regardless of treatment. Biochemistry results confirmed that antibiotic diffused throughout the disc but was more concentrated in the annulus than the nucleus. At all doses disc concentration of antibiotic was higher in lambs than sheep.

Discussion: Treatment with cephazolin sodium at a dose of 50mg/kg/day for 21 days administered from seven days after inoculation, did not prevent discitis. This does not appear to be due to inability of antibiotic diffusion into the disc.

Introduction: Although it is well recognized that the outer annulus is innervated, the relative densities of innervation of different regions of the disc have not been quantitated. We present here the first comparative analysis of the innervation of the innervation of different regions of the lumbar intervertebral disc.

Methods: A sheep model was used allowing evaluation of the whole motion segment. Four sheep spines were used. One was processed for PGP 9.5 immunoflourescence and three were processed for PGP 9.5 immunoperoxidase histochemistry. Serial sagittal sections were obtained and a count was made of the densities of innervation of different regions of the endplate and annulus. These were compared to identify which areas of the disc and endplate are most innervated.

Results: The endplate innervation is concentrated centrally adjoining the nucleus. The mean density of innervation of the central endplate was 0.44 (SEM 0.07) nerves/ mm² while the mean density of the peripheral endplate was 0.10 (SEM 0.03) nerves/ mm² (p= 0.0001). There was no significant difference between the overall endplate and annulus innervation densities 0.52 (SEM 0.1) v 0.37 (SEM 0.07) p=0.2. But the peri-annular connective tissue, external to the outer annulus contained the densest innervation of any region in the motion segment 1.05 (SEM 0.16).

Discussion: The lumbar intervertebral disc has a meagre innervation. This is concentrated in the peri-annular connective tissue and the central endplate. While receptor threshold is more closely related to nociceptive function than innervation density, these findings have important implications for any treatment of discogenic pain.

Introduction: Intradiscal electrothermal therapy (IDET) is being used increasingly as a minimally-invasive treatment for chronic discogenic low back pain, with success reported in up to 70% of cases. The mechanism of action however is poorly understood. Proposed mechanisms include the contraction of collagen and the coagulation of annular nociceptors. An ovine model was used to assess the innervation of peripheral posterolateral annular lesions and the potential for IDET to denervate this region.

Methods: Posterolateral annular incisions were made in 36 lumbar discs of 18 sheep. After twelve weeks the sheep underwent IDET at one level and a sham treatment at the other level. IDET was performed using a modified intradiscal catheter (SpineCATH™, Oratec Interventions Inc., Menlo Park, CA). Temperatures were recorded in the nucleus and the posterior annulus. The spines were harvested at intervals of up to eighteen months. Histological sections of the discs were stained with haematoxylin and eosin and an antibody to the general neuronal marker PGP 9.5.

Results: The target temperature of 90°C at the catheter tip was reached in all cases. The mean maximum T_Pa was 63.6°C and the mean maximum TN was 67.8°C. Vascular granulation tissue consistent with a healing response was observed in the region of the posterior annulus tear of all incised discs from 12 weeks. PGP 9.5 positive nerve fibres were clearly identified in the adjacent periannular tissue, but were scarce within the outer few lamellae of the annulus. There were no fewer nerve fibres identified in those specimens that had undergone IDET. From six weeks after IDET there was evidence of thermal necrosis in the inner annulus, sparing the periphery of the disc.

Discussion: IDET delivered at 90°C in the sheep consistently heats the posterior annulus and the nucleus to a temperature associated with coagulation of nociceptors and collagen contraction. Thermal necrosis was observed within the inner annulus from six weeks after IDET. In this model IDET did not appear to produce denervation of the posterior annular lesion.