Osteogenic differentiation of bone marrow stromal cells is hindered by the presence of intervertebral disc cells.

BACKGROUND: Clinical observations indicate that the presence of nucleus pulposus (NP) tissue during spinal fusion hinders the rate of disc ossification. While the underlying mechanism remains unknown, this observation could be due to incomplete removal of NP cells (NPCs) that secrete factors preventing disc calcification, such as bone morphogenetic protein (BMP) antagonists including noggin and members of the DAN (differential screening selected gene aberrative in neuroblastoma) family. METHODS: Monolayer human bone marrow-derived mesenchymal stem cells (MSCs) were cocultured withNPCs and annulus fibrosus cells (AFCs) embedded in alginate for 21 days. At the end of coculture, MSCs were stained for mineral deposition by alizarin red, and relative expression of bone-related genes [Runt-related transcription factor 2, (RUNX2), Osteopontin (OPN), and Alkaline phosphatase (ALP)] and ALP activity were analyzed. Relative expression of three BMP antagonists, chordin (CHRD), gremlin (GREM1), and noggin (NOG), was determined in primary human NPCs and AFCs. These cells were also stained for Gremlin and Noggin by immunocytochemistry. RESULTS: Alizarin red staining showed that MSC osteogenesis in monolayer cultures was inhibited by coculture with NPCs or AFCs. ALP activity and RT-PCR analyses confirmed these results and demonstrated inhibition of osteogenesis of MSC in the presence of disc cells. NOG was significantly up-regulated in MSCs after coculture. Relative gene expression of intervertebral disc (IVD) cells showed higher expression of GREM1 in NPCs than in AFCs. CONCLUSIONS: We show that primary IVD cells inhibit osteogenesis of MSCs. BMP inhibitors NOG, GREM1 and CHRD were expressed in IVD cells. GREM1 appears to be differentially expressed in NPCs and AFCs. Our results have implications for the design and development of treatments for non-union in spinal fusion.

Vertebral body stenting / stentoplasty.

Osteoporotic vertebral fractures are frequent. Although the majority of fractures follow a benign course, there are certain fracture types which result in severe spinal deformity and / or are associated with neurological complications. These patients should be detected early and undergo surgical treatment. Vertebroplasty remains an important and effective treatment option for acute painful vertebral fractures showing progressive collapse. By this procedure the fracture can be stabilised, the pain is controlled and the progression of height loss is also halted. If a vertebral body shows a higher degree of collapse and kyphotic deformity or even some posterior wall involvement, the stentoplasty procedure (further evolution of kyphoplasty) allows height resotartion by the stent and the stabilisation of the vertebral body by cement.

Cervical spine injury in the young child.

This grand rounds is about the clinical and radiological presentation, treatment and outcome of pediatric cervical spine injury. A 15-month-old girl suffers from a motor vehicle accident and is intubated on-site because of progressive agitation. Whole body trauma CT was read as normal. When sedation was discontinued after 24 h she was found to be tetraplegic below C6 level. MRI shows a total disruption between C6 and C7 that in hindsight was also visible on the initial trauma CT. She was treated surgically by an anterior and posterior reconstruction and was post-operatively treated with a halo vest. Clearing the cervical spine in young children is deceptively difficult. Meticulous review and interpretation of conventional radiographs and CT are important yet MRI should be considered in uncertain cases. Severe ligamentous injury without concomitant bony injury occurs more frequently than in older children and adults, with sometimes devastating consequences.

Expert's comment concerning grand rounds case entitled "a traumatic, high-energy and unstable fracture of the C5 vertebra managed with kyphoplasty: a previously unreported case" (by Nasir A. Quraishi and Sherief Elsayed).

Case presentation of a cervical spine fracture treated by kyphoplasty. The review of the case, especially the careful re-assessment of the x-rays reveals that the treatment goals have not been achieved (motion preservation, functional restoration of the vertebral body). The case report is an excellent example of our biased viewpoint in assessing our efforts.

Cardiovascular changes after PMMA vertebroplasty in sheep: the effect of bone marrow removal using pulsed jet-lavage.

Clinically, the displacement of intravertebral fat into the circulation during vertebroplasty is reported to lead to problems in elderly patients and can represent a serious complication, especially when multiple levels have to be treated. An in vitro study has shown the feasibility of removing intravertebral fat by pulsed jet-lavage prior to vertebroplasty, potentially reducing the embolization of bone marrow fat from the vertebral bodies and alleviating the cardiovascular changes elicited by pulmonary fat embolism. In this in vivo study, percutaneous vertebroplasty using polymethylmethacrylate (PMMA) was performed in three lumbar vertebrae of 11 sheep. In six sheep (lavage group), pulsed jet-lavage was performed prior to injection of PMMA compared to the control group of five sheep receiving only PMMA vertebroplasty. Invasive recording of blood pressures was performed continuously until 60 min after the last injection. Cardiac output and arterial blood gas parameters were measured at selected time points. Post mortem, the injected cement volume was measured using CT and lung biopsies were processed for assessment of intravascular fat. Pulsed jet-lavage was feasible in the in vivo setting. In the control group, the injection of PMMA resulted in pulmonary fat embolism and a sudden and significant increase in mean pulmonary arterial pressure. Pulsed jet-lavage prevented any cardiovascular changes and significantly reduced the severity of bone marrow fat embolization. Even though significantly more cement had been injected into the lavaged vertebral bodies, significantly fewer intravascular fat emboli were identified in the lung tissue. Pulsed jet-lavage prevented the cardiovascular complications after PMMA vertebroplasty in sheep and alleviated the severity of pulmonary fat embolism.

Hierarchical clustering of flow cytometry data for the study of conventional central chondrosarcoma.

We have investigated the use of hierarchical clustering of flow cytometry data to classify samples of conventional central chondrosarcoma, a malignant cartilage forming tumor of uncertain cellular origin, according to similarities with surface marker profiles of several known cell types. Human primary chondrosarcoma cells, articular chondrocytes, mesenchymal stem cells, fibroblasts, and a panel of tumor cell lines from chondrocytic or epithelial origin were clustered based on the expression profile of eleven surface markers. For clustering, eight hierarchical clustering algorithms, three distance metrics, as well as several approaches for data preprocessing, including multivariate outlier detection, logarithmic transformation, and z-score normalization, were systematically evaluated. By selecting clustering approaches shown to give reproducible results for cluster recovery of known cell types, primary conventional central chondrosacoma cells could be grouped in two main clusters with distinctive marker expression signatures: one group clustering together with mesenchymal stem cells (CD49b-high/CD10-low/CD221-high) and a second group clustering close to fibroblasts (CD49b-low/CD10-high/CD221-low). Hierarchical clustering also revealed substantial differences between primary conventional central chondrosarcoma cells and established chondrosarcoma cell lines, with the latter not only segregating apart from primary tumor cells and normal tissue cells, but clustering together with cell lines from epithelial lineage. Our study provides a foundation for the use of hierarchical clustering applied to flow cytometry data as a powerful tool to classify samples according to marker expression patterns, which could lead to uncover new cancer subtypes.

DensiProbe Spine: a novel instrument for intraoperative measurement of bone density in transpedicular screw fixation.

STUDY DESIGN: Cadaver study. OBJECTIVE: To determine bone strength in vertebrae by measuring peak breakaway torque or indentation force using custom-made pedicle probes. SUMMARY OF BACKGROUND DATA: Screw performance in dorsal spinal instrumentation is dependent on bone quality of the vertebral body. To date no intraoperative measuring device to validate bone strength is available. Destructive testing may predict bone strength in transpedicular instrumentations in osteoporotic vertebrae. Insertional torque measurements showed varying results. METHODS: Ten human cadaveric vertebrae were evaluated for bone mineral density (BMD) measurements by quantitative computed tomography. Peak torque and indentation force of custom-made probes as a measure for mechanical bone strength were assessed via a transpedicular approach. The results were correlated to regional BMD and to biomechanical load testing after pedicle screw implementation. RESULTS: Both methods generated a positive correlation to failure load of the respective vertebrae. The correlation of peak breakaway torque to failure load was r = 0.959 (P = 0.003), therewith distinctly higher than the correlation of indentation force to failure load, which was r = 0.690 (P = 0.040). In predicting regional BMD, measurement of peak torque also performed better than that of indentation force (r = 0.897 [P = 0.002] vs. r = 0.777 [P = 0.017]). CONCLUSION: Transpedicular measurement of peak breakaway torque is technically feasible and predicts reliable local bone strength and implant failure for dorsal spinal instrumentations in this experimental setting.

The effect of pulsed jet lavage in vertebroplasty on injection forces of PMMA bone cement: an animal study.

Percutaneous vertebroplasty, comprising of the injection of polymethylmethacrylate (PMMA) into vertebral bodies, is an efficient procedure to stabilize osteoporotic compression fractures as well as other weakening lesions. Besides fat embolism, cement leakage is considered to be one of the major and most severe complications during percutaneous vertebroplasty. The viscosity of the PMMA during injection plays a key role in this context. It was shown in vitro that the best way to lower the risk of cement leakage is to inject the cement at higher viscosity, which is requires high injection forces. Injection forces can be reduced by applying a newly developed lavage technique as it was shown in vitro using human cadaver vertebrae. The purpose of this study was to prove the in vitro results in an in vivo model. The investigation was incorporated in an animal study that was performed to evaluate the cardiovascular reaction on cement augmentation using the lavage technique. Injection forces were measured with instrumentation for 1 cc syringes, additionally acquiring plunger displacement. Averaged injection forces measured, ranged from 12 to 130 N and from 28 to 140 N for the lavage group and the control group, respectively. Normalized injection forces (by viscosity and injection speed) showed a trend to be lower for the lavage group in comparison to the control group (P = 0.073). In conclusion, the clinical relevance on the investigated lavage technique concerning lowering injection forces was only shown by trend in the performed animal study. However, it might well be that the effect is more pronounced for osteoporotic vertebral bodies.

The effect of pulsed jet lavage in vertebroplasty on injection forces of polymethylmethacrylate bone cement, material distribution, and potential fat embolism: a cadaver study.

STUDY DESIGN: In vitro testing of vertebroplasty techniques including pulsed jet-lavage for fat and marrow removal in human cadaveric lumbar and thoracic vertebrae. OBJECTIVE: To develop jet-lavage techniques for vertebroplasty and investigate their effect on cement distribution, injection forces, and fat embolism. SUMMARY OF BACKGROUND DATA: The main complications of cement vertebroplasty are cement leakage and pulmonary fat embolism, which can have fatal consequences and are difficult to prevent reliably by current vertebroplasty techniques. METHODS: Twenty-four vertebrae (Th8-L04) from 5 osteoporotic cadaver spines were grouped in triplets depending on bone mineral density (BMD). Before polymethylmethacrylate (PMMA) vertebroplasty, a pulsatile jet-lavage for removal of intertrabecular fat and bone marrow was performed in 2 groups with 8 specimens each, performing radial and axial irrigation from the biopsy needles. One hundred mL of Ringer solution were injected through 1 pedicle and regained by low vacuum via the contralateral pedicle. Eight control vertebrae were not irrigated. All specimens underwent standardized PMMA cement augmentation injecting 20% of the vertebral volume. Injection forces, cement distribution, and extravasations were quantified. RESULTS: All irrigation solution could be retrieved with the vacuum applied. A Kruskal-Wallis test revealed significantly higher injection forces of the control group as compared with the irrigated groups (P = 0.021). Dilatation of the syringe at forces above 300 N occurred in 75% of the untreated compared with 12.5% of the lavaged specimens. CT distribution analysis showed more homogenous cement distribution of the cement and significantly less extravasation in the irrigated specimens. CONCLUSION: The developed lavage technique for vertebroplasty showed to be feasible and reproducible. The reduction of injection forces would allow the use of more viscous PMMA cement lowering the risk for cement embolization and results in a safer procedure. The wash-out of bone marrow and the possible reduction of pulmonary fat embolism have to be verified with in vivo models.

Development of a computer-assisted high-pressure Injection device for vertebroplasty.

A novel computer-assisted injection device for the delivery of highly viscous bone cements in vertebroplasty is presented. It addresses the shortcomings of manual injection systems ranging from low-pressure and poor level of control to device failure. The presented instrument is capable of generating a maximum pressure of 5000 kPa in traditional 6-ml syringes and provides an advanced control interface for precise cement delivery from outside radiation fields emitted by intraoperative imaging systems. The integrated real-time monitoring of injection parameters, such as flow-rate, volume, pressure, and viscosity, simplifies consistent documentation of interventions and establishes a basis for the identification of safe injection protocols on the longer term. Control algorithms prevent device failure due to overloading and provide means to immediately stop cement flow to avoid leakage into adjacent tissues.

Lordoplasty: report on early results with a new technique for the treatment of vertebral compression fractures to restore the lordosis.

Cement augmentation using PMMA cement is known as an efficient treatment for osteoporotic vertebral compression fractures with a rapid release of pain in most patients and prevention of an ongoing kyphotic deformity of the vertebrae treated. However, after a vertebroplasty there is no chance to restore vertebral height. Using the technique of kyphoplasty a certain restoration of vertebral body height can be achieved. But there is a limitation of recovery due to loss of correction when deflating the kyphoplastic ballon and before injecting the cement. In addition, the instruments used are quite expensive. Lordoplasty is another technique to restore kyphosis by indirect fracture reduction as it is used with an internal fixateur. The fractured and the adjacent vertebrae are instrumented with bone cannulas bipediculary and the adjacent vertebrae are augmentated with cement. After curing of the cement the fractured vertebra is reduced by applying a lordotic moment via the cannulas. While maintaining the pretension the fractured vertebra is reinforced. We performed a prospective trial of 26 patients with a lordoplastic procedure. There was a pain relief of about 87% and a significant decrease in VAS value from 7.3 to 1.9. Due to lordoplasty there was a significant and permanent correction in vertebral and segmental kyphotic angle about 15.2 degrees and 10.0 degrees , respectively and also a significant restoration in anterior and mid vertebral height. Lordoplasty is a minimal invasive technique to restore vertebral body height. An immediate relief of pain is achieved in most patients. The procedure is safe and cost effective.

Radiation exposure to the surgeon during fluoroscopically assisted percutaneous vertebroplasty: a prospective study.

STUDY DESIGN: A prospective case control study design was conducted. OBJECTIVES: The purpose of the current study was to determine the intraoperative radiation hazard to spine surgeons by occupational radiation exposure during percutaneous vertebroplasty and possible consequences with respect to radiation protection. SUMMARY OF BACKGROUND DATA: The development of minimally invasive surgery techniques has led to an increasing number of fluoroscopically guided procedures being done percutaneously such as vertebroplasty, which is the percutaneous cement augmentation of vertebral bodies. METHODS: Three months of occupational dose data for two spine surgeons was evaluated measuring the radiation doses to the thyroid gland, the upper extremities, and the eyes during vertebroplasty. RESULTS: The annual risk of developing a fatal cancer of the thyroid is 0.0025%, which means a very small to small risk. The annual morbidity (the risk of developing a cancer including nonfatal ones) is 0.025%, which already means a small to medium risk. The dose for the eye lens was about 8% of the threshold dose to develop a radiation induced cataract (150 mSv); therefore, the risk is very low but not negligible. The doses measured for the skin are 10% of the annual effective dose limit (500 mSv) recommended by the ICRP (International Commission on Radiologic Protection); therefore, the annual risk for developing a fatal skin cancer is very low. CONCLUSION: While performing percutaneous vertebroplasty, the surgeon is exposed to a significant amount of radiation. Proper surgical technique and shielding devices to decrease potentially high morbidity are mandatory. Training in radiation protection should be an integral part of the education for all surgeons using minimally invasive radiologic-guided interventional techniques.

Clinical measurements of cement injection pressure during vertebroplasty.

STUDY DESIGN: Clinical study of injection pressure during vertebroplasty. OBJECTIVES: To investigate the range of injection pressures during conventional vertebroplasty interventions and to study the influence of syringe design and cement polymerization time on injection pressure. SUMMARY OF BACKGROUND DATA: Vertebroplasty is an efficient procedure for the treatment of painful vertebral fractures. However, cement leakage is a potentially serious complication. Although injection pressure has been suggested as a factor for extravasation risk, to date, there are only anecdotal reports of pressure data for cement augmentation in the clinic. METHODS: Using a syringe holder instrumented with load and displacement transducers, injection pressure and volume were recorded in vivo during conventional cement augmentation. Wide (3 mm opening) and normal (1.8 mm opening) syringes were alternated such that each type was evaluated for early (300-500 seconds postmixing) and late (>500 seconds postmixing) cement polymerization time. The influence of syringe type and polymerization time on injection pressure was evaluated using a multifactorial analysis of variance followed by Scheffe post hoc comparison. RESULTS: The maximum peak injection pressure measured was 3215 kPa. The average pressure peaks for normal and wide syringes were 1693 +/- 653 kPa and 1727 +/- 597 kPa, respectively. No statistically significant differences were found between injection pressures with wide and normal syringes. Higher injection pressures were observed later in the polymerization process. CONCLUSIONS: High injection pressures approaching 20 atmospheres are reached during conventional vertebroplasty. Widening the syringe tip diameter did not significantly change injection pressures, whereas elapsed time did. Further research is needed to improve injection equipment and materials for vertebroplasty.

Correlation of radiographic and MRI parameters to morphological and biochemical assessment of intervertebral disc degeneration.

UNLABELLED: Degenerative disc disease (DDD) is a common finding in MRI scans and X-rays. However, their correlation to morphological and biochemical changes is not well established. In this study, radiological and MRI parameters of DDD were assessed and compared with morphological and biochemical findings of disc degeneration. Thirty-nine human lumbar discs (L1-S1), age 19-86 years, were harvested from eight cadavers. Within 48 h postmortem, MRIs in various spin-echo sequences and biplanar radiographs of intact spines were obtained. Individual discs with endplates were then sectioned in the mid-sagittal plane and graded according to the morphological appearance. Samples from the nucleus of each disc were harvested for biochemical analysis including water and proteoglycan contents. On MRIs, T2-signal intensity, Modic changes, disc extension beyond the interspace (DEBIT), nucleus pulposus shape, annular tears, osteophytes and endplate integrity were graded. On radiographs, an independent observer classified the parameters disc height, endplate sclerosis, osteophytes, Schmorl's nodes, intradiscal calcifications and endplate shape. General linear-regression models were used for statistical analysis. Backward elimination with a 10% significance cut-off level was used to identify the most significant parameters, which then were summed to create composite scores for radiography, MRI and the combination of both methods. The grading was performed by three observers, and a reliability analysis using Cronbach's alpha model was used to control interobserver agreement. The three radiographic parameters height-loss, osteophytes and intradiscal calcifications correlated significantly with the morphological degree of degeneration (p<0.001, R2=642). Significant differences of even one morphological grade could also be differentiated in the composite radiological score (p<0.05), except at the extremes between grades 1 and 2 and grades 4 and 5. All MRI parameters correlated significantly with the morphological grade (p<0.05); however Modic changes, T2-intensity and osteophytes accounted for 83% of the variation in the data. T2-signal intensity correlated significantly with H2O and proteoglycan content (p<0.001), and was best for detecting highly degenerated discs. Regression showed that the combined score was better correlated with the morphological grade (p<0.001, R2=775) than either the composite radiographic (p<0.001, R2=642) or composite MRI (p<0.001, R2=696) alone. Based on the combined score, a backwards elimination of the regression was performed, in which the parameters Modic changes, and T2-intensity loss (MRI) as well as calcifications (X-ray) accounted for 87% of the variability. The interobserver validation showed a high correlation for all three scores (Cronbach's alpha values ranging from 0.95 to 0.97). CONCLUSION: Selective imaging parameters and a newly created scoring scheme were found to correlate with disc degeneration as determined in a morphological manner. Surprisingly, radiographic parameters were able to distinguish different stages of degeneration, whereas MRI could only detect advanced stages of disc degeneration. We conclude that X-rays may remain a cost-effective, non-invasive in vivo-grading method to detect early disc degeneration, and, combined with MRI, correlate best with morphological and biochemical assessment of disc degeneration.

2004 Young Investigator Award Winner: vertebral endplate marrow contact channel occlusions and intervertebral disc degeneration.

STUDY DESIGN: Intervertebral disc degeneration was evaluated by morphologic appearance, magnetic resonance imaging, and by biochemical matrix composition. Caliber and distribution of openings of the adjacent vertebral osseous endplates were measured. OBJECTIVES: Correlation between occlusion of vertebral endplate openings and intervertebral disc degeneration was quantified. SUMMARY OF BACKGROUND DATA: Calcifications of vertebral endplates with disease and age have suggested insufficient nutrition as a mechanism for intervertebral disc degeneration. It has been proposed that occlusion of endplate openings, which contain vascular sources for the disc, may limit the transport of nutrients, leading to disc degeneration. METHODS: Fresh magnetic resonance images from 39 human lumbar discs were scored. Sectioned discs with endplates were morphologically graded. Samples of nuclear and anular regions were evaluated for proteoglycan and collagen contents. Backlight microscopic images of 4 endplate regions were obtained, and caliber and distribution of endplate openings for each disc were measured. Analysis of variance regression models were used to assess correlation between endplate openings and disc degeneration. RESULTS: The decrease in opening density significantly correlated to morphologic degeneration grade, best for openings with 20 to 50 im equivalent diameter and in the nuclear region. Although the density of 20 to 50 im openings also significantly indirectly correlated to age, it was not as strong as the correlation to degeneration grade. Opening density was also significantly correlated to proteoglycan content in all regions. However, all other biochemical parameters as well as the T2 intensity score showed only weak or no correlation to opening density. CONCLUSIONS: A high indirect correlation between the density of openings in the osseous endplate (particularly of the size of the capillary buds) and the morphologic degeneration grade of the disc support the hypothesis that occlusion of these openings may deprive the cells of nutrients, leading to insufficient maintenance of the extracellular matrix and disc degeneration.

The current treatment--a survey of osteoporotic fracture treatment. Osteoporotic spine fractures: the spine surgeon's perspective.

For the spine surgeon the problems of the osteoporotic spine can be a painful simple compression fracture, the persisting instability after a fracture, the silent loss of posture due to progressive collapse of multiple vertebrae, and neurological complications related to an osteoporotic fracture. The use of polymethylmethacrylate (PMMA) for the reinforcement of osteoporotic vertebral bodies has turned out to be extremely efficient. Although the natural course of pain due to vertebral fractures decreases within the first weeks in the majority of patients, there remains a number of them with persistent pain and/or ongoing vertebral collapse. With percutaneous cement injection one can achieve fast and lasting pain reduction in 80% to 93% of patients. With multilevel injections it is possible to address the severe osteoporotic spine as a whole where we can halt the ongoing collapse and preserve posture efficiently. Rectification of lordosis can be achieved in fresh fractures with the kyphoplasty technique and, even more efficiently, with the lordoplasty procedure. Kyphosis correction ranges from 8.5 degrees to 14 degrees , and restoration of vertebral body (VB) height goes up to 90%. When surgical stabilization is required, the combination of PMMA reinforcement and fixation with screws appears the only alternative in order to anchor the implants in the severely osteoporotic bone.

Surgical treatment with the technique of Galveston-Luque of spine deformities associated with the prune belly syndrome.

In prune belly syndrome (PBS), spine deformities, particularly scoliosis, are the most frequent accompanying orthopedic problem (18%). We give a detailed description of two cases with prune belly-associated spine deformities. A combined anterior-posterior fusion procedure with posterior correction and stabilization according to the Galveston-Luque technique was carried out in both patients. In one patient insufficiency of the pelvic fixation developed with loss of balance. Reoperation was performed. In the second patient correct alignment and balance was achieved, breakage of both metal rods occurred over time. The PBS represents a model of how scoliosis may emerge. The lack of the abdominal musculature subsequently leads to congenial imbalance, which may cause lordo-scoliotic deformity. The operative treatment of spine deformities in the PBS in the presented technique is efficient. In great deformities balance is difficult to achieve and maintain especially in the lumbo-sacral junction.