Proton density fat fraction of the spinal column: an MRI cadaver study.

BACKGROUND: The increased popularity of minimally invasive spinal surgery calls for a revision of guidance techniques to prevent injuries of nearby neural and vascular structures. Lipid content has previously been proposed as a distinguishing criterion for different bone tissues to provide guidance along the interface of cancellous and cortical bone. This study aims to investigate how fat is distributed throughout the spinal column to confirm or refute the suitability of lipid content for guidance purposes. RESULTS: Proton density fat fraction (PDFF) was assessed over all vertebral levels for six human cadavers between 53 and 92 years of age, based on fat and water MR images. According to their distance to the vertebra contour, the data points were grouped in five regions of interest (ROIs): cortical bone (-1 mm to 0 mm), pre-cortical zone (PCZ) 1-3 (0-1 mm; 1-2 mm; 2-3 mm), and cancellous bone ([Formula: see text] 3 mm). For PCZ1 vs. PCZ2, a significant difference in mean PDFF of between -7.59 pp and -4.39 pp on average was found. For cortical bone vs. PCZ1, a significant difference in mean PDFF of between -27.09 pp and -18.96 pp on average was found. CONCLUSION: A relationship between distance from the cortical bone boundary and lipid content could be established, paving the way for guidance techniques based on fat fraction detection for spinal surgery.

Observations on the recovering lumbar spinal cord of lizards show multiple origins of the cells forming the bridge region including immune cells.

After transection the lumbar spinal cord of lizards forms a bridge of connective and nervous tissues between the severed proximal and distal ends of the cord. The types of proliferating cells activated in the injured spinal cord have been analyzed using light and ultrastructural immunolabeling for 5BrdU and nestin from 11 to 34 days after injury, when recovery of some hindlimb movements has occurred. At 11-22 days post-transection an intense proliferation of glial, immune and meningeal cells takes place. Nestin is almost absent in the normal spinal cord but becomes detectable at 11-34 days postinjury in ependymal and sparse glial cells located in the bridge region. At 11-22 days postinjury also numerous macrophages, lymphocytes, and some plasma cells appear proliferating during the intense inflammatory and antimicrobial phase. Phagocytosis within the injured spinal cord probably decreases inflammation and may indirectly promote axonal regeneration. Proliferating cells likely derive from precursor or stem elements of the reactive ependymal epithelium, but also from glial cells and meningeal fibroblasts. This is indicated by the presence of 5BrdU-long retaining labeling cells of glial and fibroblast types located in the stumps of the spinal cord and in the bridge. The present observations suggest that meningeal, ependymal, and numerous glial cells are the precursors of those forming the bridge region. Among glial cells, sparse oligodendrocytes myelinating the few axons present at 34 day after the injury also appear capable to proliferate. The myelinated axons are probably involved in the limited but important functional recovery of limb movements observed after 30-90 days postinjury.

Reliability of measuring the fat content of the lumbar vertebral marrow and paraspinal muscles using MRI mDIXON-Quant sequence.

PURPOSE: We aimed to assess the reliability of measuring the fat content of the lumbar vertebral marrow and the paraspinal muscles using magnetic resonance imaging (MRI) mDIXON-Quant sequence. METHODS: Thirty-one healthy volunteers were included. All participants underwent liver mDIXON-Quant imaging on a 3.0 T Philips MRI scanner by observer A. Within two weeks, observer B repeated the scan. After the examination, each observer independently measured the fat content of the third lumbar vertebra (L3), and the psoas (PS), erector spinae (ES), and multifidus (MF) muscles on central L3 axial images. After two weeks, each observer repeated the same measurements. They were blinded to their previous results. Reliability was estimated by evaluating the repeatability and reproducibility. RESULTS: The repeatability of the fat content measurements of L3, PS, ES, and MF was high. The intraclass correlation coefficients of the fat content of L3, PS, ES, and MF were 0.997, 0.984, 0.997, and 0.995 for observer A and 0.948, 0.974, 0.963, and 0.995 for observer B, respectively. The reproducibility of the measurement of the fat content of L3, PS, ES, and MF was high, and the interclass correlation coefficients were 0.984, 0.981, 0.977, and 0.998, respectively. CONCLUSION: Using mDIXON-Quant imaging to measure the fat content of the lumbar vertebral marrow and paraspinal muscles shows high reliability and is suitable for use in clinical practice.

The clinical relevance of the presence of bridging syndesmophytes on kyphosis correction and maintenance following pedicle subtraction osteotomy for thoracolumbar kyphotic deformity in ankylosing spondylitis: a comparative cohort study.

BACKGROUND: The presence of bridging syndesmophytes (BS) in spinal osteotomy region serves traditionally as one critical determinant for selection of osteotomy techniques. While nowadays the proportion of kyphotic ankylosing spondylitis (AS) patients receiving pedicle subtraction osteotomy (PSO) with yet mobile neighboring disc has seen a substantial increase. Literatures investigating the clinical relevance of the presence of BS on kyphosis correction and maintenance following PSO are scarce. METHODS: A total of 71 thoracolumbar kyphotic AS patients treated with single-level PSO at our hospital between September 2010 and August 2014 were retrospectively reviewed, 32 of whom were stratified into the BS group (BG). The operative corrections of multiple spino-pelvic sagittal parameters were assessed. Comparison of the contribution of adjacent disc wedging to total correction per PSO segment was made between the BS and non-BS groups (NBG). The correction loss were also evaluated and compared with a minimum 2-year follow-up. RESULTS: A significantly younger age (30.97 ± 8.28 vs. 40.31 ± 8.44 yrs., p < 0.001), smaller pelvic incidence (PI) (43.03 ± 10.60 vs. 49.36 ± 9.75°, p = 0.011), greater wedging index of osteotomized vertebra (1.17 ± 0.16 vs. 1.09 ± 0.08, p = 0.011) and larger local kyphosis (19.59 ± 10.84 vs. 13.56 ± 8.50°, p = 0.013) was observed in NBG preoperatively. Patients in BG and NBG accomplished comparable amount of kyphosis correction per PSO segment (40.22 ± 7.09 vs. 43.85 ± 8.71°, p = 0.062). However, the contribution of adjacent disc wedging to total correction per PSO was significantly larger in NBG [8.10 ± 6.19 (18.5%) vs. 1.09 ± 2.88° (2.7%), p < 0.001]. By ultimate follow-up, the global kyphosis (18.26 ± 10.97 vs. 21.51 ± 10.89°, p < 0.05) and thoracic kyphosis (37.95 ± 11.87 vs. 42.87 ± 11.56°, p < 0.05) deteriorated significantly in the NBG but not BG, so was further pelvic retroversion as represented by increased pelvic tilt (19.46 ± 8.13 vs. 23.44 ± 8.19°, p < 0.05) and decreased sacral slope (23.02 ± 9.12 vs. 18.62 ± 10.10°, p < 0.05). Loss of corrections concerning contribution of adjacent disc wedging was also larger in NBG (1.41 ± 3.27 vs. 0.22 ± 1.49°, p < 0.05). CONCLUSIONS: Our study might suggest that the evaluation and treatment methods of kyphotic AS patients needed to be fine-tuned with appropriate subgrouping by the presence of syndesmophytes with bamboo sign as they were potentially distinct groups with different PI, contributor of lordosing capability and prognosis that might require separate analysis.

Biomechanical properties of 3D-printed bone scaffolds are improved by treatment with CRFP.

BACKGROUND: One of the major challenges in orthopedics is to develop implants that overcome current postoperative problems such as osteointegration, proper load bearing, and stress shielding. Current implant techniques such as allografts or endoprostheses never reach full bone integration, and the risk of fracture due to stress shielding is a major concern. To overcome this, a novel technique of reverse engineering to create artificial scaffolds was designed and tested. The purpose of the study is to create a new generation of implants that are both biocompatible and biomimetic. METHODS: 3D-printed scaffolds based on physiological trabecular bone patterning were printed. MC3T3 cells were cultured on these scaffolds in osteogenic media, with and without the addition of Calcitonin Receptor Fragment Peptide (CRFP) in order to assess bone formation on the surfaces of the scaffolds. Integrity of these cell-seeded bone-coated scaffolds was tested for their mechanical strength. RESULTS: The results show that cellular proliferation and bone matrix formation are both supported by our 3D-printed scaffolds. The mechanical strength of the scaffolds was enhanced by trabecular patterning in the order of 20% for compression strength and 60% for compressive modulus. Furthermore, cell-seeded trabecular scaffolds modulus increased fourfold when treated with CRFP. CONCLUSION: Upon mineralization, the cell-seeded trabecular implants treated with osteo-inductive agents and pretreated with CRFP showed a significant increase in the compressive modulus. This work will lead to creating 3D structures that can be used in the replacement of not only bone segments, but entire bones.

A comparative study of bone biopsies from the iliac crest, the tibial bone, and the lumbar spine.

BACKGROUND: Patients with an impaired renal function show a high incidence of bone and mineral disturbances. These 'chronic kidney disease - mineral and bone disorders' (CKD-MBD) range from high turnover osteoporosis to adynamic bone disease. Currently, the histomorphometric analysis of a bone biopsy taken from the iliac crest is viewed as the gold standard for CKD-MBD subtype differentiation. However, the clinical relevance of such a biopsy is questionable since iliac crest fractures are an extremely rare finding. Therefore, we aimed to elucidate if the histomorphometric parameter 'trabecular bone volume (BV/TV)' from the iliac crest is representative for other biopsy locations. We chose two skeletal sites of higher fracture risk for testing, namely, the tibial bone and the lumbar spine, to examine if the current gold standard of bone biopsy is indeed golden. METHODS: Bone biopsies were taken from 12 embalmed body donors at the iliac crest, the proximal tibia, and the lumbar vertebral body, respectively. Masson-Goldner stained sections of methyl methacrylate embedded biopsies were used for trabecular bone volume calculation. Furthermore, exemplary µ-computed tomography (XtremeCT) scans with subsequent analysis were performed. RESULTS: Median values of trabecular bone volume were comparable between all body donors with median (interquartile range, IQR) 18.3% (10.9-22.9%) at the iliac crest, 21.5% (9.5-40.1%) at the proximal tibia, and 16.3% (11.4-25.0%) at the lumbar spine. However, single values showed extensive intra-individual variation, which were also confirmed by XtremeCT imaging. CONCLUSIONS: Distinct intra-individual heterogeneity of trabecular bone volume elucidate why a bone biopsy from one site does not necessarily predict patient relevant endpoints like hip or spine fractures. Physicians interpreting bone biopsy results should know this limitation of the current gold standard for CKD-MBD diagnostic, especially, when systemic therapeutic decisions should be based on it.

How maturity influences annulus-endplate integration in the ovine intervertebral disc: a micro- and ultra-structural study.

The annulus-endplate anchorage system plays a vital role in structurally linking the compliant disc to its adjacent much more rigid vertebrae. Past literature has identified the endplate as a region of weakness, not just in the mature spine but also in the immature spine. The aim of this structural study was to investigate in detail the morphological changes associated with annulus-endplate integration through different stages of maturity. Ovine lumbar motion segments were collected from two immature age groups: (i) newborn and (ii) spring lamb (roughly 3 months old); these were compared with a third group of previously analysed mature ewe samples (3-5 years). Sections from the posterior region of each motion segment were obtained for microstructural analysis and imaged in their fully hydrated state via differential interference contrast (DIC) optical microscopy. Selected slices were further prepared and imaged via scanning electron microscopy (SEM) to analyse fibril-level modes of integration. Despite significant changes in endplate morphology, the annular fibre bundles in all three age groups displayed a similar branching mechanism, with the main bundle splitting into several sub-bundles on entering the cartilaginous endplate. This morphology, previously described in the mature ovine disc, is thought to strengthen significantly annulus-endplate integration. Its prevalence from an age as young as birth emphasizes the critical role that it plays in the anchorage system. The structure of the branched sub-bundles and their integration with the surrounding matrix were found to vary with age due to changes in the cartilaginous and vertebral components of the endplate. Microscopically, the sub-bundles in both immature age groups appeared to fade into the surrounding tissue due to their fibril-level integration with the cartilaginous endplate tissue, this mechanism being particularly complex in the spring lamb disc. However, in the fully mature disc, the sub-bundles remained as separate entities throughout the full depth of their anchorage into the cartilaginous endplate. Cell morphology was also found to vary with maturity within the cartilaginous matrix and it is proposed that this relates to endplate development and ossification.

Freeze-Dried Platelet-Rich Plasma Accelerates Bone Union with Adequate Rigidity in Posterolateral Lumbar Fusion Surgery Model in Rats.

Fresh platelet-rich plasma (PRP) accelerates bone union in rat model. However, fresh PRP has a short half-life. We suggested freeze-dried PRP (FD-PRP) prepared in advance and investigated its efficacy in vivo. Spinal posterolateral fusion was performed on 8-week-old male Sprague-Dawley rats divided into six groups based on the graft materials (n = 10 per group): sham control, artificial bone (A hydroxyapatite-collagen composite) -alone, autologous bone, artificial bone + fresh-PRP, artificial bone + FD-PRP preserved 8 weeks, and artificial bone + human recombinant bone morphogenetic protein 2 (BMP) as a positive control. At 4 and 8 weeks after the surgery, we investigated their bone union-related characteristics including amount of bone formation, histological characteristics of trabecular bone at remodeling site, and biomechanical strength on 3-point bending. Comparable radiological bone union was confirmed at 4 weeks after surgery in 80% of the FD-PRP groups, which was earlier than in other groups (p < 0.05). Histologically, the trabecular bone had thinner and more branches in the FD-PRP. Moreover, the biomechanical strength was comparable to that of autologous bone. FD-PRP accelerated bone union at a rate comparable to that of fresh PRP and BMP by remodeling the bone with thinner, more tangled, and rigid trabecular bone.

Development of a Large Animal Long-Term Intervertebral Disc Organ Culture Model That Includes the Bony Vertebrae for Ex Vivo Studies.

Intervertebral disc (IVD) degeneration is a common cause of low back pain. Testing potential therapeutics in the regeneration of the disc requires the use of model systems. Although several animal models have been developed to investigate IVD degeneration, they are technically challenging to prepare, expensive, present with limitations when performing biomechanical studies on the disc, and are impractical in large-scale screening of novel anabolic and scaffolding agents. An IVD organ culture system offers an inexpensive alternative. In the current paradigm, the bony endplates are removed to allow for nutrient diffusion and maintenance of disc cell viability. Although this is an excellent system for testing biologics, it results in concave cartilage endplates and, as such, requires special platens for loading purposes in a bioreactor as flat ones can overload the annular disc region leading to improper loading. Furthermore, the absence of bone makes it unsuitable for applying complex cyclic loading, a topic of interest in the study of chronic progressive degeneration, as multiaxial loading is more representative of daily forces encountered by the IVD. We have developed and validated a novel long-term IVD organ culture model that retains vertebral bone and is easy to prepare. Our model is ideal for testing potential drugs and alternate-based therapies, in addition to investigating the long-term effects of loading paradigms on disc degeneration and repair.

Conditional Deletion of Prolyl Hydroxylase Domain-Containing Protein 2 (Phd2) Gene Reveals Its Essential Role in Chondrocyte Function and Endochondral Bone Formation.

The hypoxic growth plate cartilage requires hypoxia-inducible factor (HIF)-mediated pathways to maintain chondrocyte survival and differentiation. HIF proteins are tightly regulated by prolyl hydroxylase domain-containing protein 2 (Phd2)-mediated proteosomal degradation. We conditionally disrupted the Phd2 gene in chondrocytes by crossing Phd2 floxed mice with type 2 collagen-α1-Cre transgenic mice and found massive increases (>50%) in the trabecular bone mass of long bones and lumbar vertebra of the Phd2 conditional knockout (cKO) mice caused by significant increases in trabecular number and thickness and reductions in trabecular separation. Cortical thickness and tissue mineral density at the femoral middiaphysis of the cKO mice were also significantly increased. Dynamic histomorphometric analyses revealed increased longitudinal length and osteoid surface per bone surface in the primary spongiosa of the cKO mice, suggesting elevated conversion rate from hypertrophic chondrocytes to mineralized bone matrix as well as increased bone formation in the primary spongiosa. In the secondary spongiosa, bone formation measured by mineralizing surface per bone surface and mineral apposition rate were not changed, but resorption was slightly reduced. Increases in the mRNA levels of SRY (sex determining region Y)-box 9, osterix (Osx), type 2 collagen, aggrecan, alkaline phosphatase, bone sialoprotein, vascular endothelial growth factor, erythropoietin, and glycolytic enzymes in the growth plate of cKO mice were detected by quantitative RT-PCR. Immunohistochemistry revealed an increased HIF-1α protein level in the hypertrophic chondrocytes of cKO mice. Infection of chondrocytes isolated from Phd2 floxed mice with adenoviral Cre resulted in similar gene expression patterns as observed in the cKO growth plate chondrocytes. Our findings indicate that Phd2 suppresses endochondral bone formation, in part, via HIF-dependent mechanisms in mice.

Isolation and Characterization of Human Mesenchymal Stem Cells From Facet Joints and Interspinous Ligaments.

STUDY DESIGN: A descriptive in vitro study on isolation and differentiation of human mesenchymal stem cells (MSCs) derived from the facet joints and interspinous ligaments. OBJECTIVE: To isolate cells from the facet joints and interspinous ligaments and investigate their surface marker profile and differentiation potentials. SUMMARY OF BACKGROUND DATA: Lumbar spinal canal stenosis and ossification of the posterior longitudinal ligament are progressive conditions characterized by the hypertrophy and ossification of ligaments and joints within the spinal canal. MSCs are believed to play a role in the advancement of these diseases and the existence of MSCs has been demonstrated within the ligamentum flavum and posterior longitudinal ligament. The aim of this study was to investigate whether these cells could also be found within facet joints and interspinous ligaments. METHODS: Samples were harvested from 10 patients undergoing spinal surgery. The MSCs from facet joints and interspinous ligaments were isolated using direct tissue explant technique. Cell surface antigen profilings were performed via flow cytometry. Their lineage differentiation potentials were analyzed. RESULTS: The facet joints and interspinous ligaments-derived MSCs have the tri-lineage potential to be differentiated into osteogenic, adipogenic, and chondrogenic cells under appropriate inductions. Flow cytometry analysis revealed both cell lines expressed MSCs markers. Both facet joints and interspinous ligaments-derived MSCs expressed marker genes for osteoblasts, adipocytes, and chondrocytes. CONCLUSION: The facet joints and interspinous ligaments may provide alternative sources of MSCs for tissue engineering applications. The facet joints and interspinous ligaments-derived MSCs are part of the microenvironment of the human ligaments of the spinal column and might play a crucial role in the development and progression of degenerative spine conditions.

Activation of Protein Kinase A in Mature Osteoblasts Promotes a Major Bone Anabolic Response.

Protein kinase A (PKA) regulates osteoblast cell function in vitro and is activated by important bone mass modulating agents. We determined whether PKA activation in osteoblasts is sufficient to mediate a bone anabolic response. Thus, a mouse model conditionally expressing a constitutively active PKA (CA-PKA) in osteoblasts (CA-PKA-OB mouse) was developed by crossing a 2.3-kb α1 (I)-collagen promoter-Cre mouse with a floxed-CA-PKA mouse. Primary osteoblasts from the CA-PKA-OB mice exhibited higher basal PKA activity than those from control mice. Microcomputed tomographic analysis revealed that CA-PKA-OB female mice had an 8.6-fold increase in femoral but only 1.16-fold increase in lumbar 5 vertebral bone volume/total volume. Femur cortical thickness and volume were also higher in the CA-PKA-OB mice. In contrast, alterations in many femoral microcomputed tomographic parameters in male CA-PKA-OB mice were modest. Interestingly, the 3-dimensional structure model index was substantially lower both in femur and lumbar 5 of male and female CA-PKA-OB mice, reflecting an increase in the plate to rod-like structure ratio. In agreement, femurs from female CA-PKA-OB mice had greater load to failure and were stiffer compared with those of control mice. Furthermore, the CA-PKA-OB mice had higher levels of serum bone turnover markers and increased osteoblast and osteoclast numbers per total tissue area compared with control animals. In summary, constitutive activation of PKA in osteoblasts is sufficient to increase bone mass and favorably modify bone architecture and improve mechanical properties. PKA activation in mature osteoblasts is, therefore, an important target for designing anabolic drugs for treating diseases with bone loss.

Stochastic predictors from the DXA scans of human lumbar vertebrae are correlated with the microarchitecture parameters of trabecular bone.

The purpose of this study was to provide a novel stochastic assessment of inhomogeneous distribution of bone mineral density (BMD) from the Dual-energy X-ray Absorptiometry (DXA) scans of human lumbar vertebrae and identify the stochastic predictors that were correlated with the microarchitecture parameters of trabecular bone. Eighteen human lumbar vertebrae with intact posterior elements from 5 cadaveric spines were scanned in the posterior-anterior projection using a Hologic densitometer. The BMD map of human vertebrae was obtained from the raw data of DXA scans by directly operating on the transmission measurements of low- and high-energy X-ray beams. Stochastic predictors were calculated by fitting theoretical models onto the experimental variogram of the BMD map, rather than grayscale images, from DXA scans. In addition, microarchitecture parameters of trabecular bone were measured from the 3D images of human vertebrae acquired using a Micro-CT scanner. Significant correlations were observed between stochastic predictors and microarchitecture parameters. The sill variance, representing the standard deviation of the BMD map to some extent, had significantly positive correlations with bone volume, trabecular thickness, trabecular number and connectivity density. The sill variance was also negatively associated with bone surface to volume ratio and trabecular separation. This study demonstrates that the stochastic assessment of the inhomogeneous distribution of BMD from DXA scans of human lumbar vertebrae can reveal microarchitecture information of trabecular bone. However, future studies are needed to examine the potential of stochastic predictors from routine clinical DXA scans in providing bone fragility information complementary to BMD.

A comprehensive review of the sub-axial ligaments of the vertebral column: part II histology and embryology.

BACKGROUND: As important as the vertebral ligaments are in maintaining the integrity of the spinal column and protecting the contents of the spinal canal, a single detailed review of their histology and embryology is missing in the literature. METHODS: A literature search using online search engines was conducted. RESULTS: Single comprehensive reviews of the histology and embryology of the spinal ligaments are not found in the extant medical literature. CONCLUSIONS: This review will be useful to those who study or treat patients with pathology of the spine.

MicroRNA-494 inhibition protects nucleus pulposus cells from TNF-α-induced apoptosis by targeting JunD.

BACKGROUND: Human nucleus pulposus cell (HNPC) apoptosis plays an important role in the development of intervertebral disc degeneration (IVDD). Our previous research revealed that among all of the dysregulated microRNAs in the degenerated nucleus pulposus tissues of patient with IVDD, miRNA-494 (miR-494) is the most significantly increased. However, the influence of miR-494 HNPC apoptosis has not been confirmed. OBJECTIVE: This study was designed to evaluate the effect of miR-494 on the HNPC apoptosis induced by TNF-α and to explore the possible mechanism of this process. METHODS: First, HNPCs were stimulated with TNF-α at different concentrations (0 ng/ml, 10 ng/ml, 50 ng/ml, or 100 ng/ml) for 0 h, 8 h, 16 h, or 24 h. Annexin V-PE/7-AAD assays and real-time quantitative PCR were used to detect the cell apoptosis rates and miR-494 expression. Second, we successfully knocked down endogenous miR-494 in HNPCs via lentiviral antigomiR-494 vector infection and then stimulated with TNF-α (100 ng/ml, 16 h). The rates of apoptosis and miR-494 expression were then detected again. Additionally, a dual-luciferase reporter assay and western blotting were used to determine whether JunD is a target of miR-494. Finally, western blotting was used to analyze the expression of cytochrome C. RESULTS: We found that the rate of apoptosis increased with concentration, time (p < 0.05) and miR-494 expression (p < 0.05). The rate of apoptosis in the 100 ng/ml, 16 h group appeared to be suitable. After transfection, the apoptosis rate and miR-494 expression were significantly decreased in the antigomiR-494+TNF-α group compared to the controls (p < 0.05). We also revealed that JunD is a target of miR-494. Western blotting analysis demonstrated that treatment with the lentiviral antigomiR-494 vector resulted in increased expression of JunD (p < 0.05) and decreased expression of cytochrome C (p < 0.05). CONCLUSION: These results indicated that miR-494 is a novel regulator of HNPC apoptosis induced by TNF-α. The knock-out of miR-494 expression protected the HNPCs from apoptosis via the up-regulation of JunD, which was possibly mediated via cytochrome C apoptotic signaling. These findings suggest that the miR-494/JunD signaling pathway might represent a novel therapeutic target for the prevention of IVDD.

A multi-target approach for pain treatment: dual inhibition of fatty acid amide hydrolase and TRPV1 in a rat model of osteoarthritis.

The pharmacological inhibition of anandamide (AEA) hydrolysis by fatty acid amide hydrolase (FAAH) attenuates pain in animal models of osteoarthritis (OA) but has failed in clinical trials. This may have occurred because AEA also activates transient receptor potential vanilloid type 1 (TRPV1), which contributes to pain development. Therefore, we investigated the effectiveness of the dual FAAH-TRPV1 blocker OMDM-198 in an MIA-model of osteoarthritic pain. We first investigated the MIA-induced model of OA by (1) characterizing the pain phenotype and degenerative changes within the joint using X-ray microtomography and (2) evaluating nerve injury and inflammation marker (ATF-3 and IL-6) expression in the lumbar dorsal root ganglia of osteoarthritic rats and differences in gene and protein expression of the cannabinoid CB1 receptors FAAH and TRPV1. Furthermore, we compared OMDM-198 with compounds acting exclusively on FAAH or TRPV1. Osteoarthritis was accompanied by the fragmentation of bone microstructure and destroyed cartilage. An increase of the mRNA levels of ATF3 and IL-6 and an upregulation of AEA receptors and FAAH in the dorsal root ganglia were observed. OMDM-198 showed antihyperalgesic effects in the OA model, which were comparable with those of a selective TRPV1 antagonist, SB-366,791, and a selective FAAH inhibitor, URB-597. The effect of OMDM-198 was attenuated by the CB1 receptor antagonist, AM-251, and by the nonpungent TRPV1 agonist, olvanil, suggesting its action as an "indirect" CB1 agonist and TRPV1 antagonist. These results suggest an innovative strategy for the treatment of OA, which may yield more satisfactory results than those obtained so far with selective FAAH inhibitors in human OA.

Gene expression profile analysis of human mesenchymal stem cells from herniated and degenerated intervertebral discs reveals different expression of osteopontin.

Gene expression analysis provides an effective methodology to identify clinically relevant genes implicated in intervertebral disc (IVD) pathology. The analysis of gene profile in mesenchymal stem cells (MSCs) from human herniated IVD (H-IVD) and degenerated IVD (D-IVD) has not yet been investigated. We present in this study a characterization of MSCs isolated from clinically categorized H-IVD and D-IVD disc samples. H-IVD-MSCs and D-IVD-MSCs showed multipotent mesenchymal differentiation ability, expressing positivity for adipogenic, osteogenic, and chondrogenic markers with an immunophenotypical profile representative of MSCs. FACS analyses revealed a higher expression of CD44 in D-IVD-MSCs compared to H-IVD-MSCs. Gene expression profile revealed that most genes under investigation displayed large variations and were not significantly different in the two types of analyzed IVD-MSCs. Conversely, the gene expression of osteopontin (OPN), a protein involved in bone matrix mineralization and extracellular matrix destruction, was found markedly increased (more than 400-fold) in D-IVD-MSCs compared to H-IVD-MSCs. Moreover, the OPN protein expression was detectable only in D-IVD-MSCs, and its levels were directly related with D-IVD severity. These findings suggest that an abnormal expression of OPN in D-IVD-MSCs occurs and plays a pivotal role in the pathophysiological process of human disc degeneration. We speculate that the regulation of the OPN pathway might be a therapeutic target to counteract disc degeneration.

Aberrant bone density in aging mice lacking the adenosine transporter ENT1.

Adenosine is known to regulate bone production and resorption in humans and mice. Type 1 equilibrative nucleoside transporter (ENT1) is responsible for the majority of adenosine transport across the plasma membrane and is ubiquitously expressed in both humans and mice. However, the contribution of ENT1-mediated adenosine levels has not been studied in bone remodeling. With the recent identification of the importance of adenosine signaling in bone homeostasis, it is essential to understand the role of ENT1 to develop novel therapeutic compounds for bone disorders. Here we examined the effect of ENT1 deletion on bone density using X-ray, dual energy X-ray absorptiometry and micro-computerized tomography analysis. Our results show that bone density and bone mineral density is reduced in the lower thoracic and lumbar spine as well as the femur of old ENT1 null mice (>7 months) compared to wild-type littermates. Furthermore, we found increased mRNA expression of tartrate-resistant acid phosphatase (TRAP), an osteoclast marker, in isolated long bones from 10 month old ENT1 null mice compared to wild-type mice. In addition, aged ENT1 null mice displayed severe deficit in motor coordination and locomotor activity, which might be attributed to dysregulated bone density. Overall, our study suggests that ENT1-regulated adenosine signaling plays an essential role in lumbar spine and femur bone density.

Influence of bone marrow composition on measurements of trabecular microstructure using decay due to diffusion in the internal field MRI: simulations and clinical studies.

PURPOSE: Decay due to diffusion in the internal field (DDIF) MRI allows for measurements of microstructures of porous materials at low spatial resolution and thus has potential for trabecular bone quality measurements. In trabecular bone, solid bone changes (osteoporosis) as well as changes in bone marrow composition occur. The influence of such changes on DDIF MRI was studied by simulations and in vivo measurements. METHODS: Monte Carlo simulations of DDIF in various trabecular bone models were conducted. Changes in solid bone and marrow composition were simulated with numerical bone erosion and marrow susceptibility variations. Additionally, in vivo measurements were performed in the lumbar spine of healthy volunteers aged 23-62 years. RESULTS: Simulations and in vivo results showed that 1) DDIF decay times decrease with increasing marrow fat and 2) the marrow fat percentage needs to be incorporated in the DDIF analysis to discriminate between healthy and osteoporotic solid bone structures. CONCLUSIONS: Bone marrow composition plays an important role in DDIF MRI: incorporation of marrow fat percentage into DDIF MRI allowed for differentiation of young and old age groups (in vivo experiments). DDIF MRI may develop into a means of assessing osteoporosis and disorders that affect marrow composition.

Histological features of endplates of the mammalian spine: from mice to men.

STUDY DESIGN: Histological features of the intervertebral disc (IVD)-endplate interface were analyzed. OBJECTIVE: To define cartilaginous and bony vertebral endplate in commonly used laboratory animals and compare with that of the humans. SUMMARY OF BACKGROUND DATA: Endplates are crucial for the IVD nutrient supply: the IVDs have limited blood supply; most nutrients diffuse through endplates to nourish the discs. Various animal models of IVD and endplate degeneration have been used to study the etiology and treatments of spinal disorders. However, because humans are biped, the spine mechanics differ significantly from other mammals. Translation of animal research findings requires a characterization and comparison of the vertebral endplate in the respective species. In this study, we compared the endplate structure of laboratory animal species at the age range commonly used for modeling spine degeneration with that of an adult human. METHODS: Mouse, rat, rabbit, goat, and human IVDs and the adjacent vertebral bodies were isolated from the lower lumbar spine. Tissues were stained with Alcian Blue, counterstained with hematoxylin and eosin. RESULTS: Structure of the vertebral endplate varied significantly between the adult animal species and that of the humans. Growth plates persisted in all adult animals studied, whereas the growth plate is absent in the adult humans. In the mice and rats, the cartilaginous endplates are in continuation with the growth plates, with only a small bony center. Rabbits and goats have a bony layer between cartilaginous endplate and the growth plate. The human endplate consist of a cartilaginous layer and the bony endplate. CONCLUSION: Significant differences exist in histological features of the endplate across animal species and that of the humans. Consideration should be given when animal models are used to study IVD degeneration and surgical treatments. LEVEL OF EVIDENCE: 5.