Degenerative Cascade Research Articles

Roles of Ataxin-2 in Pathological Cascades Mediated by TAR DNA-binding Protein 43 (TDP-43) and Fused in Sarcoma (FUS)

The RNA-binding proteins TDP-43 and Fused in Sarcoma (FUS) play central roles in neurodegeneration associated with amyotrophic lateral sclerosis and frontotemporal lobar degeneration. Both proteins are components of messenger ribonucleoprotein (mRNP) granules and show cytoplasmic mislocalization in affected tissues. Recently, ataxin-2 was identified as a potent modifier of TDP-43 toxicity in an RNA-dependent manner. This study investigated to clarify how ataxin-2 modifies the TDP-43 and FUS pathological pathway. The expression of cytoplasmic TDP-43, the 35-kDa C-terminal fragment (TDP-p35f), and mutant FUS recruited ataxin-2 to mRNP granules, whereas increased ataxin-2 inhibited the mRNP granule formation of the 35-kDa C-terminal fragment and mutant FUS. A subcellular compartment analysis showed that the overexpressed ataxin-2 increased the cytoplasmic concentrations of both proteins, whereas it decreased their nuclear distributions. These data indicate that increased ataxin-2 impairs the assembly of TDP-43 and FUS into mRNP granules, leading to an aberrant distribution of RNA-binding proteins. Consequently, these sequences may exacerbate the impairment of the RNA-quality control system mediated by amyotrophic lateral sclerosis/frontotemporal lobar degeneration-associated RNA-binding proteins, which forms the core of the degenerative cascade.

Compression Failure in Lumbar Functional Spinal Units After Nucleus Augmentation with an Injectable In Situ Cured Hydrogel

Nucleus replacement devices are intended to replace or augment the pathologic intervertebral disc, particularly in the early stages of the degenerative cascade. In order to evaluate the clinical viability of these devices, it is necessary to conduct tests simulating potential failure modes in the physiologic setting in addition to bench-top mechanical device testing and traditional in vitro flexibility evaluation.

Development and neurodegeneration

The term “neurodegenerative” implies a period of normal development preceding cell death. Determining when decline begins and the selective vulnerability of specific brain regions to the degenerative cascade is key to development of disease-modifying therapies. Huntington disease (HD), an autosomal dominant disease with little contribution from modifier genes, and complete penetrance in individuals with CAG repeat length ≥40 in the Huntington ( Htt ) gene, permits the identification of an at-risk population for whom the probability of age at diagnosis can be modeled based on CAG repeat length and age. What if Huntington disease is both a neurodevelopmental disorder and a neurodegenerative disease? The possibility that mutant Htt ( mHtt ) might result in fundamental embryonic developmental abnormalities that correlate with the patterns of …

Influence of Autologous Mononuclear Stem-Cells over Symptomatic Disk Disease in Experimental Animal Model

Introduction Various therapeutic strategies have been used for controlling or at least slowing the degenerative cascade that leads to symptomatic disk degeneration. In this context, given its regenerative capability, stem cells (SC) therapy has shown promising results acting early on the degenerative cascade. This study aims to evaluate the influence of autologous adult mononuclear SC on the histological changes of experimentally induced disk degeneration. Materials and Methods After local Ethical Committee approval, 16 rabbits-New Zealand, male, 3.5 to 4.5 kg-underwent needle puncture of the three caudal lumbar intervertebral discs using a previously validated technique, and the remaining discs were used as controls. The animals received either autologous mesenchymal SC or saline solution after 2 weeks (group 1) or 2 months (group 2). Animals underwent euthanasia 8 weeks after the cell administration, and the discs were harvested for analysis. The slides were analyzed on hematoxylin-eosin stains for the presence of degeneration by a previously validated score, and by automated color morphometry, also previously validated by the authors, defining cellular size and cellular population within the nucleus pulposus (NP). Results Total 91 intervertebral discs (IVDs) were obtained for histological analysis, extracted from 16 animals-4 animals were excluded due to inadequate tissue preparation. The study group was divided as follows: 55 control-not punctured-IVDs and 36 experimental IVDs; were 21 have received SC and 15, isotonic saline solution. The microscopically characteristic alterations of the experimental disk degeneration have been observed in all punctured IVDs; however, some slides suggested a less intensive degeneration process and normal areas inside de nucleus pulposus, meaning recovery areas in the IVDs which have received mononuclear stem cells. Through color morphometry, significant differences favoring the SC group regarding number of cells and especially nuclear size ( p = 0.03) give indirect evidence that the injected cells survived within the NP. There was also a trend favoring the group receiving SC earlier (2 weeks). Conclusion Injection of SC was not able to interrupt the degenerative process triggered by disk puncture. However, the histological changes were less severe in the groups receiving cell therapy. Color morphometry showed clusters of cells within the NP, not seen in the control groups. I confirm having declared any potential conflict of interest for all authors listed on this abstract Yes Disclosure of Interest None declared

Correlation of Intervertebral Disk T2* MRI Presentation with 3D Kinematics

Introduction The causes of low back pain (LBP) are poorly defined and indistinct; the most often implicated tissue as the origin for pain is the intervertebral disk (IVD)1,2. Traditionally, lumbar spine range of motion measurements and IVD gross morphology (from MR images) are used to assess spinal health, but largely fail to correlate well with pain or provide clinically useful patient stratification3,4,5. Due to the inadequacies of current imaging techniques in differentiating symptomatic versus nonsymptomatic changes, a new series of quantitative protocols is emerging6. Quantitative T2* relaxation time measurements probe biochemical properties of the tissues. T2* mapping techniques are employed for cartilage assessment, probing spatial collagen architecture in conjunction with water molecule mobility7. As a first step, the goal of this work is to correlate quantitative MR mapping of the IVD with 3D kinematic data to probe the relationship between disk biochemical properties and functional kinematic parameters. Materials and Methods Total 18 osteoligamentous cadaveric lumbar spines (L3-S1), acquired from the UofM Bequest Program (aveage: 53.2 ± 15.5 years; range: 21 to 71 years) were examined using conventional and quantitative MR imaging protocols and biomechanically exercised in the cardinal planes. MR imaging was performed on a Siemens 3T scanner (Magnetom Trio; Siemens Health care). Conventional T2-weighted sagittal anatomic images were acquired for Pfirrmann disk grading (Fig. 1A)8. T2* relaxation maps were obtained using [TR(ms): 500; TE(ms): 4.18, 11.32, 18.46, 25.60, 32.74, 39.88; Voxel Size(mm): 0.5 × 0.5 × 3.0, Slices: 33]. Quantitative T2* maps were calculated (MapIt, Siemens Health care) and mean T2* values were recorded using Osirix Imaging Software across five regions of interest (ROI) (Fig. 1B). Utilizing MATLAB (Mathworks Inc. Natick, MA), the central ROI mean relaxation value was evaluated in the coronal plane as the intensity changed from lateral to medial to lateral (Fig. 1C). Using a least squares method, the slope of the transition zone (ms/mm) between the annulus fibrosus and nucleus pulposus on each side was computed (Fig. 1D). The area underneath the curve was calculated and normalized to the width of the IVD (Fig. 1D). After imaging, the specimens were embedded in polymethylmethacrylate and tested in a six-axis Spine Kinetic Simulator (8821 Biopuls, Instron, Norwood, MA, USA). Pure moments of up to 7 Nm were applied sinusoidally (0.015 Hz). Three cycles were performed and the final cycle was analyzed. Load and moment data was collected at 100 Hz. Segmental displacements were recorded using a 3D visual motion analysis 5-camera system (Vicon MX-F40NIR, Vicon Motion Systems, Centennial, CO) capturing a four-ball infrared reflecting marker set attached to each vertebral body. Kinematic outcomes (ROM, Neutral Zone, NZ/ROM, Stiffness) were correlated with imaging parameters using Pearson correlation test with alpha acceptance of 0.05. Results The T2* relaxation measurements in the nucleus pulposus were significantly correlated with flexion stiffness ( p = 0.024) and lateral bending stiffness ( p = 0.021) wherein increased stiffness was associated with lower signal and higher Pfirrmann grade. The slope of the transition zone for the 54 disk samples was found to be correlated with the flexion stiffness ( p = 0.026), flexion range of motion ( p = 0.002), lateral bending stiffness ( p = 0.029), and lateral bending range of motion ( p = 0.0028). The area under the T2* intensity curves in the coronal plane was likewise found to be significantly correlated with following biomechanical measures: flexion stiffness ( p = 0.043), flexion range of motion (p = 0.014), lateral bending stiffness (p = 0.021), and lateral bending range of motion ( p = 0.023). Conclusion MR imaging of IVD using T2* relaxation measurements provide quantitative assessment of the disk's biochemical properties which is related to its biomechanical competence. This new methodology and analysis technique may provide enhanced quantitative data on the degenerative cascade which alters disk function and in some cases leads to painful outcomes. Limitations of this research include the biomechanical testing of cadaveric samples without musculature and their postmortem imaging at room temperature. Next, we aim to evaluate this relationship in human subjects and determine its clinical efficacy. This research linked kinematic data with quantitative T2* mapping of IVD from across the degenerative spectrum. By coupling quantitative MR imaging with functional kinematic outcomes, these data may improve patient stratification and targeted therapeutic intervention for patients suffering from LBP. I confirm having declared any potential conflict of interest for all authors listed on this abstract Yes Disclosure of Interest None declared Salminen. Spine 1999 Cheung. Spine 2009 Boden. Journal of Joint and Bone surgery 1990 Savage. Spine Journal 1997 Mimura. Spine 1994 Welsch. Skeletal Radiology 2011 Mamisch. Skeletal Radiology 2011 Pfirrmann. Spine 2001. Funding: NIH/NIAMS T32 AR050938

Quantification of Intervertebral Disk Cartilaginous Endplate Morphology using MRI

Introduction The IVD cartilaginous endplate (CEP) is a 450 mm to 800 mm thick layer of hyaline cartilage,2 which functions as a mechanical barrier between the pressurized nucleus pulposus and the vertebral endplate, and as a gateway for nutrient diffusion to the avascular disk.1–3 With age and degeneration, the CEP becomes thinner, calcifies, and may have reduced porosity,4 and it has been implicated as a contributor to degeneration.1–3 As the CEP is not well imaged on MRI, key geometric parameters of the CEP have not been obtained noninvasively. Thus, the objective of this study was to measure geometric properties of the CEP with MRI. Materials and Methods Lumbar motion segments ( n = 24), from 11 human cadavers (age 64.9 ± 10.0 years) were imaged on a Siemens 7T MRI scanner using a custom RF coil. Disk degeneration was graded via mid-sagittal T2 mapping. Full volume imaging of each disk at 200 mm isotropic resolution was achieved via 3D FLASH (TR/TE = 9/3.7 ms) in 6 minutes (Fig. 1). Geometric parameters (area, circumference, anterior-posterior width, lateral width) were measured in the axial and mid-sagittal planes for both the CEP and the entire disk using OsiriX software. Disk height was measured using custom-written Matlab code.5 Evaluation of CEP thickness was done on a mid-sagittal image of each disc. Semiautomated image analysis was conducted using custom-written Matlab code in which signal intensity profiles across the CEP thickness were captured at the center of the CEP and at ± ¼ and ± ¾ of its A-P length. Comparisons between superior and inferior CEP geometry were established using paired, two-tailed t-tests ( p < 0.05). Data, except thickness, was normalized by dividing CEP geometry by its corresponding disk geometry to account for differences in disk sizes between levels. A one-way ANOVA was used to evaluate differences between disk level and CEP geometry. CEP thickness measurements were evaluated using a two-way ANOVA with repeated measures, where the factors were disk level and anterior-posterior disk location. A posthoc Bonferonni test was performed when significance was detected. Linear regression analysis was performed to compare CEP geometry with T2 mapping values and subject age. Results Directly measured whole disk and CEP geometry for each level are provided in the Table. The mean CEP thickness in the mid-sagittal plane across all disk levels and locations was 0.50 ± 0.20 mm. No significant differences were found in CEP thickness with respect to the superior or inferior endplate or disk level ( p > 0.2). There was no significant difference in CEP A-P width, lateral width, or circumference area with respect to superior/inferior location ( p > 0.7) or disk level ( p > 0.6) except between L1-L2 and L4-L5 ( p < 0.05) (Fig. 2A-C). L1-L2 A-P width, lateral width, and circumference area were larger than in L4-L5 by 14, 18, and 28%, respectively. CEP thickness varied significantly with respect to anterior-posterior location for every level ( p < 0.005) (Fig. 2D). The minimum thickness was at the center of the disk (superior = 0.39 ± 0.10 mm, inferior = 0.38 ± 0.11 mm). The thickness at the center was 19.9 and 42.4% less than that of the ± ¼ and ± ¾ A-P locations. In the linear regression analysis, no correlation was found between CEP geometries and T2 mapping or age. Conclusion This study noninvasively visualized the CEP morphology and quantified CEP circumference, area, A-P width, lateral width, and thickness using an MRI 3D FLASH sequence. The measured disk geometries in this study agree well with previous literature.5,6 CEP geometries were not linearly correlated with age or T2 mapping, which is correlated with disk degenerative grade. The lack of correlation between CEP geometries and either age or T2 mapping as a marker of degeneration was not completely expected, since it has been discussed that CEP thickness decreases with degeneration.1–3 Additional data and increased sample size may provide more insight into the potential role of the CEP in the degenerative cascade. Acknowledgment NIH grant R01 AR050052. I confirm having declared any potential conflict of interest for all authors listed on this abstract Yes Disclosure of Interest None declared Urban JPG. Arthritis Research Therapy 2003;5:120 Roberts S. Spine 1989;14:166 Moore RJ. European Spine Journal 2000;9:92 Bernick S, Spine 1982;7:97 O'Connell GD. Spine 2007:32:328 Elliott DM. Spine 2004;29:713

Influence of Autologous Mononuclear Stem-Cells over Symptomatic Disk Disease in Experimental Animal Model

IntroductionVarious therapeutic strategies have been used for controlling or at least slowing the degenerative cascade that leads to symptomatic disk degeneration. In this context, given its regenerative capability, stem cells (SC) therapy has shown promising results acting early on the degenerative cascade. This study aims to evaluate the influence of autologous adult mononuclear SC on the histological changes of experimentally induced disk degeneration. Materials and MethodsAfter local Ethical Committee approval, 16 rabbits-New Zealand, male, 3.5 to 4.5 kg-underwent needle puncture of the three caudal lumbar intervertebral discs using a previously validated technique, and the remaining discs were used as controls. The animals received either autologous mesenchymal SC or saline solution after 2 weeks (group 1) or 2 months (group 2). Animals underwent euthanasia 8 weeks after the cell administration, and the discs were harvested for analysis.The slides were analyzed on hematoxylin-eosin stains for the presence of degeneration by a previously validated score, and by automated color morphometry, also previously validated by the authors, defining cellular size and cellular population within the nucleus pulposus (NP). ResultsTotal 91 intervertebral discs (IVDs) were obtained for histological analysis, extracted from 16 animals-4 animals were excluded due to inadequate tissue preparation. The study group was divided as follows: 55 control-not punctured-IVDs and 36 experimental IVDs; were 21 have received SC and 15, isotonic saline solution.The microscopically characteristic alterations of the experimental disk degeneration have been observed in all punctured IVDs; however, some slides suggested a less intensive degeneration process and normal areas inside de nucleus pulposus, meaning recovery areas in the IVDs which have received mononuclear stem cells.Through color morphometry, significant differences favoring the SC group regarding number of cells and especially nuclear size (p = 0.03) give indirect evidence that the injected cells survived within the NP. There was also a trend favoring the group receiving SC earlier (2 weeks). ConclusionInjection of SC was not able to interrupt the degenerative process triggered by disk puncture. However, the histological changes were less severe in the groups receiving cell therapy. Color morphometry showed clusters of cells within the NP, not seen in the control groups.I confirm having declared any potential conflict of interest for all authors listed on this abstractYesDisclosure of InterestNone declared

Spine Osteoarthritis

Osteoarthritis of the spine develops as a consequence of the natural aging process and is associated with significant morbidity and health care expenditures. Effective diagnosis and treatment of the resultant pathologic conditions can be clinically challenging. Recent evidence has emerged to aid the investigating clinician in formulating an accurate diagnosis and in implementing a successful treatment algorithm. This article details the degenerative cascade that results in the osteoarthritic spine, reviews prevalence data for common painful spinal disorders, and discusses evidence-based treatment options for management of zygapophysial and sacroiliac joint arthrosis.

Axonal degeneration as a therapeutic target in the CNS

Degeneration of the axon is an important step in the pathomechanism of traumatic, inflammatory and degenerative neurological diseases. Increasing evidence suggests that axonal degeneration occurs early in the course of these diseases and therefore represents a promising target for future therapeutic strategies. We review the evidence for axonal destruction from pathological findings and animal models with particular emphasis on neurodegenerative and neurotraumatic disorders. We discuss the basic morphological and temporal modalities of axonal degeneration (acute, chronic and focal axonal degeneration and Wallerian degeneration). Based on the mechanistic concepts, we then delineate in detail the major molecular mechanisms that underlie the degenerative cascade, such as calcium influx, axonal transport, protein aggregation and autophagy. We finally concentrate on putative therapeutic targets based on the mechanistic prerequisites.

The Biomechanics of Cervical Spondylosis

Aging is the major risk factor that contributes to the onset of cervical spondylosis. Several acute and chronic symptoms can occur that start with neck pain and may progress into cervical radiculopathy. Eventually, the degenerative cascade causes desiccation of the intervertebral disc resulting in height loss along the ventral margin of the cervical spine. This causes ventral angulation and eventual loss of lordosis, with compression of the neural and vascular structures. The altered posture of the cervical spine will progress into kyphosis and continue if the load balance and lordosis is not restored. The content of this paper will address the physiological and biomechanical pathways leading to cervical spondylosis and the biomechanical principles related to the surgical correction and treatment of kyphotic progression.

Intervertebral disc viscoelastic parameters and residual mechanics spatially quantified using a hybrid confined/in situ indentation method

With advancing age, injury, musculoskeletal pathology or a combination of these, a degenerative cascade of biomechanical, biochemical, and nutritional alterations diminish the intervertebral discs’ ability to maintain its structure and function. While the biomechanics of isolated disc tissues has been investigated across this degenerative spectrum, none have attempted to retain the in situ disc-endplate morphology during compressive tissue characterization. The objective of this study was to spatially quantify the viscoelastic parameters of the intervertebral disc throughout degeneration, including the as yet unreported residual stress/strain. This required the development of a hybrid confined/in situ indentation methodology, which preserves the disc structural morphology. At four locations of the disc (anterior-AF, right and left lateral AF, and NP) stress-relaxation tests were performed using the hybrid confined/in situ indentation method, which utilizes the vertebral endplate as the porous indenter tip. This method allows the endplate to remain interwoven with the disc tissue, retaining its native orientation. Healthy disc tissue exhibited significantly higher residual stress values compared to both moderate and severe degeneration in all locations (p<0.0156). Furthermore, the equilibrium stress at 15% strain (stress relaxation) was significantly diminished with advancing disc degeneration (p<0.0241). The equilibrium viscoelastic parameters show healthy discs encounter higher forces at the same strain level, and are able to maintain this force, where degenerated discs are unable to maintain this force throughout time. This morphology-conserved method provides insight into the spatial compressive mechanical properties of the intervertebral disc across the degeneration spectrum and will aid in modeling these tissue changes.

Conjoint pathologic cascades mediated by ALS/FTLD-U linked RNA-binding proteins TDP-43 and FUS

The RNA-binding proteins TAR DNA-binding protein (TDP-43) and fused in sarcoma (FUS) play central roles in neurodegeneration associated with familial amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration with ubiquitin-positive inclusions (FTLD-U). Normally localized in the nucleus, in sites affected by ALS and FTLD-U they are mislocalized to the cytoplasm and form cytoplasmic inclusions. TDP-43 and FUS are transported to the nucleus in a Ran-GTPase-dependent manner via nuclear import receptors, but they also contribute to the formation of stress granules (SGs), which are intracytoplasmic structures incorporating RNA. C-terminal truncations of TDP-43 eliminate the nuclear transport signal and cause mislocalization of the protein to the cytoplasm, where it accumulates and forms SGs. ALS-associated FUS mutations impair nuclear transport and cause mislocalization of FUS to the cytoplasm, where it also contributes to assembly of SGs. Furthermore, the ALS susceptibility factor ataxin-2, recently identified as a potent modifier of TDP-43 toxicity, is also a predicted cytoplasmic RNA-binding protein and a constituent protein of SGs, suggesting that it is a part of the common pathologic cascade formed by TDP-43 and FUS. Thus, we propose that excessive mislocalization of the RNA-binding proteins TDP-43, FUS, and ataxin-2 into the cytoplasm leads to impairment of the RNA quality control system, forming the core of the ALS/FTLD-U degenerative cascade. In this review, we discuss the molecular basis of the novel disease spectrum of ALS/FTLD-U, including the neurodegenerative mechanism of the cytoplasmic RNA-binding proteins TDP-43 and FUS and the possibility of a novel therapeutic strategy.

Cervical spondylotic myelopathy: pathophysiology, clinical presentation, and treatment.

Age-related changes in the spinal column result in a degenerative cascade known as spondylosis. Genetic, environmental, and occupational influences may play a role. These spondylotic changes may result in direct compressive and ischemic dysfunction of the spinal cord known as cervical spondylotic myelopathy (CSM). Both static and dynamic factors contribute to the pathogenesis. CSM may present as subclinical stenosis or may follow a more pernicious and progressive course. Most reports of the natural history of CSM involve periods of quiescent disease with intermittent episodes of neurologic decline. If conservative treatment is chosen for mild CSM, close clinical and radiographic follow-up should be undertaken in addition to precautions for trauma-related neurologic sequelae. Operative treatment remains the standard of care for moderate to severe CSM and is most effective in preventing the progression of disease. Anterior surgery is often beneficial in patients with stenotic disease limited to a few segments or in cases in which correction of a kyphotic deformity is desired. Posterior procedures allow decompression of multiple segments simultaneously provided that adequate posterior drift of the cord is attainable from areas of anterior compression. Distinct risks exist with both anterior and posterior surgery and should be considered in clinical decision-making.

July 2011 Letters to the Editor-in-Chief

July 2011 Letters to the Editor-in-Chief

Complex loading affects intervertebral disc mechanics and biology

Complex loading develops in multiple spinal motions and in the case of hyperflexion is known to cause intervertebral disc (IVD) injury. Few studies have examined the interacting biologic and structural alterations associated with potentially injurious complex loading, which may be an important contributor to chronic progressive degeneration. This study tested the hypothesis that low magnitudes of axial compression loading applied asymmetrically can induce IVD injury affecting cellular and structural responses in a large animal IVD ex-vivo model. Bovine caudal IVDs were assigned to either a control or wedge group (15°) and placed in organ culture for 7 days under static 0.2MPa load. IVD tissue and cellular responses were assessed through confined compression, qRT-PCR, histology and structural and compositional measurements, including Western blot for aggrecan degradation products. Complex loading via asymmetric compression induced cell death, an increase in caspase-3 staining (apoptosis), a loss of aggrecan and an increase in aggregate modulus in the concave annulus fibrosis. While an up-regulation of MMP-1, ADAMTS4, IL-1β, and IL-6 mRNA, and a reduced aggregate modulus were induced in the convex annulus. Asymmetric compression had direct deleterious effects on both tissue and cells, suggesting an injurious loading regime that could lead to a degenerative cascade, including cell death, the production of inflammatory mediators, and a shift towards catabolism. This explant model is useful to assess how injurious mechanical loading affects the cellular response which may contribute to the progression of degenerative changes in large animal IVDs, and results suggest that interventions should address inflammation, apoptosis, and lamellar integrity.

Selective modulation of the endocannabinoid system for targeted protection in kainic acid models of excitotoxicity

The endocannabinoids anandamide (AEA) and 2‐arachidonoylglycerol (2‐AG) have been implicated in neuroprotective responses against excitotoxic events linked to seizure activity and associated neurodegeneration. Related to such responses to injury, excitotoxic kainic acid (KA) injections increase AEA levels in the brain. To study the modulation of this response in vitro and in vivo, we utilized new‐generation compounds (AM5206 and AM6702) that selectively inhibit the AEA deactivating enzyme fatty acid amide hydrolase (FAAH), as well as a dual inhibitor (AM6701) that blocks FAAH and the 2‐AG deactivating enzyme monoacylglycerol lipase (MAGL). AM5206 protected against the KA‐induced degenerative cascade in hippocampal slices assessed 24 h post‐insult. Cultured slice data also suggest AM6701 elicits more protection than AM6702 by ameliorating cytoskeletal breakdown and declines in synaptic markers and neuronal integrity. In vivo, KA administration induced seizures and the same neurodegenerative events exhibited in vitro. Protection by AM5206 and AM6701 was evident with respect to cytoskeletal damage, pre‐ and postsynaptic markers, seizure scores, and behavioral deficits. Our study has identified and characterized new‐generation inhibitors of endocannabinoid‐degrading enzymes to enhance cannabinergic signaling, and in turn prevent excitotoxic progression that cause brain damage.

The effect of needle size inducing degeneration in the rat caudal disc: evaluation using radiograph, magnetic resonance imaging, histology, and immunohistochemistry

Background context The rat caudal disc has been increasingly used in studying of disc degeneration because of its simplicity, low cost, and efficiency. However, the reproducibility and standardization are essential to facilitate the investigations of biologic therapeutics at different stages of degeneration. Purpose To identify the effect of different needle gauges to the degenerative response in rat caudal discs and to examine its pathogenesis by looking at the cellular and matrix changes. Study design In vivo study of injury-induced rat caudal disc degeneration using needle puncture. Patient sample Thirty-six Lewis rats aged 12–14 weeks. Outcome measures The induced degenerative discs were analyzed by plain radiograph, magnetic resonance imaging (MRI) and histological examination. Proteoglycan content was assessed by alcian blue stain. Immunohistochemistry using aggrecan, collagen II, and Sox-9 was also evaluated to investigate cell differentiation and matrix changes. Methods All rats were divided into three groups according to different needle gauges (18G, 20G, and 22G). Caudal discs were punctured percutaneously under image guidance. Radiographs and MRI were obtained at 2 weeks interval until 8 weeks. At each time point, three rats from each group were sacrificed for histological analysis and immunohistochemistry. Results Larger needle gauges, especially 18G, produced more deterioration of the disc when compared with smaller sizes, particularly with time. Significant differences were identified in almost all parameters compared between 18G and 22G at the 8-week time point. For the effect of time in the same needle size, the differences occurred between 2- or 4-week and 8-week time point in the 18G and 20G groups. The proteoglycan and aggrecan stain gradually decreased over time. Chondrogenic differentiation was identified within the degenerative disc by detecting Sox-9 positive cells and collagen II accumulation increased as degeneration progressed. Conclusions The puncture-induced degenerative changes in rat caudal discs can imitate the human degenerative cascade as observed in plain radiograph, MRI, histology, and immunohistochemistry. We suggest that needle size affects the occurrence of progression of degeneration; thus, the large needle size was required to accelerate the deterioration. The size of needle and time point after injury should be considered when investigating the effect of therapeutic materials to retard degeneration or regenerate the intervertebral disc.

Coordinated and sequential activation of neutral and acidic DNases during interdigital cell death in the embryonic limb

Interdigital tissue regression during embryonic development is one of the most representative model systems of morphogenetic cell death, but the degenerative cascade accounting for this process awaits clarification. Although the canonical apoptotic caspase pathway appears to be activated in the interdigital mesenchyme committed to die, neither genetic nor chemical blockage of caspases or their downstream effectors, is sufficient to prevent cell death. Hence, alternative and/or complementary dying pathways must also be responsible for this degenerative process. In this work we have chosen to study the endonucleases during the regression of the interdigital tissue of avian embryos to gain insights into the molecular mechanisms accounting for programmed cell death in this system. We show that caspase activated DNase, which is a neutral DNase associated with the caspase apoptotic pathway, appears to be the main endonuclease only at an initial phase of interdigit regression. However at peak stages of the degenerative process, the acidic DNases L-DNase II and lysosomal DNase IIB become predominant in the system and markers for cell autophagy become moderately up-regulated. Consistent with the activation of acidic endonucleases we observed that microenvironmental pH value in the interdigits decreased to levels only appropriate for acidic enzymes. Furthermore, we found that overexpression of lysosomal DNase IIB in embryonic limb mesoderm promoted cell death, which was also accompanied by up-regulation and activation of L-DNase II. Up-regulation of acidic DNases was maintained in interdigits explanted to culture dishes, where the participation of exogenous professional phagocytes of hematopoietic origin is avoided. Finally, and consistent with all our findings, up-regulation of acidic DNases was much reduced in the webbed interdigits of duck embryos, characterized by a rudimentary interdigital degenerative process. We conclude that the regression of the interdigital tissue involves a coordinated and sequential activation of the caspase and lysosomal degenerative molecular cascades.

Validated Finite Element Analysis of the Maverick Total Disc Prosthesis

Combining in vitro tests and finite element analysis to provide a more complete picture of the role that a disc prosthesis implant would play in the biomechanics of the spine. Analysis of the disc function after total disc prosthesis insertion with and without antero-posterior or lateral offset and in combination with adjacent fusion. To avoid the risk of degenerative cascade the total disc replacement may be considered as an alternative. Few finite element analysis combined with cadaver testing under loading conditions have been published today. In vitro tests were performed using 6 fresh human cadaver specimens to quantify the load-displacement behaviors before and after insertion of a total disc replacement (Maverick, Memphis) implant. A finite element (FE) spine model was validated with the data from the in vitro tests. This model is built on the basis of ANSYS software. The effect of the prosthesis positioning on the motion behavior at L4-L5 and on the inner loads over facets was evaluated in 4 configurations. The study showed that the motion behavior at the levels adjacent to the Maverick prosthesis remained the same as the intact spine, unlike a single level fusion at L5-S1. In the biomechanical study settings, Maverick prosthesis, once properly positioned, does not modify the motion behavior of the spine as compared with its intact state. The less-than-ideal positioning of the prosthesis, especially with anterior offset, affect significantly the range of motion of the spine segment and cause increase of inner load in the facets. Those results indicated a good reliability of the finite element model in representing both intact and instrumented spine segments. The in vitro test results demonstrated that Maverick disc prosthesis provides near physiologic function of a natural disc restores stability of the spine and preserves the segmental motion without undue stress on adjacent segments.To our knowledge, this study suggested for the first time the importance of the prosthesis positioning into the spine model.

Langzeitergebnisse, Studienlage und Differenzialindikation des DIAM™-Implantats

The DIAM (Device for Interspinous Assisted Motion) interspinous device offers the possibility of a viscoelastic supplementation of the lumbar motion segment with a small, reversible operation. It is intended as a restabilisation procedure after a discectomy to avoid facet joint overload and as a topping of procedure for the segment adjacent to a fusion. The device has been on the German market since 2004. There are several biomechanical studies available showing an effect mainly in extension and flexion, but hardly any in rotation and lateral inclination. Despite frequent clinical use, there have only been a few, mainly retrospective clinical studies indicating the success of the implant; however, sound scientific data are missing. Several prospective, randomised, controlled studies are now underway to fill that gap. Only then will it be possible to assess whether this implant is of true value to improve clinical results and to slow down the degenerative cascade or not.