No description available.

Objectif: Nous avons effectué une étude chez 135 patients ayant subis une chirurgie lombo-sacrée avec vissage pédiculaire sous navigation par tomographie axiale. Nous avons évalué la précision des vis pédiculaires et les résultats cliniques. Méthodes: Cette étude comporte 44 hommes et 91 femmes (âge moyen=61, intervalle 24-90 ans). Les diamètres, longueurs et trajectoires des 836 vis ont été planifiés en préopératoire avec un système de navigation (SNN, Surgical Navigation Network, Mississauga). Les patients ont subi une fusion lombaire (55), lombo-sacrée (73) et thoraco-lombo-sacrée (7). La perforation pédiculaire, la longueur des vis et les spondylolisthesis sont évalués par tomographies axiales postopératoires. Le niveau de douleur est mesuré par autoévaluations, échelles visuelles analogues et questionnaires (Oswestry et SF-36). La fusion osseuse a été évaluée par l’examen des radiographies postopératoires. Résultats: Une perforation des pédicules est présente pour 49/836 (5.9%) des vis (2.4% latéral, 1.7% inférieur, 1.1% supérieur, 0.7% médial). Les erreurs ont été mineures (0.1-2mm, 46/49) ou intermédiaires (2.1 - 4mm, 3/49 en latéral). Il y a aucune erreur majeure (≥ 4.1mm). Certaines vis ont été jugées trop longues (66/836, 8%). Le temps moyen pour insérer une vis en navigation a été de 19.1 minutes de l΄application au retrait du cadre de référence. Un an postopératoire on note une amélioration de la douleur des jambes et lombaire de 72% et 48% en moyenne respectivement. L’amélioration reste stable après 2 ans. La dégénérescence radiologique au dessus et sous la fusion a été retrouvée chez 44 patients (33%) and 3 patients respectivement (2%). Elle est survenue en moyenne 22.2 ± 2.6 mois après la chirurgie. Les fusions se terminant à L2 ont été associées à plus de dégénération (14/25, 56%). Conclusion: La navigation spinale basée sur des images tomographiques préopératoires est une technique sécuritaire et précise. Elle donne de bons résultats à court terme justifiant l’investissement de temps chirurgical. La dégénérescence segmentaire peut avoir un impact négatif sur les résultats radiologique et cliniques. / Objective: The authors studied 135 consecutive patients following a lumbo-sacral fixation using pedicle screws and CT-based navigation to evaluate pedicle screw accuracy and clinical outcomes. Methods: The series included 44 men and 91 women (mean age 61 years, range 24-90 years). All 836 screws were planned with pre-operative CT-Scans in a navigation system (SNN, Surgical Navigation Network, Mississauga, Ontario, Canada) for diameter, length and direction. Fixation included the lumbar spines only (55), the lumbo-sacral spine (73) or the thoraco-lumbo-sacral spine (7). Pedicle perforation, screw length and spondylolisthesis were assessed on post-operative CT-Scan. Pain was surveyed using self-rated scales, visual analogue scales, Oswestry and SF-36 questionnaires. Bony union was assessed on post-operative follow-up radiographs. Results: Pedicle perforation was found in 49/836 (5.9%) screws (2.4% laterally, 1.7% inferiorly, 1.1% superiorly, 0.7% medially). The errors were minor (0.1-2mm, 46/49) or intermediate (2.1 – 4 mm, 3/49). All intermediate errors were lateral. There were no major errors (≥ 4.1mm). Some screws were judged too long (66/836, 8%). The average time to insert one screw with navigation was 19.1 minutes from application to removal of the reference frame. The amount of improvement at one year post-operation for self-rated leg and back pain were 72% and 48% respectively. The improvement was stable over 2 years. Above-level and below-level radiological degenerations were found in 44 patients (33%) and 3 patients respectively (2%) and occurred on average 22.2 ± 2.6 months after the surgery. Fusions ending at L2 had the most degenerations (14/25, 56%). Conclusion: CT-based preoperative navigation for lumbo-sacral pedicle screw insertion is accurate and associated with a good short term outcome, making it worth the investment of the additional time required. Segmental degeneration may have a negative effect on radiological and clinical outcomes.

Indiana University-Purdue University Indianapolis (IUPUI). / Calcium/calmodulin-dependent protein kinase II (CaMKII) is a critical modulator of synaptic function, plasticity, and learning and memory. In neurons and astrocytes, CaMKII regulates cellular excitability, cytoskeletal structure, and cell metabolism. A rapid increase in CaMKII activity is observed within the first few minutes of ischemic stroke in vivo; this calcium-dependent process is also observed following glutamate stimulation in vitro. Activation of CaMKII during pathological conditions is immediately followed by inactivation and aggregation of the kinase. The extent of CaMKII inactivation is directly correlated with the extent of neuronal damage. The studies presented here show that these fluctuations in CaMKII activity are not correlated with neuronal death; rather, they play a causal role in neuronal death. Pharmacological inhibition of CaMKII in the time immediately surrounding glutamate insult protects cultured cortical neurons from excitotoxicity. Interestingly, pharmacological inhibition of CaMKII during excitotoxic insult also prevents the aggregation and prolonged inactivation of the kinase, suggesting that CaMKII activity during excitotoxic glutamate signaling is detrimental to neuronal viability because it leads to a prolonged loss of CaMKII activity, culminating in neuronal death. In support of this, CaMKII inhibition in the absence of excitotoxic insult induces cortical neuron apoptosis by dysregulating intracellular calcium homeostasis and increasing excitatory glutamate signaling. Blockade of the NMDA-receptors and enzymatic degradation of the extracellular glutamate signal affords neuroprotection from CaMKII inhibition-induced toxicity. Co-cultures of neurons and glutamate-buffering astrocytes also exhibit this slow-induced excitotoxicity, as CaMKII inhibitors reduce glutamate uptake within the astrocytes. CaMKII inhibition also dysregulates calcium homeostasis in astrocytes and leads to increased ATP release, which was neurotoxic when applied to naïve cortical neurons. Together, these findings indicate that during aberrant calcium signaling, the activation of CaMKII is toxic because it supports aggregation and prolonged inactivation of the kinase. Without CaMKII activity, neurons and astrocytes release stores of transmitters that further exacerbate neuronal toxicity.

<p>  </p> <p>With the discovery and treatment of any disease comes the important question of its genetic prevalence. This is especially important for animals under strict breeding control, such as dogs, because this can provide essential information regarding breeding pair decisions. Thus, the focus of this thesis is to investigate the genetic prevalence of three different diseases: 1) Factor VII Deficiency (FVIID), 2) Collie Eye Anomaly (CEA), and 3) Progressive Rod-Cone Degeneration-Progressive Retinal Atrophy (prcd-PRA). Factor VII Deficiency (FVIID) is a clotting disorder observed in both humans and dogs, characterized by impeded function of the Factor VII protein. In dogs, FVIID is caused by a single nucleotide substitution (c.407G>A) in the <em>F7 </em>gene. This mutation, identified in a colony of research Beagles, is also present in dogs with a wide variety of distantly-related breed backgrounds and in mixed-breed dogs, suggesting an ancient, ancestral origin. Given the relatively common presence of this variant, it was hypothesized that this genetic mutation could be contributing to excessive bleeding in canine autopsy cases that could not be attributed to typical causes. DNA from formalin-fixed paraffin-embedded tissues (n = 67 cases) were Sanger sequenced for the FVIID c.407G>A mutation, and all were determined to be homozygous wild-type. Therefore, the tested variant is not associated with the unexplained bleeding in these cases, and it is not a logical diagnostic test to apply to similar cases in the future.</p> <p><br></p> <p>CEA and prcd-PRA are ophthalmic genetic diseases of concern often included in commercial genetic testing panels. A large dataset spanning 15+ years provided by a commercial partner company (OptiGen/Wisdom Panel, Kinship) encompassed dogs tested for the CEA-associated <em>NHEJ1</em> deletion (n = 33,834 dogs) and the prcd-PRA causal mutation in <em>PRCD</em> (n = 86,667 dogs). Disease trends were observed graphically and analyzed with Chi-square goodness-of-fit testing and regression modeling for disease status and genotype classification. Both diseases had a statistically significant change in genotype frequencies from the first year of data to the last; both diseases also had a negative association between progression of time and overall probability of a dog being disease-positive or a carrier/heterozygous. This suggests that genetic testing results are being incorporated into breeding decisions, although affected dogs were still being identified by the end of this study. Different breeds, AKC groups, FCI groups, genetic clades, and geography were also investigated to determine impact on overall disease trend. </p>

No description available.

No / Microglial activation and behavioral abnormalities occur before neuronal loss in experimental murine prion disease; the behavioral changes coincide with a reduction in synaptic plasticity. Because synaptic plasticity depends on an intact perineuronal net (PN), a specialized extracellular matrix that surrounds parvalbumin (PV)-positive GABAergic (gamma-aminobutyric acid [GABA]) inhibitory interneurons, we investigated the temporal relationships between microglial activation and loss of PN and PV-positive neurons in ME7 murine prion disease. Anesthetized C57Bl/6J mice received bilateral intracerebral microinjections of ME7-infected or normal brain homogenate into the dorsal hippocampus. Microglial activation, PrP accumulation, the number of PV-positive interneurons, and Wisteria floribunda agglutinin-positive neurons (i.e. those with an intact PN) were assessed in the ventral CA1 and subiculum at 4, 8, 12, 16, and 20 weeks postinjection. Hippocampal areas and total neuron numbers in the ventral CA1 and subiculum were also determined. Loss of PN coincided with early microglial activation and with a reduction in synaptic plasticity. No significant loss of PV-positive interneurons was observed. Our findings suggest that the substrate of the earliest synaptic and behavioral abnormalities in murine prion disease may be inflammatory microglia-mediated degradation of the PN.

No / Retinal dystrophies are an overlapping group of genetically heterogeneous conditions resulting from mutations in more than 250 genes. Here we describe five families affected by an adult-onset retinal dystrophy with early macular involvement and associated central visual loss in the third or fourth decade of life. Affected individuals were found to harbor disease-causing variants in DRAM2 (DNA-damage regulated autophagy modulator protein 2). Homozygosity mapping and exome sequencing in a large, consanguineous British family of Pakistani origin revealed a homozygous frameshift variant (c.140delG [p.Gly47Valfs∗3]) in nine affected family members. Sanger sequencing of DRAM2 in 322 unrelated probands with retinal dystrophy revealed one European subject with compound heterozygous DRAM2 changes (c.494G>A [p.Trp165∗] and c.131G>A [p.Ser44Asn]). Inspection of previously generated exome sequencing data in unsolved retinal dystrophy cases identified a homozygous variant in an individual of Indian origin (c.64_66del [p.Ala22del]). Independently, a gene-based case-control association study was conducted via an exome sequencing dataset of 18 phenotypically similar case subjects and 1,917 control subjects. Using a recessive model and a binomial test for rare, presumed biallelic, variants, we found DRAM2 to be the most statistically enriched gene; one subject was a homozygote (c.362A>T [p.His121Leu]) and another a compound heterozygote (c.79T>C [p.Tyr27His] and c.217_225del [p.Val73_Tyr75del]). DRAM2 encodes a transmembrane lysosomal protein thought to play a role in the initiation of autophagy. Immunohistochemical analysis showed DRAM2 localization to photoreceptor inner segments and to the apical surface of retinal pigment epithelial cells where it might be involved in the process of photoreceptor renewal and recycling to preserve visual function.

Indiana University-Purdue University Indianapolis (IUPUI) / Human induced pluripotent stem (iPS) cells have the unique ability to differentiate into 200 or so somatic cell types that make up the adult human being. The use of human iPS cells to study development and disease is a highly exciting and interdependent field that holds great promise in understanding and elucidating mechanisms behind cellular differentiation with future applications in drug screening and cell replacement studies for complex and currently incurable cellular degenerative disorders. The recent advent of iPS cell technology allows for the generation of patient-specific cell lines that enable us to model the progression of a disease phenotype in a human in vitro model. Differentiation of iPS cells toward the affected cell type provides an unlimited source of diseased cells for examination, and to further study the developmental progression of the disease in vitro, also called the “disease-in-a-dish” model. In this study, efforts were undertaken to recapitulate the differentiation of distinct retinal cell affected in two highly prevalent retinal diseases, Usher syndrome and glaucoma. Using a line of Type III Usher Syndrome patient derived iPS cells efforts were undertaken to develop such an approach as an effective in vitro model for studies of Usher Syndrome, the most commonly inherited disorder affecting both vision and hearing. Using existing lines of iPS cells, studies were also aimed at differentiation and characterization of the more complex retinal cell types, retinal ganglion cells (RGCs) and astrocytes, the cell types affected in glaucoma, a severe neurodegenerative disease of the retina leading to eventual irreversible blindness. Using a previously described protocol, the iPS cells were directed to differentiate toward a retinal fate through a step-wise process that proceeds through all of the major stages of neuroretinal development. The differentiation process was monitored for a period of 70 days for the differentiation of retinal cell types and 150 days for astrocyte development. The different stages of differentiation and the individually derived somatic cell types were characterized by the expression of developmentally associated transcription factors specific to each cell type. Further approaches were undertaken to characterize the morphological differences between RGCs and other neuroretinal cell types derived in the process. The results of this study successfully demonstrated that Usher syndrome patient derived iPS cells differentiated to the affected photoreceptors of Usher syndrome along with other mature retinal cell types, chronologically analogous to the development of the cell types in a mature human retina. This study also established a robust method for the in vitro derivation of RGCs and astrocytes from human iPS cells and provided novel methodologies and evidence to characterize these individual somatic cell types. Overall, this study provides a unique insight into the application of human pluripotent stem cell biology by establishing a novel platform for future studies of in vitro disease modeling of the retinal degenerative diseases: Usher syndrome and glaucoma. In downstream applications of this study, the disease relevant cell types derived from human iPS cells can be used as tools to further study disease progression, drug screening and cell replacement strategies.

Traumatic brain injury (TBI) is a leading cause of disability worldwide. Annually, 150 to 200/1,000,000 people become disabled as a result of brain trauma. Axonal degeneration is a critical, early event following TBI of all severities but whether axon degeneration is a driver of TBI remains unclear. Molecular pathways underlying the pathology of TBI have not been defined and there is no efficacious treatment for TBI. Despite this significant societal impact, surprisingly little is known about the molecular mechanisms that actively drive axon degeneration in any context and particularly following TBI. Although severe brain injury may cause immediate disruption of axons (primary axotomy), it is now recognized that the most frequent form of traumatic axonal injury (TAI) is mediated by a cascade of events that ultimately result in secondary axonal disconnection (secondary axotomy) within hours to days. Proposed mechanisms include immediate post-traumatic cytoskeletal destabilization as a direct result of mechanical breakage of microtubules, as well as catastrophic local calcium dysregulation resulting in microtubule depolymerization, impaired axonal transport, unmitigated accumulation of cargoes, local axonal swelling, and finally disconnection. The portion of the axon that is distal to the axotomy site remains initially morphologically intact. However, it undergoes sudden rapid fragmentation along its full distal length ~72 h after the original axotomy, a process termed Wallerian degeneration. Remarkably, mice mutant for the Wallerian degeneration slow (Wlds) protein exhibit ~tenfold (for 2–3 weeks) suppressed Wallerian degeneration. Yet, pharmacological replication of the Wlds mechanism has proven difficult. Further, no one has studied whether Wlds protects from TAI. Lastly, owing to Wlds presumed gain-of-function and its absence in wild-type animals, direct evidence in support of a putative endogenous axon death signaling pathway is lacking, which is critical to identify original treatment targets and the development of viable therapeutic approaches. Novel insight into the pathophysiology of Wallerian degeneration was gained by the discovery that mutant Drosophila flies lacking dSarm (sterile a/Armadillo/Toll-Interleukin receptor homology domain protein) cell-autonomously recapitulated the Wlds phenotype. The pro-degenerative function of the dSarm gene (and its mouse homolog Sarm1) is widespread in mammals as shown by in vitro protection of superior cervical ganglion, dorsal root ganglion, and cortical neuron axons, as well as remarkable in-vivo long-term survival (>2 weeks) of transected sciatic mouse Sarm1 null axons. Although the molecular mechanism of function remains to be clarified, its discovery provides direct evidence that Sarm1 is the first endogenous gene required for Wallerian degeneration, driving a highly conserved genetic axon death program. The central goals of this thesis were to determine (1) whether post-traumatic axonal integrity is preserved in mice lacking Sarm1, and (2) whether loss of Sarm1 is associated with improved functional outcome after TBI. I show that mice lacking the mouse Toll receptor adaptor Sarm1 gene demonstrate multiple improved TBI-associated phenotypes after injury in a closed-head mild TBI model. Sarm1-/- mice developed fewer beta amyloid precursor protein (βAPP) aggregates in axons of the corpus callosum after TBI as compared to Sarm1+/+ mice. Furthermore, mice lacking Sarm1 had reduced plasma concentrations of the phosphorylated axonal neurofilament subunit H, indicating that axonal integrity is maintained after TBI. Strikingly, whereas wild type mice exhibited a number of behavioral deficits after TBI, I observed a strong, early preservation of neurological function in Sarm1-/- animals. Finally, using in vivo proton magnetic resonance spectroscopy, I found tissue signatures consistent with substantially preserved neuronal energy metabolism in Sarm1-/- mice compared to controls immediately following TBI. My results indicate that the Sarm1-mediated prodegenerative pathway promotes pathogenesis in TBI and suggest that anti-Sarm1 therapeutics are a viable approach for preserving neurological function after TBI.

Der wichtigste exzitatorische Neurotransmitter Glutamat spielt eine grosse Rolle in der Gehirnentwicklung, wie neuronale Migration und Synaptogenese. Ob glutamaterge Stimulation für das Überleben entwickelnder Neuronen notwendig ist, war bislang jedoch unbekannt. Um zu untersuchen, ob eine Hemmung von Glutamatrezeptoren im unreifen Gehirn zu Neurodegeneration führt, wurden Ratten im Alter von 1 bis 31 Tagen für 24 Stunden mit dem N-Methyl-D-Aspartat-(NMDA) Glutamatrezeptorantagonisten Dizocilpin (MK801) behandelt. Die Dichte neuronaler Degeneration wurde mikroskopisch in Kupfer-Silber- und TUNEL- gefärbten Hirnschnittpräparaten ermittelt und Unterschiede mittels ANOVA analysiert (Signifikanzniveau p / The predominant excitatory neurotransmitter glutamate plays a major role in certain aspects of neural development. However, whether developing neurons depend on glutamate for survival remains unknown. To investigate if deprivation of glutamate stimulation in the immature mammalian brain causes neuronal cell death (apoptosis), rat pups aged 0 to 30 days were treated for 24 hours with dizocilpine maleate (MK801), an N-methyl-D-aspartate-(NMDA) glutamate receptor antagonist. Density of neural degeneration was evaluated by a stereological dissector method in cupric-silver and TUNEL-stained brain slices. Groups were compared by ANOVA and significance considered at p